@article {pmid41808567,
year = {2026},
author = {Ding, X and Liu, Y and Luo, L and Cai, Y and Wang, C and Jin, J and Chen, Y},
title = {Review Genomic Hotspot Mining and Characterization for Stable Expression of Therapeutic Protein in Chinese Hamster Ovary Cells.},
journal = {ACS synthetic biology},
volume = {15},
number = {3},
pages = {1241-1247},
doi = {10.1021/acssynbio.5c00776},
pmid = {41808567},
issn = {2161-5063},
mesh = {Animals ; CHO Cells ; Cricetulus ; *Recombinant Proteins/genetics/biosynthesis ; CRISPR-Cas Systems/genetics ; Chromosome Mapping ; Cricetinae ; Promoter Regions, Genetic ; Genomics/methods ; Lentivirus/genetics ; },
abstract = {The development of rCHO cell lines that stably express therapeutic proteins is crucial for pharmaceutical protein industrial production. In this study, a systematic method was established to identify genomic hotspots for exogenous protein expression in CHO cells and construct stable recombinant CHO cell strains. Four stable monoclonal cell lines (1b7, 1d2, 2d9, and 2f7) were obtained by using the lentiviral random integration reporter gene. Chromosome mapping analysis found four stable integration sites: chr1_0 (7,30,83,299-7,32,45,508 bp) in 1b7, chr1_0 (17,69,68,187-17,69,68,191 bp) in 1d2, chr3 (4,08,81,262-4,08,99,858 bp) in 2d9, and chr5 (1,69,77,575-1,70,61,744 bp) in 2f7. Based on these sites, we developed recombinant CHO cells capable of long-term stable expression of foreign proteins through the combined application of CRISPR/Cas9 technology and Bxb1 recombinase-mediated cassette exchange. Utilizing "promoter capture technology", all screened LP cell monoclonal lines can express exogenous proteins, with the entire construction process completed in just 2∼3 weeks.},
}

Animals
CHO Cells
Cricetulus
*Recombinant Proteins/genetics/biosynthesis
CRISPR-Cas Systems/genetics
Chromosome Mapping
Cricetinae
Promoter Regions, Genetic
Genomics/methods
Lentivirus/genetics

@article {pmid41832076,
year = {2026},
author = {Gundra, SR and Jiang, W and Aouida, M and Wang, Q and Kazlak, AM and Elbehery, AHA and Saleh, A and Masood, M and Ghouneimy, A and Mahfouz, M},
title = {Characterization and engineering of highly efficient Cas12j genome editors.},
journal = {Trends in biotechnology},
volume = {},
number = {},
pages = {},
doi = {10.1016/j.tibtech.2026.02.001},
pmid = {41832076},
issn = {1879-3096},
abstract = {The large size of widely used CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats and CRISPR-associated proteins) enzymes limits their delivery for therapeutic applications. Cas12j nucleases offer a hypercompact alternative but show modest editing efficiency. To overcome this limitation, we identified eight novel Cas12j orthologs from viral metagenomes, which in their native form exhibit low editing activity in mammalian cells. We therefore engineered T5 exonuclease-Cas12j fusions, resulting in substantially enhanced genome-editing activity across multiple mammalian cell types, reaching levels comparable to established compact CRISPR-Cas editors. Intriguingly, robust cellular editing occurred in the presence of a previously unrecognized trinucleotide sequence context within the target DNA. Furthermore, we developed Cas12j-based adenine base editors by coupling catalytically inactive Cas12j orthologs with adenine deaminase, enabling efficient A-to-G base conversion in mammalian cells. This study expands the CRISPR toolbox by establishing engineering principles that convert compact Cas12j nucleases into efficient and modular genome-editing platforms well suited for delivery-constrained therapeutic applications.},
}

@article {pmid41832226,
year = {2026},
author = {Lee, SY and Park, HH},
title = {AcrIIA7 hijacks tracrRNA to block CRISPR-Cas system.},
journal = {Nature communications},
volume = {},
number = {},
pages = {},
doi = {10.1038/s41467-026-70749-w},
pmid = {41832226},
issn = {2041-1723},
support = {RS-2025-02316334//National Research Foundation of Korea (NRF)/ ; },
abstract = {The CRISPR-Cas9 system provides adaptive immunity against invading genetic elements through a dual-RNA-guided DNA cleavage mechanism. This system relies on the precise assembly of a ribonucleoprotein (RNP) complex composed of the Cas9 endonuclease, a CRISPR-derived RNA (crRNA), and a trans-activating CRISPR RNA (tracrRNA). Around 100 anti-CRISPR proteins that inhibit CRISPR-Cas systems have been identified, and the mechanisms by which they act are increasingly being elucidated. However, the inhibitory mechanisms of many Acrs, including AcrIIA7, remain poorly understood. Here, we present the structure of AcrIIA7 and uncover a previously unrecognized mechanism by which it inhibits Cas9 function. Structural and biochemical analyses reveal that AcrIIA7 specifically binds to tracrRNA, preventing its association with crRNA and thereby blocking formation of the active Cas9 RNP complex. This tracrRNA hijacking mechanism represents a unique strategy for CRISPR inhibition, in which an anti-CRISPR protein targets an RNA scaffold essential for Cas9 activation rather than interacting directly with the Cas9 protein. Our findings provide the first structural insight into tracrRNA-targeted anti-CRISPR activity and highlight RNA-RNA interaction interfaces as vulnerable nodes in CRISPR-Cas immunity.},
}

@article {pmid41834690,
year = {2026},
author = {Marpaung, DSS and Yap Sinaga, AO and Damayanti, D and Utari, NWA and Harmiansyah, and Karangan, A and Kusmali, M},
title = {Integration of CRISPR/Cas12a and Toehold-Mediated Strand Displacement for Alternative Conventional miRNA Detection.},
journal = {Chembiochem : a European journal of chemical biology},
volume = {27},
number = {6},
pages = {e202500932},
doi = {10.1002/cbic.202500932},
pmid = {41834690},
issn = {1439-7633},
mesh = {*MicroRNAs/analysis/genetics/metabolism ; *CRISPR-Cas Systems ; Humans ; *Endodeoxyribonucleases/metabolism/genetics ; Bacterial Proteins ; CRISPR-Associated Proteins ; },
abstract = {MicroRNAs (miRNAs) are short, noncoding RNAs that regulate gene expression and serve as powerful biomarkers for cancer and other diseases. Conventional detection methods such as RT-qPCR, Northern blotting, microarrays, and next-generation sequencing provide robust analytical capabilities but remain limited by complexity, cost, and poor suitability for point-of-care diagnostics. CRISPR/Cas12a has emerged as a versatile nucleic acid detection platform with high specificity and sensitivity. However, its intrinsic preference for DNA substrates restricts direct application to miRNA sensing. Early CRISPR/Cas12a-based assays relied on enzymatic amplification, direct RNA-induced activation, or split-component designs, each offering proof-of-concept feasibility but facing trade-offs in sensitivity, workflow complexity, or robustness. Toehold-mediated strand displacement (TSD) provides a powerful alternative by converting miRNA inputs into DNA activators or crRNAs that efficiently trigger Cas12a. This integration enables enzyme-free amplification, programmable logic operations, and enhanced sensitivity, while reducing reliance on multienzyme cascades. This review critically evaluates conventional, enzymatic, direct, and split-based CRISPR/Cas12a strategies and emphasizes emerging TSD-assisted platforms as next-generation solutions for sensitive, specific, and portable miRNA detection.},
}

*MicroRNAs/analysis/genetics/metabolism
*CRISPR-Cas Systems
Humans
*Endodeoxyribonucleases/metabolism/genetics
Bacterial Proteins
CRISPR-Associated Proteins

@article {pmid41834849,
year = {2026},
author = {Tenea, GN},
title = {Comprehensive genomic and metabolomic profiling of Weissella confusa UTNCys2-2 highlights bioactive potential.},
journal = {Frontiers in microbiology},
volume = {17},
number = {},
pages = {1779198},
pmid = {41834849},
issn = {1664-302X},
abstract = {INTRODUCTION: The genus Weissella comprises a diverse group of lactic acid bacteria (LAB) widely distributed across plant- and food-associated ecosystems and recognized for their functional and technological versatility. Weissella confusa UTNCys2-2, a plant-derived strain isolated from Amazonian spiral ginger (Costus sp.), that produces exopolysaccharides (EPS) with documented antioxidant activity and promising probiotic properties.

METHODS: Whole-genome sequencing of UTNCys2-2 was performed to establish its taxonomic assignment, phylogenomic analysis, while genome mining was conducted to evaluate safety, metabolic potential, and biosynthetic capabilities. Carbohydrate-active enzymes (CAZymes), Kyoto Encyclopedia of Genes and Genomes (KEGG), and MetaCyc pathways were analyzed for functional insights. Moreover, the metabolite composition of the cell-free supernatant (CFS) was examined using liquid chromatography-tandem mass spectrometry (LC-MS/MS) combined with Sequential Windowed Acquisition of all Theoretical Fragment Ion Mass Spectra (SWATH-MS).

RESULTS: The genome consists of a 2.32 Mb circular chromosome (44.59% GC) encoding 2,194 proteins, 76 tRNAs, and 10 rRNAs, with no plasmids. Phylogenomic analyses assigned the strain to the W. confusa clade, clustering closely with the reference strain DSM 20196. UTNCys2-2 harbors a complete Type II-A CRISPR-Cas system, intact prophages, and mobile elements, while lacking virulence determinants and transferable antimicrobial resistance genes. Functional annotation revealed 118 CAZymes supporting EPS biosynthesis, polysaccharide utilization, and carbohydrate metabolism. KEGG and MetaCyc pathways highlighted glycogen and riboflavin biosynthesis, stress tolerance, and metabolic versatility. Genome mining identified a Type III polyketide synthase (T3PKS) gene cluster with low similarity to known pathways, suggesting potential for novel secondary metabolites. Pangenome analysis showed extensive strain-specific genes linked to carbohydrate metabolism and EPS production. Metabolomic profiling of the CFS detected alkaloids, bioactive peptides, functional carbohydrates, and phenolics, supporting antimicrobial, probiotic, and host-interactive activities.

CONCLUSION: W. confusa UTNCys2-2 represents a biosafe and metabolically versatile strain with strong genomic capacity for EPS production, potential for novel secondary metabolite biosynthesis, and diverse bioactive properties, supporting its applicability in food fermentation, probiotic development, and microbial biotechnology.},
}

@article {pmid41834871,
year = {2026},
author = {Abdul Rehman, Y and Fayyaz, A and Alblooshi, AS and Muhammad, K and Mundra, S and Alam, MT},
title = {Molecular adaptations and engineering of extremophiles for synthetic biology and biotechnological applications.},
journal = {Frontiers in microbiology},
volume = {17},
number = {},
pages = {1754802},
pmid = {41834871},
issn = {1664-302X},
abstract = {Extremophiles are microorganisms that thrive in environments previously thought to be uninhabitable, including extreme temperature, salinity, pH, pressure, and radiation. These organisms, found in Archaea, Bacteria, and Eukarya, exhibit distinct structural, metabolic, and genetic adaptations, such as enhanced enzyme stability, efficient DNA repair mechanisms, and robust stress-response systems that enable survival under extreme conditions. Understanding these adaptation mechanisms is key to engineering similar traits in mesophilic organisms. This review discusses the diversity of extremophiles and presents phylogenetic and comparative genomic insights which may provide insights into the origins and evolution of early life on Earth We highlight recent advances in CRISPR/Cas-based genome editing, genome-scale metabolic modeling (GEM), and synthetic biology that have expanded the use of extremophiles in sustainable industrial biotechnology. The exceptional stability and catalytic efficiency of extremozymes under harsh conditions underscore their potential in various biotechnological applications. Finally, we discuss the ecological significance of extremophiles in climate change mitigation and outline current challenges and future directions in extremophile research.},
}

@article {pmid40692150,
year = {2026},
author = {Sun, Z and Cui, M and Deng, Z and Zhou, L and Zeng, F and Liu, Z and Liu, Y},
title = {Loss of CD99L2 Contributed to Temozolomide Resistance and Glioblastoma Tumorigenesis Based on Genome-scale CRISPR/Cas9 Screening.},
journal = {Current pharmaceutical design},
volume = {32},
number = {9},
pages = {696-709},
pmid = {40692150},
issn = {1873-4286},
support = {81974512//National Natural Science Foundation of China/ ; 2020JJ4894//Hunan Natural Science Foundation/ ; },
mesh = {Humans ; *Temozolomide/pharmacology ; *Glioblastoma/drug therapy/genetics/pathology/metabolism ; *Drug Resistance, Neoplasm/drug effects/genetics ; *Brain Neoplasms/drug therapy/genetics/pathology/metabolism ; *CRISPR-Cas Systems/genetics ; *Antineoplastic Agents, Alkylating/pharmacology ; *12E7 Antigen/genetics/metabolism ; Cell Proliferation/drug effects ; *Carcinogenesis/drug effects/genetics ; Cell Line, Tumor ; },
abstract = {INTRODUCTION: Glioblastoma Multiforme (GBM) is a highly aggressive and fatal brain malignancy, with Temozolomide (TMZ) serving as the first-line chemotherapeutic treatment. However, over 50% of patients do not respond to TMZ, and the underlying mechanisms remain unclear. This study utilized the GeCKO library to identify novel genes involved in TMZ resistance and to explore their functions.

METHODS: Loss-of-function genes related to TMZ resistance in GBM cells were identified using the GeCKO library and Next-Generation Sequencing (NGS), validated by qPCR and CCK-8 assays. CD99L2 function was assessed through proliferation, migration, and EdU assays in U251 and U87 cells. Tumor samples from 55 stage IV GBM patients were analyzed to explore the correlation between CD99L2 expression and Progression- Free Survival (PFS).

RESULTS: GeCKO library screening identified seven genes associated with TMZ resistance. After validation, CD99L2 was confirmed as a key contributor to TMZ resistance. Knockdown of CD99L2 increased the IC50 of U251 and U87 cells by 1.39- and 1.54-fold, respectively. Conversely, CD99L2 overexpression reduced the IC50 by 0.52- and 0.58-fold. CD99L2 knockdown also promoted tumor proliferation and aggressiveness. Additionally, higher CD99L2 expression was associated with longer PFS in GBM patients (median PFS: 7.87 months vs. 2.7 months, P=0.0003).

DISCUSSION: The functions of CD99L2 remain poorly understood. A few studies have reported that CD99L2 may serve as an adhesion molecule modulating inflammatory responses. One study has shown that CD99L2 is highly expressed in the brain and affects neuronal excitability. These findings suggest that CD99L2 may play a positive role in the body's defense against glioma.

CONCLUSION: This study demonstrated that CD99L2 knockdown promotes TMZ resistance and tumorigenesis in GBM, suggesting its potential as a novel biomarker for TMZ resistance.},
}

Humans
*Temozolomide/pharmacology
*Glioblastoma/drug therapy/genetics/pathology/metabolism
*Drug Resistance, Neoplasm/drug effects/genetics
*Brain Neoplasms/drug therapy/genetics/pathology/metabolism
*CRISPR-Cas Systems/genetics
*Antineoplastic Agents, Alkylating/pharmacology
*12E7 Antigen/genetics/metabolism
Cell Proliferation/drug effects
*Carcinogenesis/drug effects/genetics
Cell Line, Tumor

@article {pmid41319833,
year = {2026},
author = {Hu, H and Ke, X and Xiao, H and Shang, X and Yin, Q and Li, J and Li, X and Hu, Z and Qian, P and Wang, M},
title = {Genomic assembly, rescue, and characterization of a functional pseudorabies virus.},
journal = {Virologica Sinica},
volume = {41},
number = {1},
pages = {97-106},
doi = {10.1016/j.virs.2025.11.010},
pmid = {41319833},
issn = {1995-820X},
mesh = {Animals ; *Herpesvirus 1, Suid/genetics ; *Genome, Viral ; Chlorocebus aethiops ; Vero Cells ; Swine ; Mice ; Pseudorabies/virology ; Gene Editing ; Mice, Inbred BALB C ; CRISPR-Cas Systems ; Swine Diseases/virology ; },
abstract = {With recent advances in synthetic biology methods, the genomes of several large DNA viruses have been de novo synthesized and assembled, leading to the functional rescue of the respective viruses. Pseudorabies virus (PRV), a large DNA virus belonging to the family Herpesviridae, causes severe diseases in swine, resulting in significant economic losses to the global pig farming industry. Genome editing is crucial for attenuating virulence and developing safer vaccines for PRV. However, its complex repetitive sequences and extremely high GC-rich genome pose significant challenges for genetic manipulation. In this study, we developed a PRV genome assembly platform using yeast-based transformation-associated recombination (TAR) technology. The genome of a prevalent genotype II variant strain, PRV-GX-2011 (GenBank number PV405324.1), was divided into nine A-level fragments and cloned into vectors via TAR. Subsequently, three B-level fragments were generated by recombining three A-level fragments each. In vitro CRISPR/Cas9-mediated editing was introduced to insert an egfp gene into the non-coding intergenic region between UL23 and UL22 genes. Infectious viruses were rescued by co-transfection of linearized B-level fragments in Vero cells, and an isolated virus, PRV-GX-Syn1, was purified via plaque assay. While PRV-GX-Syn1 exhibited reduced viral titer and smaller plaque size compared to the parental strain, its morphological characteristics remained indistinguishable from the parental virus. In BALB/c mice, PRV-GX-Syn1 caused lethal infection, producing lung pathology comparable to the parental strain. This TAR-based platform offers faster and more flexible genomic modification of PRV, facilitating both basic research and PRV-based vaccine vectors.},
}

Animals
*Herpesvirus 1, Suid/genetics
*Genome, Viral
Chlorocebus aethiops
Vero Cells
Swine
Mice
Pseudorabies/virology
Gene Editing
Mice, Inbred BALB C
CRISPR-Cas Systems
Swine Diseases/virology

@article {pmid41611887,
year = {2026},
author = {Gordon, A and Yoon, SJ and Bicks, LK and Martín, JM and Pintacuda, G and Arteaga, S and Wamsley, B and Guo, Q and Elahi, L and Dolmetsch, RE and Bernstein, JA and O'Hara, R and Hallmayer, JF and Lage, K and Pasca, SP and Geschwind, DH},
title = {Developmental convergence and divergence in human stem cell models of autism.},
journal = {Nature},
volume = {651},
number = {8106},
pages = {707-719},
pmid = {41611887},
issn = {1476-4687},
mesh = {Humans ; *Induced Pluripotent Stem Cells/metabolism/pathology/cytology ; Organoids/metabolism/pathology/cytology ; *Autism Spectrum Disorder/genetics/pathology ; Mutation/genetics ; Cell Differentiation/genetics ; Neural Stem Cells/metabolism/cytology/pathology ; Protein Interaction Maps/genetics ; *Models, Biological ; CRISPR-Cas Systems/genetics ; Female ; Transcriptome/genetics ; Male ; Cell Line ; },
abstract = {Two decades of genetic studies in autism spectrum disorder (ASD) have identified more than 100 genes harbouring rare risk mutations[1-13]. Despite this substantial heterogeneity, transcriptomic and epigenetic analyses have identified convergent patterns of dysregulation across the ASD postmortem brain[14,15-17]. To identify shared and distinct mechanisms of ASD-linked mutations, we assembled a large patient collection of human induced pluripotent stem (hiPS) cells, consisting of 70 hiPS cell lines after stringent quality control representing 8 ASD-associated mutations, idiopathic ASD, and 20 lines from non-affected control individuals. Here we used these hiPS cell lines to generate human cortical organoids, profiling by RNA sequencing at four distinct time points up to 100 days after in vitro differentiation. Early time points harboured the largest mutation-specific changes, but different mutations converged on shared transcriptional changes as development progressed. We identified a shared RNA and protein interaction network, which was enriched in ASD risk genes and predicted to drive the observed downstream changes in gene expression. CRISPR-Cas9 screening of these candidate transcriptional regulators in induced human neural progenitors validated their downstream convergent molecular effects. These data illustrate how risk associated with genetically defined forms of ASD can propagate by means of transcriptional regulation to affect convergently dysregulated pathways, providing new insight into the convergent impact of ASD genetic risk on human neurodevelopment.},
}

Humans
*Induced Pluripotent Stem Cells/metabolism/pathology/cytology
Organoids/metabolism/pathology/cytology
*Autism Spectrum Disorder/genetics/pathology
Mutation/genetics
Cell Differentiation/genetics
Neural Stem Cells/metabolism/cytology/pathology
Protein Interaction Maps/genetics
*Models, Biological
CRISPR-Cas Systems/genetics
Female
Transcriptome/genetics
Male
Cell Line

@article {pmid41667457,
year = {2026},
author = {He, M and Wang, W and Zhou, H and Liu, C and Zhao, C and Li, J and Han, Y and Qin, Y and Chen, M},
title = {A synthetic system for RNA-responsive pyroptosis based on type III-E CRISPR nuclease-protease.},
journal = {Nature communications},
volume = {17},
number = {1},
pages = {},
pmid = {41667457},
issn = {2041-1723},
support = {//National Key R&D Program of China/ ; //National Natural Science Foundation of China/ ; //Natural Science Foundation of Wuhan/ ; },
mesh = {*Pyroptosis/genetics ; Humans ; *CRISPR-Cas Systems/genetics ; *RNA/genetics/metabolism ; Animals ; Mice ; HEK293 Cells ; },
abstract = {Pyroptosis plays a crucial role in immune defense against infections and endogenous threats by eliminating harmful cells and modulating the immune response through inflammation. However, the natural activation of pyroptosis involves intricate signaling pathways, posing significant challenges for its artificial manipulation in research and therapies. Here, we present DAMAGE (Death Manipulation Gene), an innovative system that integrates gasdermins within the type III-E CRISPR framework, enabling the specific recognition of target RNA (tgRNA) and triggering pyroptosis. This mechanism allows DAMAGE to selectively identify and eliminate virus-infected, cancerous, and senescent cells, all of which exhibit altered RNA transcriptomes. Additionally, DAMAGE exhibits considerable promise as a platform for mRNA-LNP therapy. Our study highlights the potential of this CRISPR-based system in the controllable induction of pyroptosis, offering an innovative therapeutic strategy for treating RNA-heterogeneous diseases.},
}

*Pyroptosis/genetics
Humans
*CRISPR-Cas Systems/genetics
*RNA/genetics/metabolism
Animals
Mice
HEK293 Cells

@article {pmid41679545,
year = {2026},
author = {Bahlmann, N and Alshawabkeh, M and Tsoukas, R and Schröer, K and Schellhorn, S and Sieler, M and Dittmar, T and Ehrke-Schulz, E and Ehrhardt, A and Zhang, W},
title = {Desmoglein 2 (DSG2)-knockout human respiratory epithelial cell model to study species B adenovirus receptor usage.},
journal = {Virologica Sinica},
volume = {41},
number = {1},
pages = {172-181},
doi = {10.1016/j.virs.2026.02.007},
pmid = {41679545},
issn = {1995-820X},
mesh = {Humans ; *Desmoglein 2/genetics/metabolism ; *Receptors, Virus/genetics/metabolism ; Gene Knockout Techniques ; Membrane Cofactor Protein/genetics/metabolism ; *Adenoviruses, Human/genetics/physiology ; CRISPR-Cas Systems ; *Epithelial Cells/virology/metabolism ; *Coxsackie and Adenovirus Receptor-Like Membrane Protein/genetics/metabolism ; Cell Line ; Cell Proliferation ; },
abstract = {With an increasing number of human adenoviruses identified, the selection of potential therapeutic vectors broadens. For safety reasons, achieving cell-specific gene delivery is crucial to minimize off-target effects. Therefore, it is essential to gain a systematic understanding of adenovirus receptor-usage. Our aim is to establish a human-originated in vitro model for comparative analysis of human adenoviruses receptor usage. Based on our previous work of human CD46 and coxsackievirus and adenovirus receptor (CAR) knockout cell lines, we generated desmoglein 2 (DSG2) knockout cell lines using genome-engineering technology based on Clustered Regularly Interspaced Short Palindromic Repeats and CRISPR-associated protein 9 (CRISPR/Cas9). All together, we established a panel of cell lines that carry a single, double, or triple knockout of the three major human adenovirus receptors: CAR, CD46 and DSG2. Notably, cell proliferation speed was affected by the CAR-knockout, but not the DSG2-or CD46-knockouts. In addition, the spheroid formation ability was sharply reduced in CAR- or DSG2-knockout cells, but not the CD46-knockout cells. With this receptor-knockout model, we confirmed the receptor usage of nine species B adenoviruses. Furthermore, adenovirus vectors containing a previously identified DSG2-binding affinity-enhanced mutation showed DSG2-dependent cell entry within this cell model, which indicates that they are de-targeted from CD46 - the ubiquitously expressed receptor on all nucleated cells. Collectively, our findings show that the adenovirus major receptor knockout cell lines can serve as an in vitro model to help select adenovirus types suitable for individual applications and to better understand adenovirus infection biology.},
}

Humans
*Desmoglein 2/genetics/metabolism
*Receptors, Virus/genetics/metabolism
Gene Knockout Techniques
Membrane Cofactor Protein/genetics/metabolism
*Adenoviruses, Human/genetics/physiology
CRISPR-Cas Systems
*Epithelial Cells/virology/metabolism
*Coxsackie and Adenovirus Receptor-Like Membrane Protein/genetics/metabolism
Cell Line
Cell Proliferation

@article {pmid41691970,
year = {2026},
author = {Anders, M and Hoppe, S and Eberl, H and Rebs, S and Seedorf, A and Maurer, W and Schill, T and Zibat, A and Unsöld, JK and Yigit, G and Wollnik, B and Vollmann, D and Sossalla, S and Streckfuss-Bömeke, K},
title = {Generation of pluripotent stem cell line (IPWi001-A) and a corresponding CRISPR/Cas9 modified isogenic rescue control (IPWi001-A-1) from a patient with arrhythmia-induced cardiomyopathy harboring a KCNQ1 truncating mutation.},
journal = {Stem cell research},
volume = {92},
number = {},
pages = {103921},
doi = {10.1016/j.scr.2026.103921},
pmid = {41691970},
issn = {1876-7753},
mesh = {Humans ; *KCNQ1 Potassium Channel/genetics/metabolism ; *CRISPR-Cas Systems/genetics ; *Induced Pluripotent Stem Cells/metabolism/cytology ; *Arrhythmias, Cardiac/genetics/complications/pathology ; *Cardiomyopathies/genetics/pathology ; *Mutation/genetics ; Cell Line ; Myocytes, Cardiac/metabolism ; Cell Differentiation ; Male ; },
abstract = {KCNQ1 functions as a slow rectifying potassium channel during the repolarization of the cardiac action potential, with mutations causing long-QT syndrome 1 and arrhythmias. A genetic link between KCNQ1 mutations and arrhythmia-induced cardiomyopathy (AIC) has not been identified, and the underlying pathways remain elusive. We generated human induced pluripotent stem cells (hiPSCs) from an AIC patient harboring the heterozygous truncating mutation p.W15* in KCNQ1 and corrected the mutation to wildtype using CRISPR/Cas9. The hiPSCs retained full pluripotency, genomic integrity, and differentiation ability. They were differentiated into hiPSC-cardiomyocytes (hiPSC-CM), establishing a patient-specific model to explore potential genetic connections to AIC.},
}

Humans
*KCNQ1 Potassium Channel/genetics/metabolism
*CRISPR-Cas Systems/genetics
*Induced Pluripotent Stem Cells/metabolism/cytology
*Arrhythmias, Cardiac/genetics/complications/pathology
*Cardiomyopathies/genetics/pathology
*Mutation/genetics
Cell Line
Myocytes, Cardiac/metabolism
Cell Differentiation
Male

@article {pmid41693245,
year = {2026},
author = {Kou, S and Chua, LC and Tan, JQ and Lau, OS},
title = {Stomatal XVE: an inducible system for cell-stage-specific gene expression and editing in the stomatal lineage.},
journal = {The New phytologist},
volume = {250},
number = {2},
pages = {1330-1347},
pmid = {41693245},
issn = {1469-8137},
support = {NRF-CRP22-2019-0001//National Research Foundation Singapore/ ; },
mesh = {*Plant Stomata/genetics/cytology ; *Arabidopsis/genetics/cytology ; *Gene Expression Regulation, Plant/drug effects ; *Gene Editing/methods ; *Cell Lineage/genetics ; Arabidopsis Proteins/metabolism/genetics ; Promoter Regions, Genetic/genetics ; CRISPR-Cas Systems/genetics ; Plants, Genetically Modified ; Phenotype ; },
abstract = {Stomatal development has emerged as a valuable model for studying developmental processes. Examining gene function along the stomatal lineage often requires gene perturbation in a controlled and cell-stage-specific manner, but this remains tedious without a dedicated genetic tool. Here, we describe Stomatal XVE, a modular, two-component XVE-based inducible system that enables user-controlled gene overexpression and Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-based knockout at defined stomatal cell stages in Arabidopsis thaliana. The system consists of a collection of estrogen-responsive XVE driver lines under cell-stage-specific promoters and effector vectors responsive to activated XVE. This design simplifies cloning and allows users to scale their investigation. We validated the cell-stage specificity and inducibility of the XVE driver lines and characterized key induction parameters. To test the system functionally, we employed it to study MAPKKK YODA and a pathogen effector AvrPtoB. While YODA overexpression reproduced known early- and late-stage phenotypes, stage-specific knockouts argued against its late-stage role in guard cell (GC) differentiation. Furthermore, AvrPtoB expression during later stages triggered striking disruptions in GC morphology and viability, revealing cell-type-specific effects of the pathogen protein. Overall, our Stomatal XVE system enables precise functional analysis of genes across defined stages of stomatal development and is particularly well suited for investigating genes with pleiotropic effects.},
}

*Plant Stomata/genetics/cytology
*Arabidopsis/genetics/cytology
*Gene Expression Regulation, Plant/drug effects
*Gene Editing/methods
*Cell Lineage/genetics
Arabidopsis Proteins/metabolism/genetics
Promoter Regions, Genetic/genetics
CRISPR-Cas Systems/genetics
Plants, Genetically Modified
Phenotype

@article {pmid41722369,
year = {2026},
author = {Boumpoureka, I and Gorgogietas, V and Petkovski, E and Massart, F and Mellick, GD and Krüger, R},
title = {Generation of two isogenic control iPSC lines (LCSBi001-A-2 and LCSBi001-A-3) from a Parkinson's disease patient line (LCSBi001-A) carrying the pathogenic VPS35 p.D620N mutation.},
journal = {Stem cell research},
volume = {92},
number = {},
pages = {103944},
doi = {10.1016/j.scr.2026.103944},
pmid = {41722369},
issn = {1876-7753},
mesh = {Humans ; *Parkinson Disease/genetics/pathology/metabolism ; *Induced Pluripotent Stem Cells/metabolism/cytology/pathology ; *Vesicular Transport Proteins/genetics ; *Mutation/genetics ; Cell Line ; CRISPR-Cas Systems ; Cell Differentiation ; },
abstract = {The pathogenic mutation VPS35 p.D620N has been identified to cause autosomal dominant, late-onset Parkinson's disease (PD) in multiple individuals and families worldwide. Here, we describe the generation of two new isogenic control lines (LCSBi001-A-2 and LCSBi001-A-3) from an already established patient-derived line (LCSBi001-A) carrying the heterozygous VPS35 p.D620N mutation. The control lines were generated with CRISPR/Cas9 technology, and the correction of the mutation was verified with Sanger sequencing. Both lines express pluripotency markers, are capable of in vitro differentiation into the three germ layers, and have a normal karyotype. The mutant and control iPSC lines are available for research purposes.},
}

Humans
*Parkinson Disease/genetics/pathology/metabolism
*Induced Pluripotent Stem Cells/metabolism/cytology/pathology
*Vesicular Transport Proteins/genetics
*Mutation/genetics
Cell Line
CRISPR-Cas Systems
Cell Differentiation

@article {pmid41747624,
year = {2026},
author = {Ming Wu, SC and Boheler, KR},
title = {Generation of an induced pluripotent stem cell and isogenic control line from a vascular Ehlers-Danlos Syndrome (vEDS) patient harboring a pathogenic c.755G>T in the COL3A1 gene.},
journal = {Stem cell research},
volume = {92},
number = {},
pages = {103945},
doi = {10.1016/j.scr.2026.103945},
pmid = {41747624},
issn = {1876-7753},
mesh = {Humans ; *Ehlers-Danlos Syndrome/genetics/pathology/metabolism ; *Induced Pluripotent Stem Cells/metabolism/cytology/pathology ; *Collagen Type III/genetics/metabolism ; Cell Line ; Cell Differentiation ; Fibroblasts/metabolism ; CRISPR-Cas Systems ; Ehlers-Danlos Syndrome, Type IV ; },
abstract = {We report the generation and characterization of a collagen III-mutant human iPSC line (JHUi007-A) and an isogenic gene-edited control (JHUi007-A-1). Reprogramming of dermal fibroblasts, obtained from a patient with vascular Ehlers-Danlos syndrome (vEDS) carrying the COL3A1 c.755G>T variant, was performed using integration-free Sendai virus. Isogenic controls were produced by CRISPR/Cas9 gene editing. Both lines displayed typical morphologies, expressed stemness factors, formed derivatives of all three germ layers, and maintained a normal karyotype. These lines readily differentiated into vascular smooth muscle cells with cytoskeletal differences between vEDS and control cells, confirming the utility of this resource to study disease processes.},
}

Humans
*Ehlers-Danlos Syndrome/genetics/pathology/metabolism
*Induced Pluripotent Stem Cells/metabolism/cytology/pathology
*Collagen Type III/genetics/metabolism
Cell Line
Cell Differentiation
Fibroblasts/metabolism
CRISPR-Cas Systems
Ehlers-Danlos Syndrome, Type IV

@article {pmid41747626,
year = {2026},
author = {Yoo, DH and Bayarsaikhan, D and Lee, J and Im, YS and Bayarsaikhan, G and Kang, HA and Lee, B and Kim, YO},
title = {Generation and characterization of SOX17-specific EGFP expressing human induced pluripotent stem cell line, KSCBi017-A-4, using CRISPR/Cas9.},
journal = {Stem cell research},
volume = {92},
number = {},
pages = {103943},
doi = {10.1016/j.scr.2026.103943},
pmid = {41747626},
issn = {1876-7753},
mesh = {Humans ; *Induced Pluripotent Stem Cells/metabolism/cytology ; *CRISPR-Cas Systems/genetics ; *SOXF Transcription Factors/metabolism/genetics ; *Green Fluorescent Proteins/metabolism/genetics ; Cell Line ; Cell Differentiation ; Endoderm/cytology/metabolism ; },
abstract = {We generated a human induced pluripotent stem cell (hiPSC) reporter line in which EGFP was inserted in-frame at the C-terminus of the endogenous SOX17 locus using CRISPR/Cas9-mediated homologous recombination. The targeted clone, KSCBi017-A-4, was isolated by puromycin selection and validated by PCR and Sanger sequencing. This SOX17-EGFP hiPSC line retains a normal karyotype and pluripotency and displays specific EGFP expression upon directed definitive endoderm differentiation. This reporter line provides a reliable tool for monitoring SOX17 expression during human endoderm specification.},
}

Humans
*Induced Pluripotent Stem Cells/metabolism/cytology
*CRISPR-Cas Systems/genetics
*SOXF Transcription Factors/metabolism/genetics
*Green Fluorescent Proteins/metabolism/genetics
Cell Line
Cell Differentiation
Endoderm/cytology/metabolism

@article {pmid41763219,
year = {2026},
author = {Aravind, KM and Del Vecchio, D},
title = {Resource competition shapes CRISPR-mediated gene activation.},
journal = {Cell systems},
volume = {17},
number = {3},
pages = {101511},
doi = {10.1016/j.cels.2025.101511},
pmid = {41763219},
issn = {2405-4720},
mesh = {*CRISPR-Cas Systems/genetics ; RNA, Guide, CRISPR-Cas Systems/genetics/metabolism ; *Clustered Regularly Interspaced Short Palindromic Repeats/genetics ; Humans ; Gene Editing/methods ; *Transcriptional Activation/genetics ; },
abstract = {CRISPR-mediated gene activation (CRISPRa) allows concurrent transcriptional control of many genes and is widely used in genome-wide screening, bioproduction, and therapeutics. Multi-gene control is possible due to the sequence specificity by which guide RNAs (gRNAs) recruit dCas9 and an activator protein to target genes. Still, the optimization of CRISPRa systems remains difficult. Here, we show that, despite sequence specificity, different gRNAs interfere with each other by competing for dCas9 and the activator protein. This competition breaks modularity and hinders CRISPRa. We also discover that gene activation is biphasic, wherein increased level of a gRNA leads to target repression instead of activation. We introduce a chemical reaction-network model that captures these effects and use it for improving the dynamic range of CRISPRa. Our results demonstrate that CRISPRa is not as modular or scalable as previously thought and establish a predictive modeling tool that enables systematic design and optimization of multi-gRNA CRISPRa systems. A record of this paper's transparent peer review process is included in the supplemental information.},
}

*CRISPR-Cas Systems/genetics
RNA, Guide, CRISPR-Cas Systems/genetics/metabolism
*Clustered Regularly Interspaced Short Palindromic Repeats/genetics
Humans
Gene Editing/methods
*Transcriptional Activation/genetics

@article {pmid41814120,
year = {2026},
author = {Simonneau, B and Baghdoyan, S and Cailleret, M and Simon, S and Ruckebusch, O and Vrablikova, B and Giraud-Triboult, K and Kassar, LE and Fanen, P and Duriez, B},
title = {CRISPR-Cas9 genome editing in the parental iPSC line PCIi033-A to introduce the homozygous mutation p.F508del (c.1521_1523del) in the CFTR gene.},
journal = {Stem cell research},
volume = {92},
number = {},
pages = {103947},
doi = {10.1016/j.scr.2026.103947},
pmid = {41814120},
issn = {1876-7753},
mesh = {Humans ; *Induced Pluripotent Stem Cells/metabolism/cytology ; *Cystic Fibrosis Transmembrane Conductance Regulator/genetics ; *CRISPR-Cas Systems/genetics ; *Gene Editing/methods ; Homozygote ; Cell Line ; *Mutation/genetics ; Cystic Fibrosis/genetics/pathology ; Cell Differentiation ; },
abstract = {Cystic Fibrosis (CF) is an autosomal recessive disease caused by mutations in the CFTR gene. Patients carrying the most common mutation, p.F508del, benefit from the triple therapy Kaftrio®. We genome-edited the commercially available iPSC line PCIi033-A (wild-type CFTR) to generate the subclone PCIi033-A-5, which is homozygous for the mutation c.1521_1523del (p.F508del), using CRISPR-SpCas9 tools. PCIi033-A-5 has a normal karyotype and stem cell morphology, is pluripotent, and differentiates into the three germ layers. Introducing this mutation in a parental isogenic iPSC line is essential to demonstrate the feasibility of modeling CF disease after differentiation of the iPS cells into bronchial epithelium.},
}

Humans
*Induced Pluripotent Stem Cells/metabolism/cytology
*Cystic Fibrosis Transmembrane Conductance Regulator/genetics
*CRISPR-Cas Systems/genetics
*Gene Editing/methods
Homozygote
Cell Line
*Mutation/genetics
Cystic Fibrosis/genetics/pathology
Cell Differentiation

@article {pmid41826758,
year = {2026},
author = {Deol, KK and Harris, CA and Tomlinson, SJ and Delaney, CJ and Al-Farhan, A and Mathiowetz, AJ and Doubravsky, CE and Pratt, DA and Olzmann, JA},
title = {Vitamin B2 metabolism promotes FSP1 stability to prevent ferroptosis.},
journal = {Nature structural & molecular biology},
volume = {33},
number = {3},
pages = {525-536},
pmid = {41826758},
issn = {1545-9985},
mesh = {*Ferroptosis ; Humans ; *Riboflavin/metabolism ; *S100 Calcium-Binding Protein A4/metabolism/genetics/chemistry ; Protein Stability ; Flavin-Adenine Dinucleotide/metabolism ; CRISPR-Cas Systems ; HEK293 Cells ; },
abstract = {Ferroptosis, a regulated form of cell death driven by excessive lipid peroxidation, has emerged as a promising therapeutic target in cancer. Ferroptosis suppressor protein 1 (FSP1) is a critical regulator of ferroptosis resistance, yet the mechanisms controlling its expression and stability remain mostly unexplored. To uncover regulators of FSP1 abundance, we conducted CRISPR-Cas9 screens using a genome-edited, dual-fluorescent FSP1 reporter cell line, identifying both transcriptional and post-translational mechanisms that determine FSP1 levels. Notably, we identified riboflavin kinase and flavin adenine dinucleotide (FAD) synthase, enzymes that are essential for synthesizing FAD from vitamin B2, as key contributors to FSP1 stability. Biochemical and cellular analyses revealed that FAD binding is critical for both FSP1 activity and stability. FAD deficiency and mutations blocking FSP1-FAD binding triggered FSP1 degradation through a ubiquitin-proteasome pathway involving the E3 ligase RNF8. Unlike other vitamins that inhibit ferroptosis by scavenging radicals, vitamin B2 supports ferroptosis resistance through FAD cofactor binding, ensuring proper FSP1 stability and function. This study provides a rich resource detailing mechanisms that regulate FSP1 abundance and highlights a novel connection between vitamin B2 metabolism and ferroptosis resistance, with implications for therapeutic strategies targeting FSP1 in cancer.},
}

*Ferroptosis
Humans
*Riboflavin/metabolism
*S100 Calcium-Binding Protein A4/metabolism/genetics/chemistry
Protein Stability
Flavin-Adenine Dinucleotide/metabolism
CRISPR-Cas Systems
HEK293 Cells

@article {pmid41829816,
year = {2026},
author = {Al-Sawa'eer, AS and Al-Samydai, A and Odeh, L and Haj Ahmad, F and Obekh, R and Elqader, YMA and Khaleel, A and Al-Athamneh, AM and Gabriele, M and Di Simone, SC and Ferrante, C and Menghini, L and Ali Agha, ASA},
title = {Early Plant Development as a Systems-Level Trait: Integrating Omics, Artificial Intelligence, and Emerging Biotechnologies.},
journal = {Plants (Basel, Switzerland)},
volume = {15},
number = {5},
pages = {},
pmid = {41829816},
issn = {2223-7747},
abstract = {Seed germination and early seedling development are critical determinants of crop establishment, stress tolerance, and yield stability, yet these stages remain insufficiently integrated into contemporary crop improvement strategies. Recent advances across genome editing, microbiome-assisted seed treatments, nanotechnology-enabled priming, and artificial intelligence-guided phenotyping have generated substantial but fragmented insights into early developmental regulation. This review synthesizes recent advances across early plant development research. It demonstrates that seemingly diverse technologies converge on a limited set of regulatory control nodes, including abscisic acid-gibberellin balance, redox homeostasis, and root system architectural plasticity. By integrating evidence from molecular, microbial, physicochemical, and computational studies, early plant ontogeny is presented as a tunable regulatory state governed by quantitative thresholds rather than as a strictly predetermined genetic process. Advances in deep learning, reinforcement learning, and high-throughput phenotyping further enable the modeling and optimization of early developmental trajectories across genotype by environment contexts. Together, these insights establish early development as a programmable target for crop improvement and provide a mechanistic foundation for designing integrated interventions that enhance developmental uniformity, stress resilience, and yield stability across diverse agroecological systems.},
}

@article {pmid41830331,
year = {2026},
author = {Martínez-Alvarez, L and Peng, X},
title = {Diversity and evolution of archaeal immune strategies.},
journal = {Nucleic acids research},
volume = {54},
number = {5},
pages = {},
pmid = {41830331},
issn = {1362-4962},
support = {DFF-0135-00402//Danish Council for Independent Research/ ; 10.46540/4264-00120B//Danish Council for Independent Research/ ; NNF17OC0031154//Novo Nordisk Foundation/ ; DeiC-KU-N1-2024089//Danish e-Infrastructure Consortium/ ; DFF-0135-00402//Natural Sciences/ ; 10.46540/4264-00120B//Natural Sciences/ ; },
mesh = {*Archaea/genetics/immunology/classification ; Phylogeny ; *Genome, Archaeal ; CRISPR-Cas Systems/genetics ; *Evolution, Molecular ; Bacteria/genetics/immunology ; },
abstract = {Archaeal antiviral defense systems remain poorly characterized despite recent advances in understanding prokaryotic immunity. Here, we analyze 7747 archaeal genomes, the largest and most diverse dataset to date, revealing a striking disparity in defense system prevalence and diversity compared to Bacteria. Nearly one-third of archaeal genomes have no detected systems beyond CRISPR-Cas and restriction-modification (in contrast to only 2.2% bacterial genomes), and only 50-55% contain CRISPR-Cas systems, far below previous estimates. Many known defense systems appear restricted to Bacteria, while several single-gene putative candidate systems (PDCs) recently identified through a guilt-by-embedding approach are enriched in Archaea. Phylogenetic analyses suggest that PDC-S70 and PDC-M05 likely originated in Archaea, representing rare archaeal contributions to the prokaryotic immune repertoire. Consistent with earlier studies, our findings support the existence of deep evolutionary links between archaeal and eukaryotic systems for argonautes and viperins. These analyses highlight both the underexplored nature and the evolutionary significance of archaeal immunity, calling for expanded efforts to uncover archaeal-specific systems and improve our understanding of immune evolution across domains of life.},
}

*Archaea/genetics/immunology/classification
Phylogeny
*Genome, Archaeal
CRISPR-Cas Systems/genetics
*Evolution, Molecular
Bacteria/genetics/immunology

@article {pmid41830765,
year = {2026},
author = {Leandro, K and Rufino-Ramos, D and Lopes, SM and Silva, FS and Rodrigues-Santos, P and Silva, AC and Fernandes, AR and Henriques, C and Pereira, D and Lobo, D and Gabriel, GL and Faro, R and Nobre, RJ and Perdigão, PRL and Kleinstiver, BP and de Almeida, LP},
title = {Extracellular vesicles-mediated delivery of SpCas9 RNPs for therapeutic gene editing in Spinocerebellar Ataxia Type 3.},
journal = {Biomaterials},
volume = {331},
number = {},
pages = {124119},
doi = {10.1016/j.biomaterials.2026.124119},
pmid = {41830765},
issn = {1878-5905},
abstract = {Spinocerebellar Ataxia Type 3 (SCA3) is a neurodegenerative dominantly-inherited disorder caused by an overexpansion of a CAG tract within the ATXN3 gene, conferring toxic properties to the ataxin-3 protein. Genome editing with CRISPR-Cas9 enzymes is a promising strategy to inactivate mutant ATXN3 alleles, however, in vivo delivery remains challenging. Extracellular vesicles (EVs) are promising delivery vehicles for Cas9 and single guide RNA (sgRNA) ribonucleoproteins that minimize genomic exposure to highly active endonucleases. In this study, we designed SpCas9 with a palmitoylation motif that enables SpCas9 and sgRNA enrichment into EVs. Introduction of a photocleavable linker - PhoCl - allowed the photo-inducible release of SpCas9 from the palmitoylation motif in EVs, increasing target engagement to ATXN3 in vitro. EVs loaded with SpCas9 ribonucleoproteins resulted in ATXN3 knockout in SCA3 patient-derived iPSCs and two SCA3 animal models. These findings highlight an innovative route for transient delivery of gene editing tools. This approach provides a promising therapeutic platform for the treatment of genetic diseases, including SCA3.},
}

@article {pmid41831437,
year = {2026},
author = {Yang, J and Wang, T and Liu, Z and Wu, W and Sun, Y and Zhan, Y and Zhang, S and Chen, H and Liu, B and Yue, C and Yin, Z and Shan, Z and Li, X and Li, Z and Yuan, Z and Yin, H and Zhang, H},
title = {Molecular basis for dual-spacer-guided target cleavage by the TIGR-TasH system.},
journal = {Molecular cell},
volume = {},
number = {},
pages = {},
doi = {10.1016/j.molcel.2026.02.017},
pmid = {41831437},
issn = {1097-4164},
abstract = {The RNA-directed programmable nuclease systems, exemplified by the CRISPR-Cas system, have been widely used in genome editing. In contrast to the single-spacer configuration of CRISPR RNA (crRNA), the guide RNA (tigRNA) of the tandem interspaced guide RNA (TIGR) system features a dual-spacer arrangement, thereby directing the TIGR-associated (Tas) protein to engage both strands of the target double-stranded DNA (dsDNA). Here, we determine six cryo-electron microscopy structures of the Salicola phage TIGR-TasH complex. The central coiled-coil region of TasH mediates dimerization, while the C-terminal nucleolar protein (Nop) domain is able to autonomously process precursor tigRNA. Upon target binding, the dynamic N-terminal HNH nuclease domain is recruited for cleavage through a β-hairpin, which also determines the target preference. More interestingly, the conserved box C motif of tigRNA stabilizes this β-hairpin in an adenine-specific manner, enabling us to rationally design a guide RNA-defined nickase, distinct from conventional protein-based nickase strategies used in genome editing.},
}

@article {pmid41831575,
year = {2026},
author = {Park, E and Lee, S and Kim, D and Choi, Y and Choung, S and Kim, H and Kim, SG},
title = {Virus-induced genome editing enables functional genomics across diverse plant species.},
journal = {Molecules and cells},
volume = {},
number = {},
pages = {100348},
doi = {10.1016/j.mocell.2026.100348},
pmid = {41831575},
issn = {0219-1032},
abstract = {Virus-induced genome editing (VIGE) is expanding plant functional genomics by enabling precise and heritable genome modification across diverse species. While classical model systems such as Arabidopsis thaliana have provided foundational genetic insights, many ecologically, agriculturally, and chemically important traits reside in species that remain difficult to manipulate genetically. By coupling CRISPR-Cas systems with plant viral vectors, VIGE bypasses key limitations of conventional transformation and enables rapid mutagenesis without repeated tissue culture and plant regeneration. This approach enables researchers to examine gene function in species selected for biological relevance rather than technical convenience. Here, we review the conceptual framework, technical considerations, and applications of VIGE, and discuss its current limitations and future prospects in ecological, evolutionary, and crop research.},
}

@article {pmid41831871,
year = {2026},
author = {Li, J and Cai, W and Wang, L and He, J and Meng, Y and Liu, J and Xu, L and Feng, K and Shen, G and Wei, P and He, L},
title = {Functional analysis of inducible choline/carboxylesterase CCE01 associated with fenpropathrin and abamectin detoxification in Tetranychus urticae (Koch).},
journal = {Pesticide biochemistry and physiology},
volume = {219},
number = {},
pages = {107003},
doi = {10.1016/j.pestbp.2026.107003},
pmid = {41831871},
issn = {1095-9939},
mesh = {Animals ; *Ivermectin/analogs & derivatives/metabolism/toxicity/pharmacology ; *Pyrethrins/metabolism/toxicity ; *Tetranychidae/drug effects/enzymology/genetics/metabolism ; *Acaricides/metabolism/toxicity/pharmacology ; Drosophila melanogaster/genetics ; Inactivation, Metabolic ; *Carboxylesterase/metabolism/genetics ; Arthropod Proteins/genetics/metabolism ; CRISPR-Cas Systems ; },
abstract = {The two-spotted spider mite, Tetranychus urticae, represents a globally significant and highly destructive polyphagous pest, notorious for its capacity to develop severe resistance to numerous acaricides. Our previous investigative work identified a specific esterase gene, designated TuCCE01, which was consistently found to be over-expressed in multiple resistant mite populations. This study systematically investigated the functional role of the esterase gene TuCCE01 in metabolizing acaricides. Quantitative real-time PCR revealed that TuCCE01 expression in a susceptible strain was significantly induced by sublethal doses of fenpropathrin and abamectin, showing 3.6-fold and 3.8-fold increases after 24 h, respectively. RNA interference-mediated knockdown of TuCCE01 increased mite susceptibility to both acaricides. Critically, CRISPR/Cas9-mediated gene editing was successfully employed to generate homozygous TuCCE01 knockout mutant lines. Bioassays demonstrated that this specific knockout strain exhibited significantly increased sensitivity to both fenpropathrin and abamectin, with LC50 values markedly reduced. Conversely, transgenic Drosophila melanogaster overexpressing TuCCE01 exhibited enhanced tolerance. Importantly, heterologous expression and in vitro incubation assays confirmed that recombinant TuCCE01 could effectively deplete both acaricides. Homology modeling and molecular docking demonstrated that both acaricides bind to the substrate-binding pocket of TuCCE01, indicating direct enzymatic metabolism, which was consistent with the metabolic assays. This study enhances our understanding of CCE-mediated acaricide acute detoxification and provides a candidate molecular marker for future resistance monitoring.},
}

Animals
*Ivermectin/analogs & derivatives/metabolism/toxicity/pharmacology
*Pyrethrins/metabolism/toxicity
*Tetranychidae/drug effects/enzymology/genetics/metabolism
*Acaricides/metabolism/toxicity/pharmacology
Drosophila melanogaster/genetics
Inactivation, Metabolic
*Carboxylesterase/metabolism/genetics
Arthropod Proteins/genetics/metabolism
CRISPR-Cas Systems

@article {pmid40181169,
year = {2026},
author = {Sun, Y and Cao, Y and Song, Y and Li, J and Hou, Y and Huang, W and Xie, G and Yang, W and Zhang, R},
title = {Improved RNA base editing with guide RNAs mimicking highly edited endogenous ADAR substrates.},
journal = {Nature biotechnology},
volume = {44},
number = {3},
pages = {464-476},
pmid = {40181169},
issn = {1546-1696},
support = {2021A1515012463//Guangdong Science and Technology Department (Science and Technology Department, Guangdong Province)/ ; },
mesh = {*Adenosine Deaminase/genetics/metabolism ; *RNA Editing/genetics ; Humans ; Animals ; Mice ; *RNA, Guide, CRISPR-Cas Systems/genetics ; Gene Editing/methods ; *RNA-Binding Proteins/genetics/metabolism ; Alu Elements/genetics ; HEK293 Cells ; },
abstract = {Adenosine deaminase acting on RNA (ADAR)-mediated RNA base editing offers a safer alternative to genome editing for specific clinical applications because of nonpermanent editing of targets. Current guide RNA (gRNA) designs feature a fully complementary specificity domain with an A-C mismatch at the targeted adenosine. However, perfectly matched dsRNA is not the most effective ADAR substrate. Here we introduce MIRROR (mimicking inverted repeats to recruit ADARs using engineered oligoribonucleotides), an approach that implements structural motifs derived from highly edited inverted Alu repeats in human tissues to enable rational gRNA design for ADAR recruitment. We demonstrated that MIRROR is applicable to both short chemically synthesized gRNAs with modifications and long biologically generated gRNAs and surpasses current state-of-the-art approaches in both gRNA forms. It enhances editing efficiency by up to 5.7-fold in multiple human cell types and primary hepatocytes from an alpha-1 antitrypsin deficiency mouse model. Our findings improve programmable RNA editing in vitro and in vivo by rational design through the screening of highly edited natural substrate mimics.},
}

*Adenosine Deaminase/genetics/metabolism
*RNA Editing/genetics
Humans
Animals
Mice
*RNA, Guide, CRISPR-Cas Systems/genetics
Gene Editing/methods
*RNA-Binding Proteins/genetics/metabolism
Alu Elements/genetics
HEK293 Cells

@article {pmid40258958,
year = {2026},
author = {Hu, J and Guo, M and Gao, Q and Jia, H and He, M and Wang, Z and Guo, L and Liu, G and Gao, Q and Zhao, KT},
title = {QBEmax is a sequence-permuted and internally protected base editor.},
journal = {Nature biotechnology},
volume = {44},
number = {3},
pages = {387-393},
pmid = {40258958},
issn = {1546-1696},
support = {Z241100009024035//Beijing Municipal Science and Technology Commission/ ; },
mesh = {*Gene Editing/methods ; CRISPR-Cas Systems/genetics ; *Cytosine/chemistry ; Molecular Dynamics Simulation ; INDEL Mutation ; Base Sequence ; Humans ; },
abstract = {Cytosine base editors (CBEs) show promise for multiplex gene knockout applications, but impure edits, indels and off-targets still frequently occur. We describe here QBEmax, which exhibits high efficiency, low indel and off-targets and high product purity with up to 99.8% of edits comprised of C-to-T. Through molecular dynamic modeling, QBEmax presents as a compact and stable base editor that shields protected bases from undesired repair processes.},
}

*Gene Editing/methods
CRISPR-Cas Systems/genetics
*Cytosine/chemistry
Molecular Dynamics Simulation
INDEL Mutation
Base Sequence
Humans

@article {pmid41498761,
year = {2026},
author = {Fan, X and Zhang, R and Yang, L and Chen, S and He, M and Wang, Y and Huang, L and Zeng, J and Wu, S and Gao, M and Wang, H},
title = {CRISPR-Cas9-Loaded Theranostic Liposomes for Enhancing Radiosensitization of Prostate Cancer through POLD4 Gene Editing under Real-Time MRI Monitoring.},
journal = {Advanced science (Weinheim, Baden-Wurttemberg, Germany)},
volume = {13},
number = {16},
pages = {e19704},
doi = {10.1002/advs.202519704},
pmid = {41498761},
issn = {2198-3844},
support = {SHDC22011310-A//Shanghai Municipal Hospital Development Center/ ; SHDC22025311-B//Shanghai Municipal Hospital Development Center/ ; 25ZR1401440//Science and Technology Commission of Shanghai Municipality/ ; 82271969//National Natural Science Foundation of China/ ; 202101070002E00085//Major Program of Shanghai Municipal Commission of Education/ ; },
mesh = {Male ; *Prostatic Neoplasms/genetics/radiotherapy/therapy/diagnostic imaging ; *CRISPR-Cas Systems/genetics ; Humans ; *Liposomes ; Animals ; *Magnetic Resonance Imaging/methods ; *Gene Editing/methods ; Mice ; Cell Line, Tumor ; Theranostic Nanomedicine/methods ; Genetic Therapy/methods ; },
abstract = {Radiotherapy is a fundamental treatment for prostate cancer; however, its therapeutic efficacy is frequently limited by radioresistance mediated through DNA repair mechanisms and other biological factors. Although gene therapy holds promise for overcoming such resistance, identifying effective radiosensitization targets and developing efficient gene delivery systems remain practically challenging. In this study, transcriptomic analysis of radiotherapy-treated prostate cancer cells revealed a marked upregulation of DNA polymerase delta subunit 4 (POLD4), a target that has not been systematically studied. To evaluate the potential of POLD4 for overcoming radioresistance, CRISPR-Cas9-based plasmids along with ultrasmall superparamagnetic iron oxide nanoparticles (USPIONs) were encapsulated within cationic liposomes for achieving an MRI-trackable gene delivery platform (plasmid and iron oxide co-loaded liposomes, termed PIO@Lipo). Comprehensive in vitro and in vivo studies demonstrated that PIO@Lipo enabled efficient POLD4 knockdown. Furthermore, PIO@Lipo synergized with radiotherapy to induce extensive DNA damage, promote tumor cell apoptosis, and remodel the immunosuppressive microenvironment. Notably, PIO@Lipo displayed superior MRI contrast enhancement capability and passive tumor targeting ability. In conclusion, this study has identified POLD4 as a potent target for radiosensitization, capable of disrupting DNA damage-repair homeostasis through MRI-monitored gene editing, thereby offering a promising strategy to enhance the efficacy of radiotherapy in prostate cancer.},
}

Male
*Prostatic Neoplasms/genetics/radiotherapy/therapy/diagnostic imaging
*CRISPR-Cas Systems/genetics
Humans
*Liposomes
Animals
*Magnetic Resonance Imaging/methods
*Gene Editing/methods
Mice
Cell Line, Tumor
Theranostic Nanomedicine/methods
Genetic Therapy/methods

@article {pmid41644834,
year = {2026},
author = {Zhao, C and Guo, H and Chen, J and Tan, Y and Wu, Z and Zhang, Y and Li, Y and Wang, J and Wang, R and Zhang, H and Wang, D},
title = {An AuPt nanozyme-assisted CRISPR/Cas12a system for visual nucleic acid detection of pathogens.},
journal = {Analytical and bioanalytical chemistry},
volume = {418},
number = {7},
pages = {1997-2009},
pmid = {41644834},
issn = {1618-2650},
mesh = {*CRISPR-Cas Systems ; *Gold/chemistry ; Solanum tuberosum/microbiology ; *Platinum/chemistry ; *Alternaria/genetics/isolation & purification ; *Metal Nanoparticles/chemistry ; *Plant Diseases/microbiology ; Biosensing Techniques/methods ; Limit of Detection ; *DNA, Fungal/analysis/genetics ; },
abstract = {Potato early blight, caused by Alternaria solani, presents a significant threat to the potato industry. Existing detection methods for A. solani often fail to simultaneously achieve simplicity and accuracy. A gold-platinum (AuPt) nanozyme-assisted CRISPR/Cas12a system, termed the nanoparticle enzyme-assisted CRISPR detection (NACD assay) was developed. By integrating the precise target recognition of CRISPR with the enzyme-like activity of AuPt nanozymes, this system achieves simple, sensitive, and visual detection of A. solani. The NACD assay provided visual results through a distinct color change produced by the substrate catalyzed by the AuPt nanozyme. It can detect 100 copies/μL of the target dsDNA (A. solani 5.8S rRNA gene) and 10[-][3] ng/μL A. solani genomic DNA. This detection method demonstrates high specificity, with no cross-reactivity observed with three other pathogens. Moreover, the incorporation of a filter paper-based readout enables straightforward visual detection by the naked eye, making it particularly suitable for on-site testing. Overall, these features make it an effective on-site diagnostic tool, allowing the potato industry to manage early diseases more efficiently.},
}

*CRISPR-Cas Systems
*Gold/chemistry
Solanum tuberosum/microbiology
*Platinum/chemistry
*Alternaria/genetics/isolation & purification
*Metal Nanoparticles/chemistry
*Plant Diseases/microbiology
Biosensing Techniques/methods
Limit of Detection
*DNA, Fungal/analysis/genetics

@article {pmid41730290,
year = {2026},
author = {Chen, L and Murillo-de-Ozores, AR and Park, E and Ou, SM and Knepper, MA},
title = {GFP reporter system reveals cell-to-cell variability in aquaporin-2 expression.},
journal = {American journal of physiology. Cell physiology},
volume = {330},
number = {4},
pages = {C782-C793},
doi = {10.1152/ajpcell.00936.2025},
pmid = {41730290},
issn = {1522-1563},
support = {ZIA-HL001285//HHS | NIH | NHLBI | Division of Intramural Research (DIR)/ ; ZIA-HL006129//HHS | NIH | NHLBI | Division of Intramural Research (DIR)/ ; },
mesh = {*Aquaporin 2/genetics/metabolism ; *Green Fluorescent Proteins/genetics/metabolism ; Animals ; Genes, Reporter ; Cell Line ; Mice ; *Kidney Tubules, Collecting/metabolism/drug effects/cytology ; Deamino Arginine Vasopressin/pharmacology ; CRISPR-Cas Systems ; Gene Expression Regulation ; },
abstract = {Vasopressin regulates transcription of the aquaporin-2 gene (Aqp2) in collecting duct principal cells. To investigate regulatory mechanisms in Aqp2 gene transcription, we engineered an Aqp2 reporter cell line using CRISPR/Cas9 to insert a green fluorescent protein (GFP) cassette at the endogenous Aqp2 gene locus in mpkCCD cells. In the absence of dDAVP (1-desamino-8-D-arginine-vasopressin), a vasopressin analog, these cells exhibited low or undetectable GFP and Aqp2 expression in all cells. dDAVP stimulation (1 nM dDAVP for 48 h) markedly increased both GFP and Aqp2 expression together with reversal upon dDAVP removal. These observations demonstrate that GFP faithfully tracks Aqp2 expression. Interestingly, fewer than 50% of cells express GFP and Aqp2 after dDAVP or forskolin, indicating significant variability even though they were clonally derived. We flow-sorted the GFP[-] cells (Aqp2[-]) and GFP[+] cells (Aqp2[+]), regrew them, and restimulated them separately with dDAVP. Cells originating from GFP[-] cells gave rise to both GFP[-] cells and GFP[+] cells, and GFP[+] cells similarly regenerated both GFP[-] and GFP[+] populations in the same proportion. Flow cytometry analysis of the DNA content showed variability in cell cycle phases, with most GFP[+] cells in G0/G1, and most GFP[-] cells in G2/S. RNA-seq analysis of the GFP[-] and GFP[+] cells revealed increased abundance of cell cycle-related transcripts in the GFP[-] cells. We conclude that: 1) heterogeneity in Aqp2 expression is related to cell cycle state and 2) the newly generated reporter cell line will likely serve as a useful tool to study Aqp2 transcriptional regulation.NEW & NOTEWORTHY To investigate regulatory mechanisms in Aqp2 gene transcription, we engineered an Aqp2 reporter cell line using CRISPR/Cas9 to insert a green fluorescent protein (GFP) cassette at the endogenous Aqp2 gene locus in mpkCCD cells. We demonstrate that the GFP reporter accurately and dynamically tracks the expression and regulation of endogenous Aqp2. We reveal that Aqp2 heterogeneity in mpkCCD cells is at least partly driven by differences in cell cycle phase.},
}

*Aquaporin 2/genetics/metabolism
*Green Fluorescent Proteins/genetics/metabolism
Animals
Genes, Reporter
Cell Line
Mice
*Kidney Tubules, Collecting/metabolism/drug effects/cytology
Deamino Arginine Vasopressin/pharmacology
CRISPR-Cas Systems
Gene Expression Regulation

@article {pmid41774834,
year = {2026},
author = {Chen, Z and Lin, H and Yoon, C and Huang, H and Kim, Y and Meng, C and Jang, H and Xie, Z and Li, L and Liu, Y and Kim, JS and Zhang, H},
title = {Bismuthene-Based Nanoplatform for Synergistic Thermogenetic CRISPR and Photothermal Cancer Therapy.},
journal = {Nano letters},
volume = {26},
number = {10},
pages = {3407-3416},
doi = {10.1021/acs.nanolett.5c06006},
pmid = {41774834},
issn = {1530-6992},
mesh = {Humans ; *Photothermal Therapy/methods ; *CRISPR-Cas Systems ; Animals ; Female ; Mice ; Cell Line, Tumor ; *Triple Negative Breast Neoplasms/therapy/genetics/pathology ; *Nanostructures/chemistry ; *Hyperthermia, Induced/methods ; },
abstract = {Overcoming tumor thermotolerance within clinically safe temperature ranges remains a central limitation of photothermal therapy (PTT). Here we report a closed-loop therapeutic nanoplatform that integrates topologically enhanced photothermal conversion with thermally gated CRISPR/Cas9 regulation. Rationally engineered hexagonal bismuthene nanodiscs exhibit strong near-infrared responsiveness, enabling mild hyperthermia (∼45 °C) that activates a heat-sensitive CRISPR switch targeting CDK7. The resulting disruption of the CDK7-HSP70 stress axis lowers the thermal resistance threshold and reprograms tumor adaptation, thereby amplifying photothermal efficacy and promoting immunogenic cell death. In triple-negative breast cancer models, this gene-thermal feedback achieves >93% tumor inhibition with minimal systemic toxicity. This work establishes a genetically programmable, thermogenetic nanomaterial paradigm that links material design with gene logic for next-generation precision cancer therapy.},
}

Humans
*Photothermal Therapy/methods
*CRISPR-Cas Systems
Animals
Female
Mice
Cell Line, Tumor
*Triple Negative Breast Neoplasms/therapy/genetics/pathology
*Nanostructures/chemistry
*Hyperthermia, Induced/methods

@article {pmid41790502,
year = {2026},
author = {Guo, S and Liang, Z and Liao, S and Chen, H and Xue, C and Yue, Z and Jiang, Y and Fang, X},
title = {CRISPR-Enhanced β-Carotene Biosynthesis: Droplet Microfluidics-Driven Pathway Optimization and High-Yield Terpenoid Strain Screening.},
journal = {Journal of agricultural and food chemistry},
volume = {74},
number = {10},
pages = {8497-8507},
doi = {10.1021/acs.jafc.5c10513},
pmid = {41790502},
issn = {1520-5118},
mesh = {*beta Carotene/biosynthesis ; *Terpenes/metabolism ; Metabolic Engineering/methods ; *Microfluidics/methods/instrumentation ; *Saccharomyces cerevisiae/metabolism/genetics ; Biosynthetic Pathways ; CRISPR-Cas Systems ; Mevalonic Acid/metabolism ; },
abstract = {Terpenoids are highly abundant and valuable natural products, yet high-throughput screening platforms for microbial producers remain scarce. To address this, we developed a microfluidic screening platform leveraging the intrinsic fluorescence of β-carotene. Using an engineered yeast chassis (YsL4) with a reinforced mevalonate pathway, we optimized cultivation conditions and achieved perfect (100%) phenotypic specificity in a 1:1 coencapsulation assay between producer (YsL4) and nonproducer (Ys011) strains. Screening a genome-wide CRISPRa/i library via multiround fluorescence-activated droplet sorting identified 15 target genes. Validation confirmed that 10 targets (5 for overexpression, 10 for knockout) significantly enhanced β-carotene production. A key mutant, YL19 (ΔHMO1), achieved a titer of 33.71 mg/L in shake-flasks─a 149.15% increase over the parent strain─with a concurrent lycopene reduction indicating redirected carbon flux toward β-carotene biosynthesis. This integrated platform enhances screening efficiency and provides a new paradigm for identifying critical terpenoid biosynthetic targets.},
}

*beta Carotene/biosynthesis
*Terpenes/metabolism
Metabolic Engineering/methods
*Microfluidics/methods/instrumentation
*Saccharomyces cerevisiae/metabolism/genetics
Biosynthetic Pathways
CRISPR-Cas Systems
Mevalonic Acid/metabolism

@article {pmid41827871,
year = {2026},
author = {Zheng, J and Wu, M and Wang, X and Zuo, Z and Zhou, C and Zuo, E and Lu, Y},
title = {Prime Editing Exhibits Limited Genome-Wide Off-Target Effects in Cellular and Embryonic Gene Editing.},
journal = {Cells},
volume = {15},
number = {5},
pages = {},
pmid = {41827871},
issn = {2073-4409},
support = {2023ZD0405302 and 2023ZD04074//the Biological Breeding-Major Projects/ ; 2021YFD1300100 and 2024YFC3406001//the National Key Research and Development Program of China/ ; 32371549 and W2533083//the National Natural Science Foundation of China/ ; },
mesh = {*Gene Editing/methods ; Animals ; CRISPR-Cas Systems/genetics ; Humans ; RNA, Guide, CRISPR-Cas Systems/genetics/metabolism ; *Embryo, Mammalian/metabolism ; Mice ; *Genome ; Polymorphism, Single Nucleotide/genetics ; },
abstract = {Prime editing (PE) is a precise genome-editing technology that avoids double-strand breaks, holding great promise for clinical and agricultural applications. However, its genome-wide off-target effects are not fully understood, raising safety concerns. Here, we systematically compared the safety profiles of four prime editor variants (PE2max, PE3max, PE4max, and PE5max) using PEM-seq and RNA-seq. We further applied an ultra-sensitive method, Genome-wide Off-target analysis by Two-cell embryo Injection (GOTI), to assess PE5max. Our results show that PE5max did not produce detectable sgRNA-dependent off-target single-nucleotide variants (SNVs) in the GOTI assay and induced only limited large deletions and chromosomal translocations. Collectively, this side-by-side benchmarking under matched conditions demonstrates that PE5max achieves an improved specificity profile, with no detectable increase in genome-wide off-target SNVs, advancing its potential for safer therapeutic use.},
}

*Gene Editing/methods
Animals
CRISPR-Cas Systems/genetics
Humans
RNA, Guide, CRISPR-Cas Systems/genetics/metabolism
*Embryo, Mammalian/metabolism
Mice
*Genome
Polymorphism, Single Nucleotide/genetics

@article {pmid41827889,
year = {2026},
author = {Siles, L and Ruiz-Nogales, S and Méndez-Vendrell, P and Pomares, E},
title = {Precise CRISPR/Cas9 and Cas12 Correction Using Lipoplexes in Retinal Models Derived from Patients with Inherited Retinal Dystrophies.},
journal = {Cells},
volume = {15},
number = {5},
pages = {},
pmid = {41827889},
issn = {2073-4409},
support = {Fi-201401//Fundació de Recerca de l'Institut de Microcirurgia Ocular/ ; //IMO Grupo Miranza/ ; },
mesh = {Humans ; *CRISPR-Cas Systems/genetics ; *Gene Editing/methods ; *Retinal Dystrophies/genetics/pathology ; *Retina/pathology/metabolism ; Retinal Pigment Epithelium/metabolism/pathology ; Induced Pluripotent Stem Cells/metabolism ; Bestrophins/genetics ; Organoids/metabolism ; *Models, Biological ; },
abstract = {Gene editing, particularly CRISPR/Cas technology, represents a promising approach for the treatment of rare genetic diseases, including inherited retinal dystrophies, for which effective therapies are largely unavailable. Despite extensive research investigating gene editing across a wide range of cell types, transient delivery of CRISPR/Cas components and efficient homology-directed repair (HDR) in differentiated cells remain challenging. In this study, we employed hiPSCs derived from patients with Stargardt disease or Best disease, carrying pathogenic variants in ABCA4 or BEST1, respectively, to explore gene editing in human models. CRISPR/Cas9 and Cas12 nucleases were delivered into hiPS-derived retinal pigment epithelium (RPE) and retinal organoids using lipoplexes and compared with electroporation. We evaluated transfection efficiency, sgRNA-mediated DNA cleavage, and HDR-based correction. Precise repair of the pathogenic BEST1 variant was successfully achieved in hiPS-derived RPE cells using both nucleases, with Cas12 yielding the highest efficiency, exceeding 10% of HDR correction. Edited RPE cells preserved normal morphology and expressed specific maturity markers. In contrast, retinal organoids exhibited moderate transfection efficiency but showed no detectable CRISPR/Cas-induced DNA cleavage, highlighting the need for further optimization of gene editing in more complex cellular tissues. This study demonstrates, for the first time, precise correction of a single-nucleotide mutation in patient-derived RPE using CRISPR/Cas9 and Cas12 delivered using lipoplexes. These findings underscore the therapeutic potential of CRISPR/Cas-based strategies for inherited retinal dystrophies and provide a proof of concept for future clinical approximations.},
}

Humans
*CRISPR-Cas Systems/genetics
*Gene Editing/methods
*Retinal Dystrophies/genetics/pathology
*Retina/pathology/metabolism
Retinal Pigment Epithelium/metabolism/pathology
Induced Pluripotent Stem Cells/metabolism
Bestrophins/genetics
Organoids/metabolism
*Models, Biological

@article {pmid41828338,
year = {2026},
author = {Xu, M and Ma, Y and Liu, M and Chen, Y and Duan, Z and Shen, Z and Han, Y},
title = {Bmp16 Regulates Arterial Valve Morphogenesis Through Modulation of Notch Signaling in Zebrafish.},
journal = {International journal of molecular sciences},
volume = {27},
number = {5},
pages = {},
pmid = {41828338},
issn = {1422-0067},
mesh = {Animals ; *Zebrafish/embryology/genetics/metabolism ; *Signal Transduction ; *Heart Valves/metabolism/embryology ; *Zebrafish Proteins/metabolism/genetics ; *Receptors, Notch/metabolism/genetics ; *Morphogenesis/genetics ; *Bone Morphogenetic Proteins/metabolism/genetics ; Gene Expression Regulation, Developmental ; CRISPR-Cas Systems ; },
abstract = {Congenital valve defects account for a substantial proportion of cardiovascular malformations, yet the molecular mechanisms orchestrating cardiac valve development remain incompletely elucidated. While Bone morphogenetic protein (BMP) signaling is essential for valvulogenesis, the specific contributions of individual BMP ligands, particularly the teleost-specific bmp16, have not been characterized. Using the CRISPR/Cas9 system, we generated a bmp16 null knockout and delineated critical roles of this ligand in valvular morphogenesis. bmp16 knockout embryos display a significant reduction in Sox9-positive valvular cells and exhibit severely dysplastic arterial valves, characterized by increased interleaflet distance, thickened leaflets, and shortened leaflet lengths. These morphological abnormalities correlate with impaired valve function, culminating in progressive blood regurgitation, ventricular dilation, and pericardial edema. Mechanistically, loss of bmp16 or pharmacological inhibition of BMP signaling significantly downregulates notch1b expression in developing valves, while pharmacological activation of Notch signaling rescues the regurgitation phenotype in bmp16 mutants. Collectively, our findings establish bmp16 as a novel regulator of valve development and uncover a functional BMP-Notch signaling axis required for vertebrate valvulogenesis, providing new insights into the molecular mechanisms that govern cardiac valve formation and pathogenesis.},
}

Animals
*Zebrafish/embryology/genetics/metabolism
*Signal Transduction
*Heart Valves/metabolism/embryology
*Zebrafish Proteins/metabolism/genetics
*Receptors, Notch/metabolism/genetics
*Morphogenesis/genetics
*Bone Morphogenetic Proteins/metabolism/genetics
Gene Expression Regulation, Developmental
CRISPR-Cas Systems

@article {pmid41828461,
year = {2026},
author = {Ferrero, M and Acquadro, A and Moglia, A},
title = {From Lab to Field: CRISPRing Major Cultivated Solanaceae for Crop Improvement.},
journal = {International journal of molecular sciences},
volume = {27},
number = {5},
pages = {},
pmid = {41828461},
issn = {1422-0067},
mesh = {*Gene Editing/methods ; *CRISPR-Cas Systems ; *Crops, Agricultural/genetics/growth & development ; *Solanaceae/genetics ; Plants, Genetically Modified/genetics ; Genome, Plant ; Plant Breeding/methods ; },
abstract = {The Solanaceae family includes some of the most economically and agronomically important crops, such as tomato, potato, pepper and eggplant. Recently, CRISPR/Cas-based genome editing has emerged as a powerful tool for functional genomics and crop improvement, enabling precise and efficient genetic modifications. This review provides an overview of CRISPR/Cas-mediated genome editing technologies and their applications in the major cultivated Solanaceae crops. The use of CRISPR/Cas9 systems for targeted gene knockout and knock-in approaches is described, together with advances in precision editing strategies such as base editing and prime editing, which allow precise nucleotide substitutions and small sequence changes. The expanding CRISPR toolbox is further explored through alternative Cas proteins, such as Cas12a and Cas13 with distinct targeting features and potential applications. Emerging delivery strategies, including ribonucleoprotein-mediated editing in protoplasts, virus-induced gene editing (VIGE), de novo induction of meristems and genome editing by grafting, represent promising approaches to generate transgene-free edited plants. In addition, the current status of field trials involving genome-edited Solanaceae crops in Europe is outlined, considering the regulatory landscape and legislative requirements for their release in the environment. Despite regulatory constraints, some genome-edited crops have reached the market, highlighting their potential to contribute to sustainable agriculture and crop improvement.},
}

*Gene Editing/methods
*CRISPR-Cas Systems
*Crops, Agricultural/genetics/growth & development
*Solanaceae/genetics
Plants, Genetically Modified/genetics
Genome, Plant
Plant Breeding/methods

@article {pmid41828635,
year = {2026},
author = {Dagdelen, B and Arikoglu, H and Erkoc-Kaya, D and Bozkurt, B},
title = {Precise CRISPR-Mediated Editing of the TGFBI R555W Mutation in Patient-Derived Peripheral Blood Mononuclear Cells.},
journal = {International journal of molecular sciences},
volume = {27},
number = {5},
pages = {},
pmid = {41828635},
issn = {1422-0067},
support = {19102037//Selçuk University/ ; },
mesh = {Humans ; *Leukocytes, Mononuclear/metabolism ; *Gene Editing/methods ; *CRISPR-Cas Systems ; *Corneal Dystrophies, Hereditary/genetics ; *Mutation ; *Transforming Growth Factor beta/genetics ; *Extracellular Matrix Proteins/genetics ; RNA, Guide, CRISPR-Cas Systems/genetics ; },
abstract = {Over 70 mutations in the transforming growth factor beta-induced (TGFBI) gene are associated with corneal dystrophies that impair vision. The R555W hotspot mutation is a major cause of granular corneal dystrophy type 1 (GCD1). Here, we evaluated the technical feasibility of CRISPR/Cas9-mediated editing of the R555W mutation in peripheral blood mononuclear cells (PBMCs) obtained from a patient with GCD1. Three single guide RNAs (sgRNA1-3) and matched single-stranded oligodeoxynucleotide donors (ssODN1-3) were designed and co-transfected into PBMCs. Transfected cells were enriched by flow cytometric sorting, with GFP-positive cells representing approximately 2-4% of the total electroporated population. Editing outcomes were initially screened using high-resolution melting (HRM) analysis, and the sgRNA3-ssODN3 combination identified as the most promising candidate was subsequently validated by next-generation sequencing (NGS). Sequencing revealed a homology-directed repair efficiency of 98.2% among GFP-positive sorted cells, demonstrating efficient and precise genome editing within the enriched population. Because PBMCs are not disease-relevant corneal epithelial cells and only genomic endpoints were assessed, the clinical applicability of this study is limited and the work should be considered a technical proof-of-concept. This framework supports optimization of CRISPR-based strategies prior to studies in biologically relevant corneal models.},
}

Humans
*Leukocytes, Mononuclear/metabolism
*Gene Editing/methods
*CRISPR-Cas Systems
*Corneal Dystrophies, Hereditary/genetics
*Mutation
*Transforming Growth Factor beta/genetics
*Extracellular Matrix Proteins/genetics
RNA, Guide, CRISPR-Cas Systems/genetics

@article {pmid41828895,
year = {2026},
author = {Gu, X and Zhou, Y},
title = {Unlocking the Potential of Macroalgae: Innovative Pretreatment Strategies for Efficient Biorefinery.},
journal = {Molecules (Basel, Switzerland)},
volume = {31},
number = {5},
pages = {},
pmid = {41828895},
issn = {1420-3049},
support = {No. 31900088//the National Natural Science Foundation of China/ ; No. ZR2025MS493//the Natural Science Foundation of Shandong Province of China/ ; },
mesh = {*Seaweed/chemistry/metabolism/genetics ; Biomass ; *Biofuels ; Fermentation ; Cell Wall/chemistry/metabolism ; Gene Editing ; Biotechnology/methods ; CRISPR-Cas Systems ; },
abstract = {Macroalgae represent a promising third-generation feedstock for biorefinery due to their high biomass productivity and non-reliance on arable land. However, their complex cell wall structure poses a significant barrier to efficient bioconversion. This review integrates current pretreatment methods, including physical, chemical, biological, and combined approaches, with a focus on their mechanisms, effectiveness, and limitations. Furthermore, it explores the conversion of pretreated macroalgal biomass into bioenergy and biochemicals, such as bioethanol, organic acid and polyhydroxyalkanoate, via microbial fermentation. The review also examines the application of genetic editing tools (e.g., CRISPR-Cas systems) for the targeted modification of macroalgae to improve their inherent characteristics for biorefinery, such as reducing biomass recalcitrance or increasing the content of target carbohydrates. Finally, future perspectives on technological innovations and integrated industrial chains of macroalgal biorefinery are discussed. This review serves as a systematic reference for deepening the understanding of macroalgal cell wall deconstruction processes and supports the development of efficient and environmentally benign pretreatment strategies to advance macroalgal biorefinery toward industrialization.},
}

*Seaweed/chemistry/metabolism/genetics
Biomass
*Biofuels
Fermentation
Cell Wall/chemistry/metabolism
Gene Editing
Biotechnology/methods
CRISPR-Cas Systems

@article {pmid39901476,
year = {2026},
author = {Clarissa, EM and Karmacharya, M and Choi, H and Kumar, S and Cho, YK},
title = {Nature Inspired Delivery Vehicles for CRISPR-Based Genome Editing.},
journal = {Small (Weinheim an der Bergstrasse, Germany)},
volume = {22},
number = {16},
pages = {e2409353},
doi = {10.1002/smll.202409353},
pmid = {39901476},
issn = {1613-6829},
support = {IBS-R020-D1//Institute for Basic Science/ ; RS-2024-00508821//Korean government/ ; },
mesh = {*Gene Editing/methods ; Humans ; *CRISPR-Cas Systems/genetics ; Nanoparticles/chemistry ; *Clustered Regularly Interspaced Short Palindromic Repeats/genetics ; Animals ; *Gene Transfer Techniques ; Liposomes/chemistry ; Extracellular Vesicles/metabolism ; },
abstract = {The advent of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-based genome editing technologies has opened up groundbreaking possibilities for treating a wide spectrum of genetic disorders and diseases. However, the success of these technologies relies heavily on the development of efficient and safe delivery systems. Among the most promising approaches are natural and synthetic nanocarrier-mediated delivery systems, including viral vectors, extracellular vesicles (EVs), engineered cellular membrane particles, liposomes, and various nanoparticles. These carriers enhance the efficacy of the CRISPR system by providing a unique combination of efficiency, specificity, and reduced immunogenicity. Synthetic carriers such as liposomes and nanoparticles facilitate CRISPR delivery with high reproducibility and customizable functions. Viral vectors, renowned for their high transduction efficiency and broad tropism, serve as powerful vehicles for delivering CRISPR components to various cell types. EVs, as natural carriers of RNA and proteins, offer a stealth mechanism to evade immune detection, allowing for the targeted delivery of genome editors with minimal off-target effects. Engineered cellular membrane particles further improve delivery by simulating the cellular environment, enhancing uptake, and minimizing immune response. This review explores the innovative integration of CRISPR genome editors with various nanocarrier systems, focusing on recent advancements, applications, and future directions in therapeutic genome editing.},
}

*Gene Editing/methods
Humans
*CRISPR-Cas Systems/genetics
Nanoparticles/chemistry
*Clustered Regularly Interspaced Short Palindromic Repeats/genetics
Animals
*Gene Transfer Techniques
Liposomes/chemistry
Extracellular Vesicles/metabolism

@article {pmid40987370,
year = {2025},
author = {Sarwar, A and Shakeel, F and Fatima, T and Amin, R and Rizvi, SNB and Hussain, T and Afzal, A},
title = {DNA nanotechnology for next-generation biosensors: Principles, strategies, and challenges.},
journal = {International journal of biological macromolecules},
volume = {329},
number = {Pt 2},
pages = {147825},
doi = {10.1016/j.ijbiomac.2025.147825},
pmid = {40987370},
issn = {1879-0003},
mesh = {Humans ; Animals ; *Nanotechnology/methods ; *Biosensing Techniques/methods ; *DNA/chemistry ; Fluorescence ; Electrochemical Techniques/methods ; CRISPR-Cas Systems ; },
abstract = {The unique structural and functional properties of nucleic acids make DNA nanotechnology a powerful platform for constructing programmable nanostructures with precise control over size, shape, and biorecognition. This review highlights how DNA-based biosensors achieve unparalleled sensitivity (e.g., attomolar detection via CRISPR systems) and multiplexing capabilities, overcoming limitations of conventional methods like PCR and ELISA. We critically evaluate design strategies, including aptamers, DNAzymes, DNA origami, and CRISPR integration, for detecting diverse analytes (nucleic acids, proteins, pathogens) in clinical, environmental, and food safety applications. While challenges such as nuclease susceptibility and scalability persist, emerging solutions like backbone modifications and microfluidic automation are discussed. By synthesizing recent advances, this review underscores DNA nanotechnology's transformative potential in next-generation diagnostics, offering a roadmap for bridging lab-scale innovations to real-world deployment.},
}

Humans
Animals
*Nanotechnology/methods
*Biosensing Techniques/methods
*DNA/chemistry
Fluorescence
Electrochemical Techniques/methods
CRISPR-Cas Systems

@article {pmid41135835,
year = {2026},
author = {Chiu, CN and Tam, BM and Burns, P and Moritz, OL},
title = {RPE65 knockout Xenopus laevis have a compromised but detectable electroretinogram and altered visual responses, without retinal degeneration or altered melanophore dispersion.},
journal = {Experimental eye research},
volume = {262},
number = {},
pages = {110699},
doi = {10.1016/j.exer.2025.110699},
pmid = {41135835},
issn = {1096-0007},
mesh = {Xenopus laevis/genetics ; Animals ; Gene Knockout Techniques ; *Retinoid Isomerohydrolase/genetics ; Disease Models, Animal ; *Rhodopsin ; Retinal Rod Photoreceptor Cells/pathology ; Electroretinography ; *Photophobia/physiopathology ; *Melanophores/physiology ; Retinal Cone Photoreceptor Cells ; CRISPR-Cas Systems ; Larva ; Behavior, Animal ; },
abstract = {The isomerohydrolase RPE65 is a critical element of the visual cycle, the series of enzymatic reactions by which the chromophore of the visual pigments is regenerated following light exposure. In humans, mutations in the RPE65 gene cause a severe form of blindness called Leber's congenital amaurosis. Studies of Rpe65[-/-] mice have shown dramatic depletion of 11-cis-retinal in the retina, resulting in a slow retinal degeneration. However, a number of studies suggest that RPE65 may not be necessary for the regeneration of photopigment in all photoreceptor types. Using CRISPR/Cas9 technology, we previously generated RPE65 knockout Xenopus laevis in order to test the involvement of rhodopsin chromophore in the cell death mechanisms associated with rhodopsin mutations and rhodopsin quality control. Here we further characterize the effects of RPE65 knockout in these animals, and show their rod photoreceptors have shortened outer segments that lack detectable rhodopsin photopigment. However, there is no progressive degeneration of rods or cones. Via electroretinography we found greatly reduced but significant responses to light under scotopic and photopic conditions. We also found reduced behavioral sensitivity to light, while light-induced melanophore dispersion was unaffected. RPE65 knockout X. laevis may be a useful system for examining RPE65-independent photosensation mechanisms in vertebrates.},
}

Xenopus laevis/genetics
Animals
Gene Knockout Techniques
*Retinoid Isomerohydrolase/genetics
Disease Models, Animal
*Rhodopsin
Retinal Rod Photoreceptor Cells/pathology
Electroretinography
*Photophobia/physiopathology
*Melanophores/physiology
Retinal Cone Photoreceptor Cells
CRISPR-Cas Systems
Larva
Behavior, Animal

@article {pmid41652229,
year = {2026},
author = {Xu, X and Fang, J and Chen, J and Yang, J and Yang, X and Hou, S and Sun, W and Champer, J},
title = {Assessing target genes for homing suppression gene drive.},
journal = {The EMBO journal},
volume = {45},
number = {6},
pages = {2074-2094},
pmid = {41652229},
issn = {1460-2075},
support = {32302455//MOST | National Natural Science Foundation of China (NSFC)/ ; 32270672//MOST | National Natural Science Foundation of China (NSFC)/ ; },
mesh = {Animals ; *Drosophila melanogaster/genetics ; Female ; *Gene Drive Technology/methods ; Male ; *Fertility/genetics ; CRISPR-Cas Systems ; Drosophila Proteins/genetics ; RNA, Guide, CRISPR-Cas Systems/genetics ; },
abstract = {Gene drives are engineered alleles that bias their own inheritance in offspring, enabling the spread of specific traits throughout a population. Targeting female fertility genes in a gene drive can be an efficient strategy for population suppression. In this study, we investigated nine female fertility genes in Drosophila melanogaster using CRISPR-based homing gene drives. Employing a multiplexed gRNA approach to prevent the formation of functional resistance alleles, we aimed to maintain high drive-conversion efficiency with low fitness costs in female drive-carriers. Drive efficiency was assessed in individual crosses and had varied performance across different target genes. Notably, drives targeting the octopamine β2 receptor (oct) and stall (stl) genes exhibited the highest drive-conversion rates and were further tested in cages. A drive targeting stl successfully suppressed a cage population with a high release frequency, though suppression failed in another replicate cage with a lower initial release frequency. Fitness costs in female drive carriers were observed in test cages, impacting the overall efficiency of population suppression. Further tests on the fertility of these lines using individual crosses indicated that some fitness costs were due to maternal deposition of Cas9 combined with new gRNA expression, which would only occur in progeny of drive males when testing split drives with separate Cas9 (when mimicking cages with complete drives) but not for complete drive systems. This could enable success in complete drives with higher maternal Cas9 deposition, even if cage experiments in split drives fail. Overall, our findings identify oct and stl as promising fertility targets and demonstrate both the potential and the constraints of fertility-based suppression drives, providing empirical evidence to guide the design and assessment of more efficient population control strategies.},
}

Animals
*Drosophila melanogaster/genetics
Female
*Gene Drive Technology/methods
Male
*Fertility/genetics
CRISPR-Cas Systems
Drosophila Proteins/genetics
RNA, Guide, CRISPR-Cas Systems/genetics

@article {pmid41654520,
year = {2026},
author = {Zhang, J and Hołubowicz, R and Smidak, R and Hu, Y and Du, SW and Felgner, JH and Palczewska, G and Rodrigues Menezes, C and Risaliti, E and Dong, Z and Ma, X and Shayegan, MH and Chen, PZ and Xing, L and Hołubowicz, M and Li, B and Liu, DR and Felgner, PL and Tochtrop, GP and Palczewski, K},
title = {A combinatorial synthetic strategy for developing genome-editing protein-delivery agents targeting mouse retina.},
journal = {Nature communications},
volume = {17},
number = {1},
pages = {},
pmid = {41654520},
issn = {2041-1723},
support = {T32GM008620; F30EY033642//U.S. Department of Health & Human Services | NIH | National Eye Institute (NEI)/ ; R01EY009339; R01EY030873, P30EY034070//U.S. Department of Health & Human Services | NIH | National Eye Institute (NEI)/ ; CHE-1904530//National Science Foundation (NSF)/ ; P30CA062203//U.S. Department of Health & Human Services | NIH | NCI | Division of Cancer Epidemiology and Genetics, National Cancer Institute (National Cancer Institute Division of Cancer Epidemiology and Genetics)/ ; P30 CA062203/CA/NCI NIH HHS/United States ; },
mesh = {Animals ; *Gene Editing/methods ; Liposomes/chemistry ; Mice ; CRISPR-Cas Systems/genetics ; *Retina/metabolism ; Ribonucleoproteins/genetics/metabolism/administration & dosage ; Disease Models, Animal ; Rosaniline Dyes/chemistry ; Retinal Pigment Epithelium/metabolism ; },
abstract = {CRISPR/Cas9-based gene-editing technologies offer promise for treating inherited retinal diseases (IRDs), however safe and efficient ocular delivery of precision editors remains challenging. To address this challenge, we report a class of Coomassie brilliant blue (CBB)-derived lipidoids that bind and deliver proteins. Subretinal injection of Cre complexed with these lipidoids into mT/mG mice leads to robust recombination in the retinal pigment epithelium and photoreceptors. We employ the CBB-lipidoid platform to deliver adenine base editor (ABE) ribonucleoproteins (RNP). Incorporating CBB lipidoids into liposomes improves delivery efficiency. CBB11 stands out for facilitating precise in vivo ABE-mediated gene editing. Delivery of liposome-CBB11-RNP complexes results in a 120-fold increase in base editing compared to RNP alone and restores the scotopic ERG b-wave response in the rd12 mouse model. These results demonstrate the potential of CBB-augmented, liposome-RNP systems for therapeutic gene editing in the eye, paving the way for single-dose precision medicines to treat IRDs.},
}

Animals
*Gene Editing/methods
Liposomes/chemistry
Mice
CRISPR-Cas Systems/genetics
*Retina/metabolism
Ribonucleoproteins/genetics/metabolism/administration & dosage
Disease Models, Animal
Rosaniline Dyes/chemistry
Retinal Pigment Epithelium/metabolism

@article {pmid41700547,
year = {2026},
author = {Su, T and Zhu, D and Li, X and Qu, Z and Li, F and Zhao, D and Shao, G and Feng, Z and Su, S and Chao, J and Zuo, X and Fan, C and Wang, L},
title = {Accelerated CRISPR/Cas12a-Based Point-of-Care Diagnostics Through Critical Coupling Distance Control.},
journal = {Advanced materials (Deerfield Beach, Fla.)},
volume = {38},
number = {16},
pages = {e23381},
doi = {10.1002/adma.202523381},
pmid = {41700547},
issn = {1521-4095},
support = {62071251//National Natural Science Foundation of China/ ; 62235008//National Natural Science Foundation of China/ ; 62227803//National Natural Science Foundation of China/ ; 62288102//National Natural Science Foundation of China/ ; BK20212012//Natural Science Foundation of Jiangsu Province-Major Project/ ; BZ2022011//"Belt and Road" Innovation Cooperation Project of Jiangsu/ ; SKT2306//Project of State Key Laboratory of Transducer Technology/ ; GZR2022010026//Project of State Key Laboratory of Organic Electronics and Information Displays, Nanjing University of Posts and Telecommunications/ ; BK20243057//Basic Research Program of Jiangsu/ ; BK20253006//Basic Research Program of Jiangsu/ ; },
mesh = {*CRISPR-Cas Systems ; *Point-of-Care Systems ; *Endodeoxyribonucleases/metabolism/genetics ; *Bacterial Proteins/metabolism ; *CRISPR-Associated Proteins/metabolism ; DNA/chemistry ; Humans ; },
abstract = {Prompt pathogen detection in resource-limited settings remains constrained by energy-intensive instrumentation and a shortage of trained personnel. The CRISPR/Cas12a-based diagnostic technology, despite its robustness as a promising tool, is constrained by suboptimal detection speed and sensitivity. Here we designed triblock DNA-mediated spherical nucleic acids (tSNA) that acts as a spatially confined reporter with critical coupling distances between substrates, enabling Cas12a protein to rapidly identify concentrated and stretched single-stranded substrates with size-matching intervals. Precise control of distances on tSNA of varying sizes revealed a direct correlation between trans-cleavage efficiency and coupling distance, indicating that only when the distance exceeds the protein size can it offer an appropriate reaction space. It demonstrates a rapid "scooting" reaction model on tSNA, resulting in a trans-cleavage rate of 10 nm A30-tSNA 12 times faster and a sensitivity that is two orders of magnitude higher than that in bulk solution. Furthermore, tSNA can serve as a novel recognition and colorimetric element in lateral-flow strips, thereby reducing the detection time for pathogen nucleic acids to just 3 min. This "size-matching" model of tSNA offers a new perspective on the regulation of Cas12a enzymatic activity, establishing a versatile platform to advance diagnostic development through ultrafast, CRISPR-powered POC systems.},
}

*CRISPR-Cas Systems
*Point-of-Care Systems
*Endodeoxyribonucleases/metabolism/genetics
*Bacterial Proteins/metabolism
*CRISPR-Associated Proteins/metabolism
DNA/chemistry
Humans

@article {pmid41766140,
year = {2026},
author = {Hartig, AM and Dai, W and Zhang, K and Rottinghaus, AG and Moon, TS and Parker, KM},
title = {Genetic Markers Remain Detectable in Genetically Engineered Microbes Biocontained with a CRISPR Kill Switch.},
journal = {Environmental science & technology},
volume = {60},
number = {10},
pages = {7983-7994},
doi = {10.1021/acs.est.6c00321},
pmid = {41766140},
issn = {1520-5851},
mesh = {Escherichia coli/genetics ; Genetic Markers ; *CRISPR-Cas Systems ; Genetic Engineering ; Clustered Regularly Interspaced Short Palindromic Repeats/genetics ; *Microorganisms, Genetically-Modified/genetics ; },
abstract = {Biocontainment strategies, such as kill switches, have been developed to avoid the unintended proliferation of genetically engineered microbes (GEMs) intended for open-release environmental applications. However, the presence of GEM DNA after successful biocontainment presents new environmental risks and challenges for monitoring. In this study, we investigated whether biocontainment using a CRISPR-Cas9 kill switch, which causes double-strand breaks in target genes essential for GEM growth, could resolve this challenge in a model Escherichia coli GEM. Surprisingly, the escape rates of the GEM as determined by CRISPR-targeted gene abundances were as high as 10[-1.6] to 10[-1.0] in LB media, despite the escape rates measured by colony forming units (cfu) being only 10[-6.2] under the same condition. This discrepancy suggested that the CRISPR-Cas9 kill switch prevents colony growth while still leaving a large fraction of target genes intact for detection by molecular methods. Within 1 h after biocontainment, these target genes remained predominantly inside an intact cell membrane and were resistant to degradation by DNase, though degradation was observed in river water over multiple days. Overall, a detailed understanding of the impact of the biocontainment mechanism on both the GEM and its DNA is needed to minimize unintended environmental risks.},
}

Escherichia coli/genetics
Genetic Markers
*CRISPR-Cas Systems
Genetic Engineering
Clustered Regularly Interspaced Short Palindromic Repeats/genetics
*Microorganisms, Genetically-Modified/genetics

@article {pmid41773916,
year = {2026},
author = {Sui, Z and Chen, B and Zhao, J and Deng, R and Xu, J},
title = {Pronounced Fluorescence Polarization Enhancement Driven by RPA-CRISPR/Cas12a Induced Nucleoprotein Assembly for Salmonella Analysis in Animal-Derived Food Matrices.},
journal = {Analytical chemistry},
volume = {98},
number = {10},
pages = {7822-7831},
doi = {10.1021/acs.analchem.5c08279},
pmid = {41773916},
issn = {1520-6882},
mesh = {*Fluorescence Polarization/methods ; *CRISPR-Cas Systems ; *Salmonella/isolation & purification/genetics ; Animals ; *Nucleoproteins/metabolism/chemistry ; *Nucleic Acid Amplification Techniques/methods ; *Food Microbiology ; *CRISPR-Associated Proteins/metabolism ; *Recombinases/metabolism ; *Endodeoxyribonucleases/metabolism ; Bacterial Proteins/metabolism/genetics ; },
abstract = {Salmonella is one of the most hazardous foodborne pathogens, posing a serious threat to public health and food safety worldwide. Conventional recombinase polymerase amplification (RPA)-CRISPR/Cas12a detection assays predominantly rely on the trans-cleavage of fluorescent reporters; however, such signal-generation modes are inherently susceptible to photobleaching, signal drift, and fluctuation, thereby compromising quantitative accuracy and long-term signal stability in practical pathogen detections. To overcome these limitations, we developed a trans-cleavage-independent fluorescence polarization (FP) sensing platform for the rapid and quantitative detection of Salmonella. Unlike conventional reporter-cleavage-based readouts, the proposed system exploits target-induced nucleoprotein assembly to achieve direct, physical signal amplification. In this design, a FAM-labeled forward primer serves as an intrinsic molecular reporter, while exonuclease I (Exo I) selectively degrades unincorporated primers, effectively suppressing background interference. Upon recognition of Salmonella genomic DNA, RPA produces rigid double-stranded amplicons that restrict fluorophore rotational freedom, and subsequent crRNA-guided Cas12a binding further increases molecular size and hydrodynamic volume, resulting in a stepwise enhancement of FP signals. The assay exhibits excellent linearity over a concentration range of 3 × 10[1]-3 × 10[6] CFU mL[-1], with an ultralow detection limit of 5 CFU mL[-1]. In addition, it demonstrates outstanding photostability, reproducibility, and high specificity against nontarget bacteria. Importantly, reliable Salmonella detection was achieved in complex food matrices, including meat, eggs, and dairy products, with consistently high recoveries and strong tolerance to matrix interference, offering a promising alternative to conventional fluorescence-intensity-based CRISPR diagnostics in complex food systems.},
}

*Fluorescence Polarization/methods
*CRISPR-Cas Systems
*Salmonella/isolation & purification/genetics
Animals
*Nucleoproteins/metabolism/chemistry
*Nucleic Acid Amplification Techniques/methods
*Food Microbiology
*CRISPR-Associated Proteins/metabolism
*Recombinases/metabolism
*Endodeoxyribonucleases/metabolism
Bacterial Proteins/metabolism/genetics

@article {pmid41785043,
year = {2026},
author = {Xu, X and Zhang, R and Hai, G and Wang, Y and Zheng, H and Cui, P and Kou, B and Jin, X and Peng, J},
title = {DNA Logic-Gated CRISPR/Cas13a and PNTs-Hemin Biomimetic Nanozyme for Ratiometric Detection of BRCA1 and circROBO1.},
journal = {Analytical chemistry},
volume = {98},
number = {10},
pages = {7580-7589},
doi = {10.1021/acs.analchem.5c07475},
pmid = {41785043},
issn = {1520-6882},
mesh = {*Hemin/chemistry ; Humans ; *BRCA1 Protein/analysis ; *Biosensing Techniques/methods ; *CRISPR-Cas Systems ; *Biomimetic Materials/chemistry ; *DNA/chemistry ; Gold/chemistry ; Metal Nanoparticles/chemistry ; Logic ; Electrochemical Techniques ; },
abstract = {Although nanozyme-based biosensors show great promise for the early diagnosis of cancer, their application is often limited by poor catalytic activity at neutral pH and susceptibility to matrix interference. This study involved the construction of a biomimetic nanozyme through coordination-driven self-assembly on peptide nanotubes. This rigid framework enables the periodic arrangement of histidine residues to achieve precise axial coordination with the iron center of the hemin molecule, effectively mimicking the active site and catalytic microenvironment of natural horseradish peroxidase. Consequently, PNTs-hemin exhibits peroxidase activity 2.7 times that of free hemin under near-physiological conditions. To ensure detection specificity, an AND logic gate design was integrated, triggering CRISPR/Cas13a-mediated trans-cleavage only when both targets (BRCA1 and circROBO1) are present simultaneously. Furthermore, combining the nanozyme with graphdiyne-supported gold nanoparticles resulted in the formation of a cascade catalytic system that produced a ratiometric electrochemical reading (IFc/Ihemin). This effectively corrects for environmental fluctuations and false positive signals. This study presents an effective strategy that combines the specificity of molecular logic gates with biomimetic catalysis. This opens up new avenues for the precise diagnosis of multiple targets in complex biological samples.},
}

*Hemin/chemistry
Humans
*BRCA1 Protein/analysis
*Biosensing Techniques/methods
*CRISPR-Cas Systems
*Biomimetic Materials/chemistry
*DNA/chemistry
Gold/chemistry
Metal Nanoparticles/chemistry
Logic
Electrochemical Techniques

@article {pmid41787951,
year = {2026},
author = {Chatla, K and Roper, B and Ayalew, L and Ko, P and Lippold, S and Doma, M and Camperi, J},
title = {Assessing mRNA and sgRNA Quality for Cell and Gene Therapy Applications Using Nanopore Direct RNA Sequencing.},
journal = {Analytical chemistry},
volume = {98},
number = {10},
pages = {7452-7461},
doi = {10.1021/acs.analchem.5c06819},
pmid = {41787951},
issn = {1520-6882},
mesh = {*RNA, Messenger/genetics/analysis ; Humans ; *Sequence Analysis, RNA/methods ; *RNA, Guide, CRISPR-Cas Systems/genetics/analysis ; *Genetic Therapy ; *Nanopore Sequencing/methods ; *Nanopores ; },
abstract = {Recent advances in RNA technology have enabled the development of diverse therapeutics spanning vaccines, immunotherapies, and genome-editing platforms. Ensuring clinical efficacy and safety requires precise characterization and control of RNA critical quality attributes (CQAs). Nanopore direct RNA sequencing (NDRS) has emerged as a powerful single-molecule analytical approach capable of simultaneously resolving sequence and structural features consistent with regulatory expectations. In this study, we establish NDRS as a comprehensive, multiattribute analytical platform by integrating novel strategies to assess key CQAs in a single assay. Following workflow optimization, NDRS accurately determined full-length mRNA sequences and evaluated transcript integrity. Notably, we developed the first sequencing-based method for quantifying 5' capping efficiency directly from native RNA molecules. Additionally, we demonstrated, for the first time, full-length sequencing of 100-nucleotide single-guide RNA (sgRNA) molecules by incorporating a 5' RNA oligo adapter, enabling complete identity verification. Quantitative results for poly(A) tail length, integrity, and capping efficiency were consistent with established orthogonal techniques, including chromatography and mass spectrometry. Moreover, functional correlation studies with Cas9 mRNA and sgRNA used in CRISPR-Cas9 editing revealed that increased mRNA degradation led to decreased knockout efficiency. Together, these findings position NDRS as a versatile and unified analytical platform for comprehensive characterization of mRNA and sgRNA, supporting quality assurance, comparability, and control in the development and manufacturing of next-generation RNA therapeutics.},
}

*RNA, Messenger/genetics/analysis
Humans
*Sequence Analysis, RNA/methods
*RNA, Guide, CRISPR-Cas Systems/genetics/analysis
*Genetic Therapy
*Nanopore Sequencing/methods
*Nanopores

@article {pmid41824451,
year = {2026},
author = {Yuan, L and Liu, Q and Xiao, X and Xu, L and Liang, L and Guo, Y and Yao, Y and Wang, H and Feng, Y and Hua, X and Feng, Y},
title = {AlphaFold 3-powered discovery of phage proteins that inhibit bacterial transcription initiation.},
journal = {Cell reports},
volume = {45},
number = {3},
pages = {117082},
doi = {10.1016/j.celrep.2026.117082},
pmid = {41824451},
issn = {2211-1247},
abstract = {Many phages encode proteins that specifically inhibit host RNA polymerase activity, thereby sabotaging and, in some cases, hijacking the host transcription machinery to serve their needs. Traditional methods for identifying new phage proteins that inhibit bacterial transcription are labor intensive and require access to live phages. To overcome these limitations, we develop a highly efficient pipeline for AlphaFold 3-guided discovery of phage proteins that inhibit bacterial transcription initiation. Using this pipeline, three phage proteins are identified and characterized. Structural and biochemical analyses demonstrate that these phage proteins bind to distinct sites on RNA polymerase and inhibit transcription initiation via different mechanisms. This study showcases the power of AlphaFold 3 in discovering novel binders of large protein complexes, and the pipeline developed here could be readily adapted to screen modulators of other large targets, such as the ribosome, proteasome, and CRISPR-Cas systems.},
}

@article {pmid41824934,
year = {2026},
author = {Arif, MA and Mubashir, F and Raffay, A and Talha, M and Zohaib, M and Khan, AR and Naseer, MU and Khan, MH and Idrees, M and Rafique, S and Afzal, S and Amin, I and Shahid, M},
title = {Recent Advances in CRISPR-Cas Systems for Dengue Virus Detection.},
journal = {Critical reviews in eukaryotic gene expression},
volume = {36},
number = {1},
pages = {19-35},
doi = {10.1615/CritRevEukaryotGeneExpr.2025062420},
pmid = {41824934},
issn = {2162-6502},
mesh = {*Dengue Virus/genetics/isolation & purification ; *CRISPR-Cas Systems/genetics ; Humans ; *Dengue/diagnosis/virology/genetics ; RNA, Viral/genetics ; Nucleic Acid Amplification Techniques/methods ; },
abstract = {Dengue virus (DENV) infections persist as a significant global health threat despite decades of surveillance and control efforts. The disease may progress to severe dengue, marked by hemorrhage, plasma leakage, and vital organ impairment, contributing to substantial worldwide morbidity and mortality. The rapidly escalating DENV burden demands rapid and innovative diagnostic approaches that move beyond conventional detection methods. CRISPR-Cas-based technologies have emerged as a revolutionary approach, offering next-generation solutions for DENV diagnostics. This review outlines the recent advances in the use of CRISPR based technologies for robust and more sensitive detection of dengue virus nucleic acids, critically evaluating their advantages over conventional diagnostics, current limitations, and future prospects. The roles of Cas12 and Cas13 in DENV-RNA detection are discussed in detail. Additional key areas mentioned include field-deployable and portable CRISPR-Cas technologies, serotype-specific detection, hybrid and isothermal amplification-based approaches, and a combination of CRISPR with electrochemical sensing techniques and nanotechnology. Collectively, these advances highlight the potential of CRISPR-based diagnostics in evolving future strategies for rapid and effective dengue virus detection and control of infections.},
}

*Dengue Virus/genetics/isolation & purification
*CRISPR-Cas Systems/genetics
Humans
*Dengue/diagnosis/virology/genetics
RNA, Viral/genetics
Nucleic Acid Amplification Techniques/methods

@article {pmid41825677,
year = {2026},
author = {Xu, Z and Haghighat, M and Shafiabady, N and Wu, R and Sang, R and Deng, W},
title = {Nuclear-targeted CRISPR/Cas delivery by using aptamer-modified polymer lipid nanoparticles.},
journal = {Nanomedicine : nanotechnology, biology, and medicine},
volume = {74},
number = {},
pages = {102928},
doi = {10.1016/j.nano.2026.102928},
pmid = {41825677},
issn = {1549-9642},
abstract = {Efficient nuclear delivery of CRISPR/Cas ribonucleoproteins (RNP) remains a significant hurdle for non-viral systems. To address this, we developed a polymer-lipid hybrid nanoparticle functionalized with the AS1411 aptamer, targeting nucleolin to facilitate nucleus-directed delivery. Confocal imaging confirmed the accumulation of these aptamer-modified nanoparticles within the cell nuclei. For precise quantification, we utilized an AI-assisted segmentation approach based on deep convolutional neural networks (CNN) to analyse nanoparticle and DAPI colocalization. We further evaluated in vitro gene knockout efficiency of Cas9/sgRNA by using this nucleus-targeted system. Aptamer-functionalised nanoparticles reduced GFP-positive cells to 30.0%, compared with 40.8% for untargeted nanoparticles. Further evaluation targeting the Lcn2 gene demonstrated higher knockout efficacy and a more potent inhibition of breast cancer cell proliferation. These findings indicate that aptamer-mediated nuclear targeting enhances CRISPR/Cas9 editing efficacy and may offer the potential to advance the performance of non-viral gene therapies.},
}

@article {pmid41759757,
year = {2026},
author = {Schmidt, GE and Weaver, EA and Kim, TH},
title = {CRISPR-based functional analysis of chicken IRF9 reveals distinct modulation of dsRNA stimulated innate immune pathways.},
journal = {Developmental and comparative immunology},
volume = {177},
number = {},
pages = {105577},
doi = {10.1016/j.dci.2026.105577},
pmid = {41759757},
issn = {1879-0089},
mesh = {Animals ; *Chickens/immunology/genetics ; Immunity, Innate/genetics ; *Interferon-Stimulated Gene Factor 3, gamma Subunit/genetics/metabolism ; *RNA, Double-Stranded/immunology ; *Avian Proteins/genetics/metabolism ; Interferon Type I/metabolism ; Signal Transduction ; Clustered Regularly Interspaced Short Palindromic Repeats/genetics ; CRISPR-Cas Systems ; Gene Expression Regulation ; },
abstract = {The chicken immune system is distinct from mammalian models due to its reduced immune gene repertoire, yet it retains the ability to mount a highly effective immune response. In mammals, interferon regulatory factor 9 (IRF9) is a key transcriptional regulator of the type I interferon (IFN) pathway, stimulating the expression of hundreds of antiviral genes. Although IRF9 was previously thought to be absent in chickens, current chicken reference genome annotation (bGalGal1.mat.broiler.GRCg7b) lists a putative chicken IRF9. To investigate the function of this gene in chickens, we utilized a clustered regularly interspace short palindromic repeats (CRISPR) based transcriptional modulation platform to elucidate the role of the putative chicken IRF9 in the innate immune response. We analyzed the transcriptomes of IRF9 repressed cells stimulated with double stranded RNA at 0, 0.5, 1, and 6 h post-stimulation. Gene set enrichment analysis revealed that IRF9 repression resulted in the enrichment of pathways associated with regulating the type I IFN response, including the retinoic acid inducible gene I like (RIG-I like) receptor pathway and the Toll-like receptor pathway. Furthermore, concurrent transcriptional repression of type I IFN modulator IRF7 with transcriptional activation of IRF9 failed to rescue the expression of downstream IFN-stimulated genes. These results suggest chicken IRF9 plays a distinct regulatory role from canonical mammalian IRF9 in the type I IFN response and demonstrate a need for functional evidence-based classification of chicken IRFs.},
}

Animals
*Chickens/immunology/genetics
Immunity, Innate/genetics
*Interferon-Stimulated Gene Factor 3, gamma Subunit/genetics/metabolism
*RNA, Double-Stranded/immunology
*Avian Proteins/genetics/metabolism
Interferon Type I/metabolism
Signal Transduction
Clustered Regularly Interspaced Short Palindromic Repeats/genetics
CRISPR-Cas Systems
Gene Expression Regulation

@article {pmid41820207,
year = {2026},
author = {Porwal, S and Malviya, R and Belagodu Srighar, S and Shareef, J},
title = {Ribosome-targeted and Adjuvant Strategies to Combat Antibiotic Resistance.},
journal = {Infectious disorders drug targets},
volume = {},
number = {},
pages = {},
doi = {10.2174/0118715265396934250805055608},
pmid = {41820207},
issn = {2212-3989},
abstract = {INTRODUCTION: This study examines ribosome-targeted and adjuvant strategies to combat the growing threat of antimicrobial resistance (AMR), with a focus on novel therapeutic approaches, including phage therapy, monoclonal antibodies, CRISPR systems, and AI-driven drug discovery. The objective is to review current challenges and evaluate innovative strategies targeting bacterial ribosomes, a primary site for antibiotic action.

METHODS: A systematic literature review was conducted using databases, such as PubMed, ScienceDirect, Scopus, and Google Scholar.

RESULTS: The study indicates that bacteria evade ribosome-targeting antibiotics through various mechanisms, including porin modification, efflux pumps, ribosomal mutations, and enzymatic degradation. Innovative strategies, including AI-enabled virtual screening, phage-antibiotic synergy, ribosomal protein-targeted monoclonal antibodies and vaccines, and CRISPR-Cas systems, have shown potential in overcoming these mechanisms and restoring antibiotic efficacy.

DISCUSSION: These advanced strategies represent a significant shift from traditional approaches, as they directly target ribosomal functions or resistance genes. While promising, limitations such as phage specificity, challenges in CRISPR delivery, and regulatory concerns must be addressed to ensure clinical translation. The integration of AI with molecular techniques enhances therapeutic precision and development speed.

CONCLUSION: Ribosome-targeted therapies and adjunctive strategies, such as AI, phage therapy, monoclonal antibodies, and CRISPR, offer precise and innovative solutions to overcome antibiotic resistance.},
}

@article {pmid41820622,
year = {2026},
author = {Padureanu, T and Cocoș, R and Matache, IM and Bucur, O},
title = {Gene-sized editing for the therapy of genetic diseases.},
journal = {Functional & integrative genomics},
volume = {26},
number = {1},
pages = {},
pmid = {41820622},
issn = {1438-7948},
abstract = {Programmable genome editing technologies have reshaped the landscape of biomedical sciences, enabling the development of methods with great translational potential. CRISPR-Cas represents one of the most important and widely adopted genome editing tools, although its reliance on double-stranded DNA breaks implies inherent limitations on the precision and safety of genomic insertions. Thus, several research groups have focused on the development of new editing technologies, among which prime editing has emerged as a cutting-edge system. Ongoing advancements in prime editing, including protein engineering, have enhanced its efficiency and expanded its functionality. However, prime editing cannot achieve integration of large DNA sequences larger than 5 kilobases. To overcome this limitation, PASTE and PASSIGE methods were developed as novel genome editing methods that merge precise genome rewriting with efficient recombinase-mediated gene insertion. In this review, we investigate the mechanistic principles of these systems, compare their performance in cellular and animal models, and discuss the ongoing efforts to enhance system components and delivery. We extended our investigation to recent progress supporting their translational potential, assessing efficient delivery methods, genome site specificity, safety, and long-term efficacy, which are crucial for successful in vivo applications.},
}

@article {pmid41821244,
year = {2026},
author = {Elamin Eltom, A and Kareem, AK and Shaaban, Z and Sanaan Jabbar, H and Sharma, MK and Ghazi Al-Shawi, S and AlMohamadi, H and Abbas, Z and S Jabir, M and Ahmed AbdRabou, M},
title = {Progress of Magnetic Particles-Integrated CRISPR/Cas Biosensors for Pathogen Bacteria Detection: Design, Mechanism and Application.},
journal = {Critical reviews in analytical chemistry},
volume = {},
number = {},
pages = {1-19},
doi = {10.1080/10408347.2026.2636658},
pmid = {41821244},
issn = {1547-6510},
abstract = {Ensuring food safety through rapid, sensitive, and point-of-care (POC) detection of microbial pathogens is crucial for protecting public health and minimizing the socio-economic losses associated with foodborne diseases. Despite stringent regulatory measures, foodborne illnesses caused by microbial contamination continue to pose a significant global challenge. In this context, the emergence of CRISPR/Cas systems has significantly improved the performance of biosensors due to their programmability, high specificity, and precise recognition of target RNA and DNA sequences. Following target recognition, Cas proteins exhibit both cis- and trans-cleavage activities, enabling highly sensitive signal amplification. To achieve rapid analysis and low detection limits, recent studies have increasingly focused on integrating CRISPR/Cas system with magnetic particles (MPs). MPs offer key advantages, including superparamagnetism, biocompatibility, and facile surface functionalization, which enhance target enrichment, assay speed, and analytical sensitivity. Accordingly, substantial progress has been made in MP-conjugated CRISPR/Cas biosensors for the detection of diverse foodborne microbial pathogens. This review comprehensively summarizes recent advances in the integration strategies of magnetic particles with CRISPR/Cas-based biosensing platforms for the quantitative detection of microbial pathogens. Particular emphasis is placed on performance metrics, assay design, and the feasibility of these systems for POC applications, highlighting their potential to enhance food safety monitoring.},
}

@article {pmid41823840,
year = {2026},
author = {Loweree-Rivera, FD and Pérez-Álvarez, S and Castillo, AM and Vega Mares, JH and Leyva-Hernández, HA and Sánchez Chávez, E and Escobedo-Bonilla, CM and Uranga-Valencia, LP and Chávez Medina, JA},
title = {Pelecyphora chihuahuensis (Britton & Rose) D. Aquino & Dan. Sánchez: A Review on Its Taxonomy, Ecology and Conservation of an Endemic Mexican Cactus Species with Biotechnological Perspectives.},
journal = {Biology},
volume = {15},
number = {5},
pages = {},
pmid = {41823840},
issn = {2079-7737},
abstract = {The cactus Pelecyphora chihuahuensis is endemic to northern Mexico and represents an interesting subject on the integration of classical taxonomy with modern biotechnological tools to solve conservation issues. Because of its narrow ecological range and high ornamental value, the species is increasingly at risk from degradation of its habitats, climate change, and plant poaching. This review includes current knowledge on its taxonomic status, ecological distribution, and conservation needs, with a focus on biotechnological means to aid its preservation. Aspects such as molecular markers, next-generation sequencing, and previously reported GIS-based species distribution models provide valuable insights into its identity and ecological niche. Biotechnological tools for ex situ conservation include in vitro propagation and cryopreservation. Potential applications of CRISPR-Cas and synthetic biology in preserving rare cacti are also discussed. By uncovering gaps, this review opens a window of opportunity to urgently promote the sustainable management of P. chihuahuensis and related endangered cacti by merging biotechnology with ecology and taxonomy, the results presented here underscore the importance of integrating scientific findings into future research that supports the development and implementation of effective policies that prioritize the conservation and biocultural preservation of arid-land flora, ensuring that both ecological integrity and cultural values are maintained for these unique ecosystems.},
}

@article {pmid41824054,
year = {2026},
author = {Ren, C and Lin, Y and Mohamed, MSM and Liu, C and Liang, Z},
title = {A simplified RUBY reporter-enabled hairy-root system for rapid evaluation of genome editing and gene function in grapevine.},
journal = {Planta},
volume = {263},
number = {4},
pages = {},
pmid = {41824054},
issn = {1432-2048},
support = {YSBR-093//CAS Project for Young Scientists in Basic Research/ ; 32530090//National Natural Science Foundation of China/ ; 2022078//Youth Innovation Promotion Association of the Chinese Academy of Sciences/ ; },
mesh = {*Vitis/genetics/metabolism ; *Gene Editing/methods ; *Plant Roots/genetics/metabolism ; Genes, Reporter ; CRISPR-Cas Systems ; Lignin/metabolism/biosynthesis ; Plants, Genetically Modified ; Plant Proteins/genetics/metabolism ; Glucosyltransferases/genetics ; Gene Expression Regulation, Plant ; },
abstract = {The simplified RUBY represents an efficient reporter for monitoring plant transformation and, when combined with hairy-root system in grapevine, can facilitate the application of novel CRISPR technologies and gene functional study. Monitoring successful transformation events is essential for plant transformation and genome editing. The development and application of the RUBY reporter enable effective selection of transformation events based solely on distinct red pigmentation. Here, we report that a simplified version of RUBY (siRUBY), lacking the glucosyltransferase gene, also functions effectively as a visual selection marker for plant transformation. Furthermore, the siRUBY-assisted hairy-root system was established as a rapid and efficient platform for evaluating activity of adenine base editor (ABE) in grapevine. Targeted A-to-G base editing was achieved using ABE8e, with an average efficiency of approximately 14%. Moreover, this platform is also suitable for functional investigation of genes of interest. Knockout of the MYB4a gene promoted lignin deposition, likely through upregulating key lignin biosynthesis genes while repressing transcription of downstream flavonoid biosynthesis genes. Collectively, these results demonstrate that siRUBY coupled with grapevine hairy roots provides a robust platform for rapid and efficient genome editing in grapevine.},
}

*Vitis/genetics/metabolism
*Gene Editing/methods
*Plant Roots/genetics/metabolism
Genes, Reporter
CRISPR-Cas Systems
Lignin/metabolism/biosynthesis
Plants, Genetically Modified
Plant Proteins/genetics/metabolism
Glucosyltransferases/genetics
Gene Expression Regulation, Plant

@article {pmid40588030,
year = {2026},
author = {Guo, Z and Tang, J and Ling, S and Zheng, C and Bass, C and He, S and Wan, H and Li, J and Ma, K},
title = {Upregulation of a cytochrome P450 gene, CYP6B50, confers multi-insecticide resistance in Spodoptera frugiperda.},
journal = {Journal of advanced research},
volume = {82},
number = {},
pages = {99-111},
doi = {10.1016/j.jare.2025.06.083},
pmid = {40588030},
issn = {2090-1224},
mesh = {Animals ; *Spodoptera/genetics/drug effects ; *Insecticide Resistance/genetics ; *Cytochrome P-450 Enzyme System/genetics/metabolism/chemistry ; Insecticides/pharmacology ; Up-Regulation ; *Insect Proteins/genetics/metabolism ; Drosophila melanogaster/genetics ; Larva/genetics/drug effects ; Molecular Docking Simulation ; Animals, Genetically Modified ; CRISPR-Cas Systems ; Oxazines ; },
abstract = {INTRODUCTION: The fall armyworm, Spodoptera frugiperda, is a globally invasive pest that has developed resistance to numerous insecticides. Cytochrome P450 enzymes have been commonly implicated in insecticide resistance in insect pests, including in S. frugiperda.

OBJECTIVE: Our previous study revealed that the P450 gene CYP6B50 was significantly overexpressed in a multi-insecticide resistant strain of S. frugiperda. However, the functional role of CYP6B50 in conferring multiple resistance remains unclear.

METHODS: Here we used reverse genetic (RNAi, CRISPR/Cas9, and transgenic Drosophila melanogaster) and computational modelling approaches to investigate the role of CYP6B50 in resistance to insecticides.

RESULTS: Functional analysis revealed that CYP6B50 is highly expressed during the larval stage of S. frugiperda, particularly in key tissues associated with detoxification such as the fat body, midgut, hemolymph, and Malpighian tubules. The expression of CYP6B50 was significantly upregulated after exposure to the insecticides indoxacarb, chlorantraniliprole, lambda-cyhalothrin, tetrachlorantraniliprole, and deltamethrin. Functional validation of CYP6B50 confirmed its contribution to resistance against the five tested insecticides. Molecular docking revealed a strong binding affinity of CYP6B50 to these insecticides.

CONCLUSIONS: These results identify CYP6B50 as a key generalist detoxification P450 in S. frugiperda and its role in mediating resistance to multiple insecticides. These findings provide insights into the role of cytochrome P450s in the evolution of insecticide resistance and will inform the development of strategies to manage or overcome resistance in a globally distributed crop pest.},
}

Animals
*Spodoptera/genetics/drug effects
*Insecticide Resistance/genetics
*Cytochrome P-450 Enzyme System/genetics/metabolism/chemistry
Insecticides/pharmacology
Up-Regulation
*Insect Proteins/genetics/metabolism
Drosophila melanogaster/genetics
Larva/genetics/drug effects
Molecular Docking Simulation
Animals, Genetically Modified
CRISPR-Cas Systems
Oxazines

@article {pmid40712808,
year = {2026},
author = {Jin, M and Huang, S and Zhou, S and Shen, W and Cao, J and Song, S and Wei, Y and Kalds, P and Hua, J and Ma, B and Ross, PJ and Wang, X and Chen, Y},
title = {Efficient derivation of stable sheep embryonic stem cells opens a new avenue for agricultural and biomedical application.},
journal = {Journal of advanced research},
volume = {82},
number = {},
pages = {155-169},
doi = {10.1016/j.jare.2025.07.036},
pmid = {40712808},
issn = {2090-1224},
mesh = {Animals ; Sheep ; *Embryonic Stem Cells/cytology/metabolism ; Gene Editing/methods ; Cell Differentiation/genetics ; CRISPR-Cas Systems ; Agriculture/methods ; Cell Culture Techniques/methods ; Blastocyst/cytology ; Pluripotent Stem Cells/cytology ; },
abstract = {INTRODUCTION: Embryonic stem cells (ESCs), capable of generating all adult cell types, hold transformative potential for agriculture and biomedicine. However, stable sheep ESCs derivation remains challenging, limiting their application in further research.

OBJECTIVES: We aimed to establish stable sheep ESCs using a simplified protocol, validate their pluripotency, and demonstrate genome editing utility.

METHODS: Sheep ESCs were derived from blastocyst inner cell masses via an IWR-1-containing culture system (termed TePR). Pluripotency was assessed through long-term culture (>100 passages), trilineage differentiation, transcriptomic analysis (E0-E6 embryos), and cross-species comparisons. ATAC-seq and WGBS mapped chromatin accessibility and methylation patterns, respectively. Genome editing utilized PiggyBac transposition and CRISPR/Cas9.

RESULTS: Sheep ESCs derived under TePR condition (termed TePR-sESCs) exhibited stable morphology and trilineage differentiation. Transcriptomics showed TePR-sESCs resemble 8-cell/morula embryos, consistent with sheep genome activation timing. ATAC-seq revealed accessible chromatin at pluripotency loci (e.g., POU5F1, NANOG). WGBS identified hypomethylation in pluripotency-associated regions. Efficient mCherry integration and MSTN knockouts confirmed editing compatibility.

CONCLUSION: The TePR system enables robust derivation of sheep ESCs with embryonic-like pluripotency. TePR-sESCs' editing proficiency supports applications in livestock trait engineering and biomedical modeling, overcoming a major barrier in ungulate stem cell research.},
}

Animals
Sheep
*Embryonic Stem Cells/cytology/metabolism
Gene Editing/methods
Cell Differentiation/genetics
CRISPR-Cas Systems
Agriculture/methods
Cell Culture Techniques/methods
Blastocyst/cytology
Pluripotent Stem Cells/cytology

@article {pmid41544897,
year = {2026},
author = {Hossain, NS and Tasnim, N and Ferdoush, J and Roy, A and Saha, SC and Saha, A},
title = {Next-generation lung-cancer-on-a-chip: Toward personalized therapy, AI, and CRISPR-driven models.},
journal = {Drug discovery today},
volume = {31},
number = {2},
pages = {104604},
doi = {10.1016/j.drudis.2026.104604},
pmid = {41544897},
issn = {1878-5832},
mesh = {Humans ; *Lung Neoplasms/genetics/drug therapy/pathology/therapy ; *Precision Medicine/methods ; *Artificial Intelligence ; Clustered Regularly Interspaced Short Palindromic Repeats ; Gene Editing ; Animals ; CRISPR-Cas Systems ; *Lab-On-A-Chip Devices ; Tumor Microenvironment ; },
abstract = {Lung-cancer-on-a-chip (LCOC) technologies have advanced rapidly, yet most models evaluate mechanical strain, patient-derived tumors, multi-organ interactions, artificial intelligence (AI) analytics, and clustered regularly interspaced short palindromic repeats (CRISPR) editing in isolation. In this review, we uniquely integrate these emerging components into a unified framework centered on the breathing LCOC. We highlight how embedding patient-derived lung tumor fragments into cyclically stretched microenvironments, then linking them to downstream organ compartments, enables patient-specific mapping of metastatic routes under physiologically relevant mechanics. We further describe how continuous high-resolution imaging from these platforms can feed AI pipelines for automated drug-response prediction and metastatic trajectory simulation, and how on-chip CRISPR editing enables accurate investigation of metastatic drivers within dynamic, strain-modulated microenvironments. By synthesizing these technologies, we outline a next-generation, personalized multi-organ-on-chip architecture capable of predicting individual disease progression without direct patient risk. We also address practical barriers, including tumor fragility under strain, imaging domain shift, and gene-editing delivery challenges, and how to overcome such barriers.},
}

Humans
*Lung Neoplasms/genetics/drug therapy/pathology/therapy
*Precision Medicine/methods
*Artificial Intelligence
Clustered Regularly Interspaced Short Palindromic Repeats
Gene Editing
Animals
CRISPR-Cas Systems
*Lab-On-A-Chip Devices
Tumor Microenvironment

@article {pmid41654238,
year = {2026},
author = {Jo, SJ and Park, SC and Kim, SG},
title = {CRISPR/Cas9-engineered Salmonella phage displaying antimicrobial peptide LL37 for enhanced antibacterial activity.},
journal = {International journal of antimicrobial agents},
volume = {67},
number = {4},
pages = {107734},
doi = {10.1016/j.ijantimicag.2026.107734},
pmid = {41654238},
issn = {1872-7913},
mesh = {Animals ; *Salmonella typhimurium/virology/drug effects ; *Cathelicidins/genetics/pharmacology ; *CRISPR-Cas Systems ; *Salmonella Phages/genetics ; *Anti-Bacterial Agents/pharmacology ; Moths/microbiology ; *Antimicrobial Cationic Peptides/genetics/pharmacology ; Epithelial Cells/microbiology ; Humans ; Larva/microbiology ; Phage Therapy ; *Antimicrobial Peptides/genetics/pharmacology ; Salmonella Infections/therapy ; Gene Editing ; },
abstract = {OBJECTIVES: The increasing prevalence of antibiotic-resistant Salmonella Typhimurium has highlighted the urgent need for alternative therapeutic strategies. This study engineered a lytic S. Typhimurium bacteriophage to present the antimicrobial peptide LL-37 on the virion surface, followed by evaluation of its enhanced antibacterial efficacy.

METHODS: A recombinant lytic bacteriophage displaying LL-37 on its capsid was constructed using CRISPR/Cas9-mediated genome editing. The engineered phage was characterized for structural stability, adsorption kinetics, and lytic activity. Antibacterial efficacy was evaluated through bacterial growth inhibition assays, assessment of phage resistance rates, and host-pathogen interaction studies using intestinal epithelial cells. Intracellular bacterial survival was assessed in vitro, and prophylactic efficacy was further examined in a Galleria mellonella infection model.

RESULTS: The engineered phage exhibited thermal and pH stability comparable to that of the wild-type phage, while demonstrating enhanced adsorption efficiency. In a cell lysis assay, the engineered phage sustained bacterial suppression for 24-72 h, whereas the wild-type phage permitted bacterial regrowth due to the emergence of phage-resistant mutants. The engineered phage significantly reduced bacterial attachment and intracellular survival in an intestinal epithelial cell model. Furthermore, it improved larval survival in a Galleria mellonella infection model in a dose-dependent manner, without inducing significant cytotoxicity.

CONCLUSIONS: LL-37-engineered bacteriophages demonstrated improved antibacterial activity and intracellular infection control against S. Typhimurium. These findings support antimicrobial peptide-armed phages as a promising strategy for enhancing phage therapy and mitigating antibiotic-resistant bacterial infections.},
}

Animals
*Salmonella typhimurium/virology/drug effects
*Cathelicidins/genetics/pharmacology
*CRISPR-Cas Systems
*Salmonella Phages/genetics
*Anti-Bacterial Agents/pharmacology
Moths/microbiology
*Antimicrobial Cationic Peptides/genetics/pharmacology
Epithelial Cells/microbiology
Humans
Larva/microbiology
Phage Therapy
*Antimicrobial Peptides/genetics/pharmacology
Salmonella Infections/therapy
Gene Editing

@article {pmid41672309,
year = {2026},
author = {Terakawa, Y and Koshi, D and Kawauchi, M and Nakazawa, T and An, GH and Honda, Y},
title = {Establishment of a CRISPR/Cas9 protocol by biolistic transformation of the filamentous basidiomycete Pleurotus ostreatus.},
journal = {Journal of microbiological methods},
volume = {243},
number = {},
pages = {107430},
doi = {10.1016/j.mimet.2026.107430},
pmid = {41672309},
issn = {1872-8359},
mesh = {*Pleurotus/genetics ; *CRISPR-Cas Systems ; *Transformation, Genetic ; *Gene Editing/methods ; *Biolistics/methods ; Genetic Vectors ; Genome, Fungal ; },
abstract = {Genome editing was successfully performed by introducing a CRISPR/Cas9 expression vector through biolistic transformation of Pleurotus ostreatus, an oyster mushroom. This method can be applied for genome editing in filamentous basidiomycetes without protoplasting. To our knowledge, this is the first report of genome editing with biolistic transformation in filamentous basidiomycetes.},
}

*Pleurotus/genetics
*CRISPR-Cas Systems
*Transformation, Genetic
*Gene Editing/methods
*Biolistics/methods
Genetic Vectors
Genome, Fungal

@article {pmid41759376,
year = {2026},
author = {Xu, Y and Wu, Y and Zheng, H and Zhao, J and Chen, J and Liu, S and Han, M and Li, F and Zhou, F and Zhang, X and Cao, Y and Zhang, H and Zhang, C},
title = {CRISPR-based metabolic screening identifies PLCE1 as a pivotal regulator of oncolytic viral antitumor immunity via tumor immune microenvironment remodeling.},
journal = {Biochemical and biophysical research communications},
volume = {810},
number = {},
pages = {153505},
doi = {10.1016/j.bbrc.2026.153505},
pmid = {41759376},
issn = {1090-2104},
mesh = {*Tumor Microenvironment/immunology ; Animals ; *Oncolytic Virotherapy/methods ; *Oncolytic Viruses/immunology/genetics ; Mice ; *CRISPR-Cas Systems ; Cell Line, Tumor ; *Colorectal Neoplasms/therapy/immunology/genetics ; Humans ; *Phosphoinositide Phospholipase C/genetics/metabolism/antagonists & inhibitors ; Female ; Mice, Inbred BALB C ; },
abstract = {As a promising cancer immunotherapeutic agent, oncolytic viruses (OV) can specifically kill tumor cells and elicit systemic antitumor immune responses. However, the intrinsic resistance of tumors to oncolytic virotherapy severely limits its therapeutic efficacy. This study identified phospholipase C epsilon 1 (PLCE1) as a key negative regulator of OV antitumor effects via CRISPR-Cas9 metabolic gene screening in MC38 colorectal cancer model. PLCE1 inhibitor U-73122 enhanced OV infection efficiency and immunogenic cell death in vitro. In vivo, U-73122 combined with OV synergistically reduced tumor volume and prolonged survival. The combination therapy has been shown to remodel the tumor immune microenvironment, leading to an increase in CD45[+] immune cells and CD8[+] T cells, including naïve subsets, and a decrease in FOXP3[+] Treg cells. This shift promotes T cell activation by modulating relevant genes and signaling pathways. This study provides a novel target for optimizing OV immunotherapy.},
}

*Tumor Microenvironment/immunology
Animals
*Oncolytic Virotherapy/methods
*Oncolytic Viruses/immunology/genetics
Mice
*CRISPR-Cas Systems
Cell Line, Tumor
*Colorectal Neoplasms/therapy/immunology/genetics
Humans
*Phosphoinositide Phospholipase C/genetics/metabolism/antagonists & inhibitors
Female
Mice, Inbred BALB C

@article {pmid41813887,
year = {2026},
author = {Tou, CJ and Xie, K and Ferreira da Silva, J and Kalailingam, P and Amar-Lewis, E and Rufino-Ramos, D and Sawyer, W and Eller, ML and Starzyk, J and Majumdar, I and Wang, J and Lee, D and Yang, S and Meis, RJ and Dahl, GA and Li, J and Shan, R and Artzi, N and Musolino, PL and Wu, H and Kleinstiver, BP},
title = {Immune evasive DNA donors and recombinases license kilobase-scale writing.},
journal = {Nature},
volume = {},
number = {},
pages = {},
pmid = {41813887},
issn = {1476-4687},
abstract = {Genome-editing technologies that use recombinases to insert kilobase-scale DNA sequences into mammalian genomes canonically require large double-stranded DNA (dsDNA) donors[1,2]. However, dsDNA molecules evoke problematic and toxic innate immune responses, limiting integration efficiencies and generally constraining applicability to ex vivo or immune-deficient contexts. By harnessing mechanisms of integrative prokaryotic viruses and mobile genetic elements, here we demonstrate that recombinases are compatible with immune evasive circular single-stranded DNA molecules optimally bearing a partial-duplex region that reconstitutes the recombinase recognition sequence. This approach, which we term integration through nucleus-synthesized template addition of large lengths (INSTALL), is compatible with diverse protein and RNA-guided recombinases for high-fidelity kilobase-scale human genome writing. INSTALL minimizes innate immune responses in primary human cells and in mice, improving recombinase-mediated integration efficiencies and supporting systemic in vivo non-viral DNA delivery by substantially increasing tolerability and broadening the dosing range compared with lipid nanoparticle-delivered dsDNA molecules. Together, INSTALL overcomes fundamental challenges for DNA delivery and integration methods by synergizing immune-stealth nucleic acids with recombinases to enable kilobase-scale integration strategies without viral vectors.},
}

@article {pmid41816357,
year = {2026},
author = {Pan, Z and Xu, L and Fan, Z and Cao, Y and Ren, F},
title = {CRISPR-based diagnostics for infectious diseases: mechanisms, advancements and clinical transformation prospects.},
journal = {Frontiers in cellular and infection microbiology},
volume = {16},
number = {},
pages = {1769226},
pmid = {41816357},
issn = {2235-2988},
mesh = {Humans ; *CRISPR-Cas Systems ; *Communicable Diseases/diagnosis ; Gene Editing/methods ; *Clustered Regularly Interspaced Short Palindromic Repeats ; *Molecular Diagnostic Techniques/methods ; Sensitivity and Specificity ; },
abstract = {Infectious diseases continue to pose significant global public health challenges, necessitating the development of rapid, sensitive, specific, and field-deployable diagnostic platforms. The discovery of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and CRISPR-associated proteins (Cas) has revolutionized genome editing and concurrently enabled a new generation of molecular diagnostic tools. Leveraging the inherent trans-cleavage activities of Cas enzymes, platforms such as SHERLOCK (Specific High-sensitivity Enzymatic Reporter unLOCKing) and DETECTR (DNA Endonuclease-Targeted CRISPR Trans Reporter) have emerged, combining target recognition precision with reporter systems to achieve ultra-sensitive detection of pathogen-specific nucleic acids. This review systematically examines the mechanistic foundations of CRISPR diagnostics, synthesizes recent advancements in infectious disease applications, evaluates their advantages in sensitivity, specificity, operational simplicity, and multiplexing capacity, and critically analyzes current implementation barriers and future translational pathways.},
}

Humans
*CRISPR-Cas Systems
*Communicable Diseases/diagnosis
Gene Editing/methods
*Clustered Regularly Interspaced Short Palindromic Repeats
*Molecular Diagnostic Techniques/methods
Sensitivity and Specificity

@article {pmid41816914,
year = {2026},
author = {Zhao, H and Zhou, T and Zhang, M and Wang, C and Wang, R and Shu, X and Cheng, F and Xue, Q and Liu, C and Xu, J and Cao, X and Du, J and Wang, L and Liu, H and Li, M},
title = {Associate toxin-antitoxin with CRISPR-Cas to harness (ATTACH) engineered microbes.},
journal = {Nucleic acids research},
volume = {54},
number = {5},
pages = {},
pmid = {41816914},
issn = {1362-4962},
support = {2024YFA0918500//National Key Research and Development Project/ ; XDB0810000//Chinese Academy of Sciences/ ; 32370090//National Natural Science Foundation of China/ ; 32150020//National Natural Science Foundation of China/ ; 32400063//National Natural Science Foundation of China/ ; 32270092//National Natural Science Foundation of China/ ; 32200057//National Natural Science Foundation of China/ ; 32370120//National Natural Science Foundation of China/ ; 2020090//Youth Innovation Promotion Association of CAS/ ; },
mesh = {*CRISPR-Cas Systems ; Animals ; Mice ; *Toxin-Antitoxin Systems/genetics ; Escherichia coli/genetics/metabolism ; RNA, Guide, CRISPR-Cas Systems/genetics ; Plasmids/genetics ; Gene Editing/methods ; Promoter Regions, Genetic ; },
abstract = {Robust biocontainment is essential for the safe use of engineered microbes, but existing strategies suffer from genetic instability and/or laborious construction. Here, we present ATTACH, a kill switch that associates toxin-antitoxin with CRISPR-Cas to harness engineered microbes. Our approach employs a CRISPR-repressed toxin-antitoxin (CreTA) module to make microbes addicted to the type I-F Cas effector proteins, and places both the Cas3 nuclease and the chromosome-targeting guide RNA under inducible promoters, thereby improving the genetic stability and stringency of the CRISPR-based suicidal program. Additionally, we have developed a single-plasmid, antibiotic-independent ATTACH device, which shows robust, stringent containment of a microbial chassis in murine gut, and negligible impacts on culture growth or lycopene production during batch fermentation. Our data highlight the potential of CreTA to stabilize CRISPR-based kill switches, advancing their development into more portable and reliable biocontainment tools for engineered microbes.},
}

*CRISPR-Cas Systems
Animals
Mice
*Toxin-Antitoxin Systems/genetics
Escherichia coli/genetics/metabolism
RNA, Guide, CRISPR-Cas Systems/genetics
Plasmids/genetics
Gene Editing/methods
Promoter Regions, Genetic

@article {pmid41818662,
year = {2026},
author = {Wu, L and Chen, W and Huang, R and Zhou, F},
title = {EXAGO: An Argonaute-Based Primer-Free Exponential Amplification Strategy for Ultrasensitive Zero-Background Detection of Point Mutation.},
journal = {Analytical chemistry},
volume = {},
number = {},
pages = {},
doi = {10.1021/acs.analchem.5c06961},
pmid = {41818662},
issn = {1520-6882},
abstract = {Ultrasensitive gene detection is crucial for precise molecular diagnostics. Conventional nucleic acid amplification methods frequently encounter nonspecific amplification triggered by exogenous primers, which limits their utility in ultrasensitive detection. Programmable gene editing tools, such as CRISPR/Cas and Argonaute (Ago), provide new avenues for developing next-generation detection technologies. Here, we develop an Ago-mediated exponential amplification strategy, EXAGO, for ultrasensitive detection of the epidermal growth factor receptor (EGFR) L858R mutation─a critical biomarker in nonsmall cell lung cancer. Leveraging the flexible programmability and single-base-resolution cleavage activity of Ago, EXAGO can specifically initiate DNA polymerase reactions at the target mutation site. The mechanism of dual-circuit operation allows the system to achieve an efficient exponential amplification under thermal cycling. By circumventing exogenous primers, it also prevents nonspecific amplification caused by primer misidentification at the source. Moreover, wild-type genes are entirely unable to trigger signal amplification, thereby underscoring their superior specificity and the potential for practical application. Experimental results demonstrated that EXAGO achieves femtomolar-level sensitivity and exhibits favorable recovery rates for detecting plasma-diluted samples. Moreover, the use of a thermostable enzyme allows direct compatibility with cell thermal lysis, enabling detection in cell lysates. In summary, EXAGO offers a robust and practical solution for accurate genetic mutation analysis in complex samples and promotes the application of Ago-based tools in molecular diagnostics.},
}

@article {pmid41819069,
year = {2026},
author = {Stephan, TL and Hoodless, PA},
title = {The ups and downs of maturing zonated hepatoctyes.},
journal = {Developmental cell},
volume = {61},
number = {3},
pages = {462-463},
doi = {10.1016/j.devcel.2026.02.009},
pmid = {41819069},
issn = {1878-1551},
mesh = {Animals ; *Hepatocytes/metabolism/cytology ; *Liver/metabolism/embryology/cytology ; Mice ; CRISPR-Cas Systems ; *Cell Differentiation ; },
abstract = {Current in vitro protocols differentiating hepatocytes fail to activate mature metabolic genes, induce zone-specific phenotypes, and suppress fetal liver signatures. In this issue, Taguchi, Magalhães et al.[1] used CRISPR-Cas9 screening in a mouse model of hepatic development to identify Nr1i3 and Nfix as regulators of hepatocyte maturation and zonation.},
}

Animals
*Hepatocytes/metabolism/cytology
*Liver/metabolism/embryology/cytology
Mice
CRISPR-Cas Systems
*Cell Differentiation

@article {pmid41819070,
year = {2026},
author = {Wang, J and Lai, L},
title = {Repurposing Cas13's collateral cleavage activity to mitigate host cell dominance in interspecies chimera formation.},
journal = {Developmental cell},
volume = {61},
number = {3},
pages = {464-465},
doi = {10.1016/j.devcel.2026.02.010},
pmid = {41819070},
issn = {1878-1551},
mesh = {Animals ; Humans ; Mice ; *Chimera ; *CRISPR-Cas Systems/genetics ; *CRISPR-Associated Proteins/metabolism/genetics ; },
abstract = {The insufficient contribution of human cells is a key obstacle to interspecies chimera. In this issue of Developmental Cell, He et al. harnessed the RNA collateral cleavage activity of Cas13 to diminish the competitive advantage of host cells, increasing integration ratio of human cells to 1% in host mice.},
}

Animals
Humans
Mice
*Chimera
*CRISPR-Cas Systems/genetics
*CRISPR-Associated Proteins/metabolism/genetics

@article {pmid41819296,
year = {2026},
author = {Wang, C and Zhu, C and Liu, Q and Yang, L},
title = {Prokaryotic argonaute proteins: From ancient defense mechanisms to modern biosensing applications.},
journal = {Biotechnology advances},
volume = {89},
number = {},
pages = {108869},
doi = {10.1016/j.biotechadv.2026.108869},
pmid = {41819296},
issn = {1873-1899},
abstract = {Prokaryotic Argonaute (pAgo) proteins constitute an evolutionarily ancient nuclease family that is rapidly maturing into a versatile molecular toolkit rivaling CRISPR-Cas. This review synthesizes recent advances in pAgo biology and biotechnology, tracing their phylogeny across thermophilic, mesophilic, and psychrotolerant lineages and highlighting temperature-adapted catalytic signatures that diverge from eukaryotic Agos. In vivo studies reveal pAgo roles in gDNA-guided host defense, transcriptional silencing and recombination, all executed through programmable DNA- or RNA-guided nuclease activity. We detail how guide length, 5' nucleotide identity, divalent cations and accessory factors modulate cleavage efficiency, enabling rational optimization. The review then maps the explosion of pAgo-based biosensing platforms, including selective nucleic acid enrichment platforms, ultrasensitive pathogen detection methods, programmable DNA cloning systems, and high-resolution imaging techniques. Their independence from protospacer adjacent motifs (PAMs), stable DNA guides, multi-turnover kinetics, and broad thermal tolerance position pAgos as ideal complements to CRISPR systems. Finally, we outline current limitations and future directions, including the discovery and engineering of novel variants, elucidation of guide-generation mechanisms, and development of next-generation gene-editing tools, aiming to accelerate translation of these versatile enzymes into practical biotechnological and therapeutic translation.},
}

@article {pmid41813543,
year = {2026},
author = {Qiao, JH and Gao, Q and Wang, XB},
title = {Virus-induced genome editing: toward crop breeding applications.},
journal = {Trends in plant science},
volume = {},
number = {},
pages = {},
doi = {10.1016/j.tplants.2026.01.007},
pmid = {41813543},
issn = {1878-4372},
abstract = {CRISPR-Cas-based genome editing has revolutionized precise genome manipulation in plants, yet its practical application is still constrained by the inefficient delivery of editing reagents across different genotypes. Plant viruses are promising vehicles for delivering genome-editing components, bypassing plant transformation and/or tissue culture. Virus-induced genome editing (VIGE) has provided powerful tools for achieving heritable edits in model plants such as Arabidopsis thaliana and Nicotiana benthamiana. VIGE has now progressed from proof-of-concept to practical applications in agricultural crops. Notably, a recent breakthrough in VIGE in tiller has successfully achieved heritable genome editing in hexaploid wheat. This review outlines the latest advances in VIGE across diverse plant species, highlights its potential for crop improvement, and discusses future research directions.},
}

@article {pmid41811974,
year = {2026},
author = {Baba, S and Oncul, O and Aktas, Z},
title = {CRISPR-Cas Based Plasmid Design for Multidrug Resistant Klebsiella Pneumoniae Isolates.},
journal = {FEMS microbiology letters},
volume = {},
number = {},
pages = {},
doi = {10.1093/femsle/fnag026},
pmid = {41811974},
issn = {1574-6968},
abstract = {Antimicrobial resistance (AMR) is a major global health concern that requires innovative therapeutic strategies. This study aimed to address this challenge by designing Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR-associated protein (CRISPR-Cas)-based plasmid systems for potential genome editing applications in multidrug-resistant (MDR) Klebsiella pneumoniae clinical isolates. Minimum inhibitory concentrations (MICs) of imipenem, meropenem, and ertapenem were determined according to EUCAST guidelines. All isolates (n = 5) were resistant, with MIC ranges of 4-128 μg/mL for imipenem, 8-64 μg/mL for meropenem, and 8-256 μg/mL for ertapenem. Resistance gene analysis revealed blaOXA-48-like and blaCTX-M-15 in all isolates, while blaNDM-1 was detected in one isolate. Two CRISPR-based plasmid systems, CRISPR-Cas9 and CRISPR-assisted cytidine deaminase, were designed. Target genes were amplified by PCR, and guide RNA (gRNA) sequences were designed from selected regions. Apramycin (50 μg/mL) was identified as a suitable selection marker. The pSGKP-AmpR(Pro)-ApmR plasmid was successfully constructed, whereas Cas9 and APOBEC constructs could not be cloned. Overall, this study highlights technical challenges in developing CRISPR-based tools for MDR K. pneumoniae and emphasizes the need for isolate-specific plasmid design and gRNA optimization.},
}

@article {pmid41811515,
year = {2026},
author = {Thiebaut, F and Urquiaga, MC and de Araújo, PM and de Carvalho Vivarini, A and Grativol, C},
title = {Small but big player: the important role of microRNAs in legume crops.},
journal = {Molecular genetics and genomics : MGG},
volume = {301},
number = {1},
pages = {},
pmid = {41811515},
issn = {1617-4623},
mesh = {*MicroRNAs/genetics ; *Crops, Agricultural/genetics/growth & development ; *Fabaceae/genetics/growth & development ; Gene Expression Regulation, Plant ; *RNA, Plant/genetics ; Symbiosis/genetics ; RNA Interference ; Gene Editing ; CRISPR-Cas Systems ; },
abstract = {Legumes are essential components of global cropping systems due to their nutritional value and contribution to sustainable agriculture. Among the regulatory molecules, small RNAs (sRNAs), particularly microRNAs (miRNAs), play crucial roles in plant development and in responses to biotic and abiotic stresses. miRNAs regulate genes involved in diverse developmental processes, including nodule formation, which is fundamental for the nitrogen-fixing symbiosis that characterizes legumes. Functional studies have demonstrated that miRNAs are key modulators of plant defense, contributing to resistance against pathogens and environmental challenges. Moreover, miRNAs also participate in cross-kingdom communication, such as plant-bacteria interactions, influencing symbiotic efficiency. Advances in molecular biology have enabled the manipulation of miRNAs and their targets for crop improvement. Current approaches include the design of artificial miRNAs (amiRNA), modulation of miRNA expression through miRNA-encoded peptides, genome editing of non-coding genes using CRISPR/Cas9, and the application of RNA interference (RNAi) technology. Together, these strategies highlight the potential of miRNA-based tools in plant biotechnology. A deeper understanding of the molecular mechanisms governing miRNA-mediated gene silencing will provide powerful resources for optimizing legume productivity and resilience within sustainable agricultural systems.},
}

*MicroRNAs/genetics
*Crops, Agricultural/genetics/growth & development
*Fabaceae/genetics/growth & development
Gene Expression Regulation, Plant
*RNA, Plant/genetics
Symbiosis/genetics
RNA Interference
Gene Editing
CRISPR-Cas Systems

@article {pmid41811507,
year = {2026},
author = {Thakur, MK and Pandey, S and Singh, SK and Singh, SK and Singh, A},
title = {Hybrid seed production: new paradigms and challenges in the twenty-first century.},
journal = {Planta},
volume = {263},
number = {4},
pages = {},
pmid = {41811507},
issn = {1432-2048},
mesh = {*Seeds/genetics/growth & development ; *Plant Breeding/methods ; *Hybridization, Genetic ; *Crops, Agricultural/genetics ; Hybrid Vigor ; Genomics ; Climate Change ; },
abstract = {Hybrid seed technology future depends on integrating advanced genomics, AI-driven breeding, and enabling policies to sustainably delivery climate-resilient, high-performing hybrids with broad accessibility and equitable benefits worldwide. Hybrid seeds, which exploit heterosis, have driven agricultural productivity gains since the 1920s. Understanding the genetics and molecular biology of hybrid generation led to the development of modern hybrid systems. With time, modern hybrid systems integrated advanced genomic tools such as CRISPR/Cas, marker-assisted selection (MAS), and genomic selection (GS) with established technologies like cytoplasmic male sterility (CMS), restorer-of-fertility (Rf) systems, and chemical hybridizing agents (CHAs) for better hybrid production in a shorter time. Moreover, the integration of emerging approaches leveraging artificial intelligence and machine learning (AI/ML) for trait prediction, multi-parent populations to expand genetic diversity, and epigenetics to engineer climate-resilient hybrids with enhanced stress tolerance is also being explored. However, regulatory hurdles, such as divergent global policies for genetically modified (GM) hybrids, intellectual property (IP) disputes, and restricted germplasm exchange under access-and-benefit-sharing frameworks like the Nagoya Protocol, hinder innovation. Climate change exacerbates both biotic and abiotic stresses, disrupts production zones, and threatens pollinator-dependent crops, while socio-economic barriers limit the adoption of smallholder farming. Case studies of different crops demonstrate the success of hybrids, yet gaps in scalability and accessibility persist. Overall, realizing the potential of hybrid technology hinges on sustained collaboration across scientific, industrial, and policy domains to overcome technical, environmental, and socio-economic constraints. This review examines various techniques for hybrid production that incorporate genomics, future advancements, and synergies between synthetic biology, automation, and predictive breeding, as well as policies that strike a balance between intellectual property protection and germplasm accessibility for hybrid seed production.},
}

*Seeds/genetics/growth & development
*Plant Breeding/methods
*Hybridization, Genetic
*Crops, Agricultural/genetics
Hybrid Vigor
Genomics
Climate Change

@article {pmid41811196,
year = {2026},
author = {Vieira, CSD and Wang, W and Sanchez-Valdez, F and Lim, J and White, BE and Souza, CGS and Tarleton, RL and Paes, MC and Nogueira, NPA},
title = {Glycosomal Phosphoenolpyruvate Carboxykinase CRISPR/Cas9-Deletion and Its Role in Trypanosoma cruzi Metacyclogenesis and Infectivity in Mammalian Host.},
journal = {FASEB journal : official publication of the Federation of American Societies for Experimental Biology},
volume = {40},
number = {6},
pages = {e71672},
pmid = {41811196},
issn = {1530-6860},
support = {00x0ma614//Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)/ ; 88887.311601/2018-00-2629/2018//Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)/ ; E26/010.001706/2019//Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ)/ ; E26/010.100623/2018//Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ)/ ; E26/211.815/2021//Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ)/ ; 402419/2022-7//Conselho Nacional Pesquisa (CNPq) SWE (Sanduíche no Exterior)/ ; },
mesh = {Animals ; *Trypanosoma cruzi/pathogenicity/genetics/enzymology/growth & development ; *CRISPR-Cas Systems ; Mice ; *Chagas Disease/parasitology ; Glucose/metabolism ; *Phosphoenolpyruvate Carboxykinase (ATP)/genetics/metabolism ; *Microbodies/enzymology/metabolism ; Mitochondria/metabolism ; *Protozoan Proteins/genetics/metabolism ; },
abstract = {Trypanosoma cruzi, the causative agent of Chagas disease, possesses glycosomes-unique organelles that house key metabolic enzymes, several of which are promising therapeutic targets. Among them, phosphoenolpyruvate carboxykinase (PEPCK) plays a central role in succinic fermentation, the main pathway for NAD[+] regeneration within the organelle. Using CRISPR/Cas9 editing, the PEPCK gene was disrupted in T. cruzi, producing single-allele knockout epimastigotes (TcPEPCK-sKO) with reduced PEPCK expression and enzyme activity. In a high glucose environment, PEPCK disruption impaired glucose consumption and mitochondrial respiration, particularly oxidative phosphorylation, reducing dependence on mitochondrial ATP production when glucose was supplied. To compensate, pyruvate phosphate dikinase was upregulated, increasing alanine production, possibly to maintain redox balance in glycosomes. Despite this metabolic adaptation, the growth of TcPEPCK-sKO epimastigotes was partially reduced compared with non-deleted parasites. In contrast, under low glucose conditions, PEPCK activity was not critical for mitochondrial bioenergetics, ATP production, or proliferation. Although TcPEPCK-sKO epimastigotes exhibited a minor reduction in growth in high glucose medium, their differentiation (metacyclogenesis) and invasion were severely compromised. However, once inside the host cell, TcPEPCK-sKO amastigotes increased their replication, leading to enhanced trypomastigote production. The same was observed in in vivo infection, where TcPEPCK-sKO infection in IFNγ-deficient mice caused uncontrolled parasitemia and severe pathology, highlighting the critical role of PEPCK in host-pathogen interactions. However, an intact immune system effectively contained TcPEPCK-sKO infection. Taken together, our findings demonstrate that glycosomal PEPCK is crucial for coupling glycolysis to mitochondrial bioenergetics, enabling the parasite differentiation within the insect vector and controlling the infection of mammalian host cells.},
}

Animals
*Trypanosoma cruzi/pathogenicity/genetics/enzymology/growth & development
*CRISPR-Cas Systems
Mice
*Chagas Disease/parasitology
Glucose/metabolism
*Phosphoenolpyruvate Carboxykinase (ATP)/genetics/metabolism
*Microbodies/enzymology/metabolism
Mitochondria/metabolism
*Protozoan Proteins/genetics/metabolism

@article {pmid41811192,
year = {2026},
author = {Shen, Y and Yeung, AT and Ditchfield, P and Korn, E and Clements, R and Chen, X and Wang, B and Huang, Z and Sheen, M and Jarman, PA and Han, C},
title = {A genome-wide MAGIC kit for recombinase-independent mosaic analysis in Drosophila.},
journal = {eLife},
volume = {14},
number = {},
pages = {},
pmid = {41811192},
issn = {2050-084X},
support = {R24OD031953//NIH Office of the Director/ ; },
mesh = {Animals ; *Mosaicism ; *Drosophila/genetics ; *RNA, Guide, CRISPR-Cas Systems/genetics ; *Crossing Over, Genetic ; CRISPR-Cas Systems ; *Genome, Insect ; },
abstract = {Mosaic analysis has been instrumental in advancing developmental and cell biology. Most current mosaic techniques rely on exogenous site-specific recombination sequences that need to be introduced into the genome, limiting their application. Mosaic analysis by gRNA-induced crossing-over (MAGIC) was recently developed in Drosophila to eliminate this requirement by inducing somatic recombination through CRISPR/Cas9-generated DNA double-strand breaks. However, MAGIC has not been widely adopted because gRNA markers, a required component for this technique, are not yet available for most chromosomes. Here, we present a complete, genome-wide gRNA-marker kit that incorporates optimized designs for enhanced clone induction and more effective clone labeling in both positive MAGIC (pMAGIC) and negative MAGIC (nMAGIC). With this kit, we demonstrate clonal analysis in a broad range of Drosophila tissues, including cell types that have been difficult to analyze using recombinase-based systems. Notably, MAGIC enables clonal analysis of pericentromeric genes, deficiency chromosomes and in interspecific hybrid animals, opening new avenues for gene function study, rapid gene discovery, and understanding cellular basis of speciation. This MAGIC kit complements existing systems and makes mosaic analysis accessible to address a wider range of biological questions.},
}

Animals
*Mosaicism
*Drosophila/genetics
*RNA, Guide, CRISPR-Cas Systems/genetics
*Crossing Over, Genetic
CRISPR-Cas Systems
*Genome, Insect

@article {pmid41810550,
year = {2026},
author = {Rahmanian, M and Khoshandam, M and Mousazadeh, M and Yang, P and Soltaninejad, H and Karami Dehkordi, P and Sadeghizadeh, M and Hedayati Goudarzi, MT and Azimi, AH and Sheykhhasan, M},
title = {CRISPR in Medicine: A Systematic Review of Clinical Trials and Therapeutic Applications.},
journal = {Human gene therapy},
volume = {37},
number = {5-6},
pages = {170-182},
doi = {10.1177/10430342251409714},
pmid = {41810550},
issn = {1557-7422},
mesh = {Humans ; *CRISPR-Cas Systems ; *Gene Editing/methods ; *Genetic Therapy/methods ; Clinical Trials as Topic ; Neoplasms/therapy/genetics ; *Hematologic Diseases/therapy/genetics ; Clustered Regularly Interspaced Short Palindromic Repeats ; },
abstract = {Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR associated protein 9 (CRISPR/Cas9) technology has become a revolutionary tool in medicine, offering substantial potential for treating a wide range of diseases, including hematological disorders, cancers, genetic conditions, and ophthalmological diseases. This systematic review evaluates the efficacy, safety, and applicability of CRISPR/Cas9 in clinical trials. A comprehensive search of the PubMed, Scopus, Web of Science, and Cochrane databases was conducted. All studies, up to November 2024, meeting the eligibility criteria assessing the application of CRISPR for the treatment of diseases were included. A quality assessment of the included studies was conducted using the Cochrane risk of bias tool. The Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) 2020 statement for systematic reviews and meta-analyses was followed, and a total of 17 studies were included. This systematic review of CRISPR/Cas9 technology focused on its effectiveness and safety across various diseases. In nonmalignant hematological disorders, CRISPR successfully treated β-thalassemia and sickle cell disease, resulting in high transfusion independence and the elimination of disease crises. In malignant hematological disorders, B-cell acute lymphoblastic leukemia, CRISPR-engineered chimeric antigen receptor T (CAR-T) cells achieved an 83.3% complete remission rate. Furthermore, CRISPR-based CAR-T cells showed promising results in B-cell non-Hodgkin's lymphoma. In oncology, lung cancer and other solid tumors are among the diseases that have been safely engineered using CRISPR gene editing technology. For genetic disorders, CRISPR improved vision in retinal degeneration and reduced symptoms in hereditary angioedema and transthyretin amyloidosis with mild side effects. The results demonstrated CRISPR's potential across a wide range of conditions. In conclusion, the findings underscore the potential role of CRISPR/Cas9 technology across a wide range of diseases. However, challenges remain, including optimizing delivery systems, minimizing off-target effects, addressing immunogenicity concerns, and ethical considerations.},
}

Humans
*CRISPR-Cas Systems
*Gene Editing/methods
*Genetic Therapy/methods
Clinical Trials as Topic
Neoplasms/therapy/genetics
*Hematologic Diseases/therapy/genetics
Clustered Regularly Interspaced Short Palindromic Repeats

@article {pmid41810315,
year = {2026},
author = {Guo, Z and Hu, R and Wang, J and Zhou, M and Zhu, K and Xu, Y},
title = {Research Progress on Point-of-Care Testing Technology for Mycoplasma Pneumonia.},
journal = {International journal of general medicine},
volume = {19},
number = {},
pages = {584824},
pmid = {41810315},
issn = {1178-7074},
abstract = {Mycoplasma pneumoniae (MP) is a significant respiratory pathogen in children, often causing refractory and severe pneumonia. Sensitive, rapid, and portable diagnostic tools are crucial for guiding clinical management. Although traditional methods like culture, ELISA, and PCR are widely used, they suffer from drawbacks such as lengthy turnaround times, complex procedures, or reliance on laboratory equipment. Consequently, Point-of-care testing (POCT) technologies, valued for their speed, portability, and ease of use, have emerged as a key research focus for MP diagnosis. This review systematically summarizes advancements in POCT platforms, covering: (1) immunological methods; (2) molecular biology methods; and (3) biosensor technologies. Their sensitivity, specificity, and clinical performance are comparatively analyzed. Colloidal gold immunochromatography delivers results within 15 minutes but exhibits limited sensitivity. Molecular methods like LAMP and CRISPR-coupled systems achieve single-copy detection limits via isothermal amplification and gene editing, with processing times under 1 hour. Biosensors enable high-sensitivity automated detection through integrated signal amplification and microfluidics. Despite these advantages, POCT development faces challenges including cost-sensitivity tradeoffs, standardization barriers, and sample matrix interference. Future directions encompass multi-modal detection, AI-assisted interpretation, multiplex pathogen screening, and dynamic drug resistance gene monitoring. These innovations will expand POCT device deployment in primary care and home settings. This will ultimately improve effectiveness in controlling respiratory infections.},
}

@article {pmid41656303,
year = {2026},
author = {Lee, CJ and Nam, Y and Rim, YA and Ju, JH},
title = {Harnessing induced pluripotent stem cells and organoids for disease modeling and precision medicine.},
journal = {Stem cell research & therapy},
volume = {17},
number = {1},
pages = {},
pmid = {41656303},
issn = {1757-6512},
support = {HI22C1314//Ministry of Health and Welfare/ ; },
mesh = {Humans ; *Organoids/metabolism/cytology ; *Induced Pluripotent Stem Cells/metabolism/cytology ; *Precision Medicine/methods ; Gene Editing ; CRISPR-Cas Systems ; Animals ; },
abstract = {The convergence of CRISPR genome editing, patient-derived organoids, and induced pluripotent stem cells (iPSCs) has reshaped in vitro disease modeling by enabling mechanistic investigations of human pathophysiology within genetically matched, tissue-relevant systems. Together, these technologies provide a synergistic platform for precise manipulation of disease-associated variants and support the generation of isogenic organoid models that reproduce key phenotypic and functional hallmarks across cancer, neurodegenerative, inflammatory, and monogenic disorders. In this review, we highlight how diverse CRISPR modalities-including knock-out, knock-in, CRISPRa/i, and genome-scale screening-have been applied to dissect gene function, model disease progression, and guide therapeutic development using iPSC- and organoid-based systems. We further discuss the application of these platforms in genotype- and phenotype-driven precision medicine, enabling patient stratification, drug-response prediction, and individualized treatment design. We illustrate these convergent applications with representative case studies spanning mechanistic research and early clinical translation. By combining the scalability of genome engineering with the physiological fidelity of organoids, CRISPR-integrated platforms are redefining the frontiers of experimental medicine. These approaches accelerate the discovery of disease mechanisms and actionable therapeutic targets while establishing individualized clinical strategies for complex human diseases. Collectively, they position CRISPR-enabled organoid systems as a foundational infrastructure that bridges genome editing to individualized therapy and supports next-generation precision medicine.},
}

Humans
*Organoids/metabolism/cytology
*Induced Pluripotent Stem Cells/metabolism/cytology
*Precision Medicine/methods
Gene Editing
CRISPR-Cas Systems
Animals

@article {pmid41810060,
year = {2026},
author = {Shen, Z and Liu, Y and Hao, Y and Bo, Y and Dai, X and Wang, S and Xia, T and Su, X and Liu, H},
title = {Advances in Double-Stranded DNA Targeting Technologies.},
journal = {Exploration (Beijing, China)},
volume = {6},
number = {1},
pages = {20250065},
pmid = {41810060},
issn = {2766-2098},
abstract = {Double-stranded DNA (dsDNA) serves as a fundamental repository of genetic information and plays a pivotal role in the diagnosis and therapeutic management of diseases. However, the inherent stability of the DNA double helix under physiological conditions presents a challenge in accessing internal bases. To address this, various molecular targeting technologies have been developed, offering high specificity while destabilizing the DNA structure. This review provides a comprehensive overview of current dsDNA targeting tools, such as hybridization probes, modified nucleic acid probes, zinc finger proteins (ZFPs), transcription activator-like effector nucleases (TALENs), the CRISPR/Cas system, Argonaute proteins (Agos), and the lambda exonuclease-pDNA system (λ Exo-pDNA), and some cutting-edge molecular tools. It delves into the mechanisms behind these technologies. It highlights their applications in diverse areas, including in vitro detection, in situ imaging, gene editing, and their integration with artificial intelligence (AI)-driven tools. Additionally, the review compares these techniques, discusses future technological opportunities, and identifies challenges in integrating these tools into diagnostic and therapeutic practices. By providing a holistic view of these rapidly evolving technologies, this review aims to fill a gap in the current literature and explore the future potential of dsDNA targeting innovations.},
}

@article {pmid41809894,
year = {2025},
author = {Yang, P and Khoshandam, M and Bhia, I and Raji, S and Soltaninejad, H and Hosseinkhani, S and Sani, M and Hamidieh, AA and Sheykhhasan, M},
title = {Integrating CRISPR/Cas technology with clinical trials: Principles, progress and challenges.},
journal = {Asian journal of pharmaceutical sciences},
volume = {20},
number = {6},
pages = {101068},
pmid = {41809894},
issn = {2221-285X},
abstract = {CRISPR represent a groundbreaking genome-editing technology that has revolutionized genetic modification. This innovative tool offers an unparalleled revolution in the future treatment of genetic disorders, neurological diseases, infectious diseases and cancer. Despite the rapid expansion of CRISPR applications, its clinical use in humans is still relatively limited, with only 69 active clinical trials and 6 completed studies reported so far. This review examined current clinical trials and their processes in addressing various diseases via the CRISPR/Cas system. While earlier literatures have focused mainly on delivery methods and materials for CRISPR/Cas9, our review emphasized innovative targeting conditions and approaches for novel and functional therapeutic designs. In addition, we reviewed recent research to increase the efficiency of CRISPR editing in the management of genetic disorders and cancer, while exploring their future challenges and potential. This review provided a unique perspective on the advancement of CRISPR technology. By addressing these aspects, we aim to contribute to ongoing efforts to improve CRISPR-based therapies and expand their clinical applications, ultimately striving to transform the future of medical treatment.},
}

@article {pmid41808396,
year = {2026},
author = {Zargul, A and Liu, H and Zhang, W and Wang, H and Liu, J and Chen, C},
title = {Advances in Pathogen Detection by Nanosensors: Biorecognition Strategies, Signal Amplification, and Platform Engineering.},
journal = {ACS nano},
volume = {},
number = {},
pages = {},
doi = {10.1021/acsnano.5c22148},
pmid = {41808396},
issn = {1936-086X},
abstract = {The escalating global threat of infectious diseases, compounded by antimicrobial resistance (AMR), calls for improved diagnostic strategies. Conventional pathogen detection techniques─culture, enzyme-linked immunosorbent assay (ELISA), and microscopy─remain hindered by prolonged turnaround times, suboptimal sensitivity for low-abundance analytes, and operational intricacy. Nanosensor technologies have emerged as powerful enablers of rapid, ultrasensitive, and field-deployable diagnostics. This review delineates the convergence of three transformative domains: (1) advanced biorecognition strategies─including monoclonal antibodies, aptamers, bacteriophages, antimicrobial peptides, molecularly imprinted polymers, and lectins─that confer high-fidelity molecular selectivity within complex biological matrices; (2) multimodal signal amplification technologies, encompassing nanomaterial-enhanced mechanisms, enzymatic cascades, and isothermal nucleic acid amplification that drive detection down to the single-cell and femtomolar regimes; and (3) integrated platform engineering, uniting clustered regularly interspaced short palindromic repeats-Cas (CRISPR-Cas) systems, artificial intelligence (AI), and microfluidics to achieve multiplexed, real-time, point-of-care deployment. Advances are critically evaluated through standardized performance metrics─limit of detection, assay time, specificity, and operational simplicity─to reveal both synergistic opportunities and enduring translational bottlenecks. Collectively, these developments define a strategic framework for next-generation nanosensor diagnostics poised to revolutionize infectious disease surveillance and enable precision-guided therapeutic intervention.},
}

@article {pmid41807728,
year = {2026},
author = {Godsil, M and Wei, N and Meeske, AJ},
title = {Conditional activation of Cas13 enforces lysogeny in a native type VI-A CRISPR host.},
journal = {Nature microbiology},
volume = {},
number = {},
pages = {},
pmid = {41807728},
issn = {2058-5276},
support = {R35 GM142460/GM/NIGMS NIH HHS/United States ; FAIN2235762//NSF | BIO | Division of Molecular and Cellular Biosciences (MCB)/ ; R35GM142460//U.S. Department of Health & Human Services | NIH | National Institute of General Medical Sciences (NIGMS)/ ; },
abstract = {CRISPR-Cas (clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins) systems present a barrier to prophage acquisition by restricting invading phages or by inducing autoimmune cleavage of integrated prophage DNA. The RNA-sensing type VI CRISPR nuclease Cas13 mediates non-specific RNA cleavage upon recognition of phage lytic transcripts, but how this system influences the temperate phage life cycle remains unknown. Here we report that the Listeria seeligeri type VI-A CRISPR system restricts the lytic cycle of temperate phages but tolerates prophage acquisition and interferes with prophage induction through a non-abortive mechanism. During attempts at induction, Cas13 activation forces prophage re-integration, thus maintaining lysogeny. We also find that during polylysogenic induction, Cas13 acts specifically, restricting only the targeted phage, in contrast to its behaviour during lytic replication. Our findings show that Cas13 elicits a unique response to each stage of the temperate phage life cycle, enabling type VI CRISPR hosts to acquire potentially beneficial prophages while mitigating lysis.},
}

@article {pmid41807051,
year = {2026},
author = {Gur Dedeoglu, B and Noyan, S and İlhan, KNK},
title = {Non-coding RNAs regulation in breast cancer pathogenesis.},
journal = {Epigenomics},
volume = {},
number = {},
pages = {1-20},
doi = {10.1080/17501911.2026.2642583},
pmid = {41807051},
issn = {1750-192X},
abstract = {Breast cancer represents a molecularly heterogeneous disease shaped by complex genetic, epigenetic, and transcriptional dysregulation. Non-coding RNAs (ncRNAs) including microRNAs (miRNAs), long non-coding RNAs (lncRNAs), and circular RNAs (circRNAs) as well as small nucleolar RNAs (snoRNAs), piwi-interacting RNAs (piRNAs), and small nuclear RNAs (snRNAs), have emerged as key epigenetic regulators that integrate multiple layers of gene control. Through interactions with chromatin-modifying enzymes, RNA-binding proteins, and signaling effectors, ncRNAs modulate transcriptional activity, chromatin accessibility, and post-transcriptional stability of target genes. miRNAs predominantly act as post-transcriptional repressors, whereas lncRNAs and circRNAs exert transcriptional and epigenetic control via scaffolding, miRNA sponging, and chromatin remodeling; some circRNAs even encode functional peptides. Aberrant ncRNA expression contributes to proliferation, metastasis, metabolic reprogramming, immune evasion, and therapeutic resistance, with distinct expression signatures associated with triple-negative, HER2-positive, and hormone receptor - positive breast cancers. Owing to their stability and detectability in plasma and exosomes, ncRNAs hold promise as minimally invasive biomarkers for early detection and disease monitoring. Moreover, therapeutic strategies targeting ncRNAs, such as antisense oligonucleotides, RNA interference, CRISPR/Cas-based editing, and ncRNA-derived vaccines, are advancing toward clinical translation. Collectively, ncRNAs redefine the epigenetic landscape of breast cancer, offering a framework for integrated diagnostic and therapeutic approaches in precision oncology.},
}

@article {pmid41806830,
year = {2026},
author = {Escobar, M and Malik, SA and Srinivasa, MA and Mendez-Sosa, MA and Miller, JM and Lydon, SL and Luong, SN and Mathew, PR and Abouleisa, RRE and Chakravarty, S and Pathan, S and Mohamed, TMA and Ghanta, RK and Hilton, IB},
title = {CRISPR-Cas-based activation of PPARGC1A boosts endogenous mitochondria and enhances cardiac function after myocardial infarction.},
journal = {Molecular therapy : the journal of the American Society of Gene Therapy},
volume = {},
number = {},
pages = {},
doi = {10.1016/j.ymthe.2026.02.027},
pmid = {41806830},
issn = {1525-0024},
support = {R01 HL147921/HL/NHLBI NIH HHS/United States ; R01 HL178610/HL/NHLBI NIH HHS/United States ; R61 HL177472/HL/NHLBI NIH HHS/United States ; },
abstract = {Insufficient energy supply due to impaired mitochondria has emerged as a key pathological factor in the development of heart failure (HF) after myocardial infarction (MI). Unfortunately, no current therapeutic strategies directly augment myocardial energy production. While mitochondrial biogenesis is orchestrated by the activity of multiple genes, activation of PPARGC1A, a key regulator, can increase cellular mitochondria; however, supraphysiological levels of PPARGC1A result in adverse tissue remodeling and heart dysfunction. CRISPR activation (CRISPRa) technologies present a unique opportunity to address these shortcomings, as they enable tunable control over endogenous target gene expression. Here, we demonstrate that transcriptional activation of PPARGC1A using CRISPRa increases cellular mitochondria in human cell types. This effect is mediated through the activation of transcriptional programs driving mitochondrial biogenesis, mitochondrial function, and cellular bioenergetics. These activated transcriptional programs synergize to increase ATP production and reserve capacity in human cardiomyocytes. CRISPRa targeting of PPARGC1A in vivo increases cardiac mitochondria to recover heart ejection fraction in an acute MI model. Furthermore, CRISPRa acts on the adult human heart to increase PPARGC1A protein and cellular mitochondria, elevating mitochondrial function in both normal and HF-diagnosed hearts. These results provide the first proof of concept that endogenous gene activation via CRISPRa can improve heart function after MI.},
}

@article {pmid41713565,
year = {2026},
author = {Shashikala, T and Yogi, D and Akshay, K and Nagesh, SN and Manamohan, M and Venkataravanappa, V and Jha, GK and Ashok, K and Asokan, R},
title = {First report of CRISPR/Cas13a-based rapid detection of groundnut bud necrosis virus without amplification.},
journal = {Methods (San Diego, Calif.)},
volume = {249},
number = {},
pages = {9-22},
doi = {10.1016/j.ymeth.2026.02.010},
pmid = {41713565},
issn = {1095-9130},
mesh = {*CRISPR-Cas Systems/genetics ; *Plant Diseases/virology ; RNA, Viral/genetics/isolation & purification ; *Tospovirus/genetics/isolation & purification ; Vigna/virology ; RNA, Guide, CRISPR-Cas Systems/genetics ; },
abstract = {Globally, the groundnut bud necrosis virus (GBNV) (Bunyaviridae), pose a serious threat to solanaceous and leguminous crops causing serious crop loss. This requires a rapid and sensitive diagnostics for initially identifying at the earliest stage and further to initiate disease management. This study presents the first report of a CRISPR/Cas13a-based diagnostic assay for GBNV detection without amplification. GBNV was maintained in cowpea cv. C152 through mechanical inoculation, further viral RNA was isolated to clone the two target genes viz. nucleocapsid (NP) and movement protein (MP) genes. These genes were ligated to pTZ57R/T vector and sequenced. Similarly, the LshCas13a gene was cloned from pUC19 into pET28a, expressed in E. coli BL21, and purified using Ni-NTA affinity chromatography. Guide RNAs targeting conserved regions of NP and MP genes were synthesized by in vitro transcription and mixed with Cas13a protein to form ribonucleoprotein (RNP) complex. Target RNA, obtained either by in vitro-transcription or crude extract of infected tomato was used to detect GBNV, using a fluorescence-based reporter assay. This method found to be highly sensitive that could detect GBNV at as low as 0.01 ng. From the field perspective, GBNV could be detected from the crude extract of the GBNV infected tomato leaves using an alkaline PEG buffer. Thus CRISPR/Cas13a-based assay provides a rapid, amplification-free, and field-deployable diagnostic platform for GBNV. This lays the groundwork for a field adoptable CRISPR diagnostics for other plant RNA viruses also.},
}

*CRISPR-Cas Systems/genetics
*Plant Diseases/virology
RNA, Viral/genetics/isolation & purification
*Tospovirus/genetics/isolation & purification
Vigna/virology
RNA, Guide, CRISPR-Cas Systems/genetics

@article {pmid41678668,
year = {2026},
author = {Kim, J and Yoon, J and Chen, J and Lee, J and Lee, HJ and Whitworth, K and Redel, B and Prather, RS and Lee, K},
title = {Enhancing the specificity of gene editing outcomes by using Cas9 variants in porcine embryos.},
journal = {Journal of animal science},
volume = {104},
number = {},
pages = {},
doi = {10.1093/jas/skag030},
pmid = {41678668},
issn = {1525-3163},
mesh = {Animals ; *Gene Editing/veterinary/methods ; Swine/embryology/genetics ; *CRISPR-Cas Systems ; *CRISPR-Associated Protein 9/genetics/metabolism ; *Embryo, Mammalian ; Female ; RNA, Guide, CRISPR-Cas Systems/genetics ; },
abstract = {The CRISPR/Cas9 technology has improved the ability to introduce targeted modifications in cells and embryos in diverse species. The use of this technology enables the establishment of genetically modified livestock models to study human diseases or improve food production. However, one of the main concerns with employing this technology is the possibility of introducing unintended genome modifications induced by the Streptococcus pyogenes Cas9 (SpCas9), a commonly used Cas9 protein. Recent advancements in CRISPR/Cas9 technology offer Cas9 variants that are designed to improve gene editing specificity. Here, three high-fidelity SpCas9 variants (eSpCas9, HiFi Cas9, and LZ3 Cas9) were employed to examine their efficacy and specificity in pig embryos. To introduce targeted modifications, mRNA coding for each Cas9 variant was mixed with IGH single guide RNA (sgRNA) and were injected into fertilized pig zygotes. The frequency of on- and off-targeting was calculated by amplifying IGH, AR, and RBFOX1 regions from genomic DNA derived from the injected embryos at the blastocyst stage and sent for Sanger sequencing. The sgRNA targeting IGH locus resulted in a 100% on-target editing rate using SpCas9. However, SpCas9 introduced off-targeting events in AR and RBFOX1 at a high frequency (> 60%) in embryos. Injecting each Cas9 variant at 20 ng/µl could modify the target gene (IGH) at 100% efficiency except for LZ3 Cas9 (59.1%). Importantly, off-target events on AR and RBFOX1 were not detected in any Cas9 variant groups. Gradually reducing the concentration of Cas9 mRNAs lowered the efficacy of on-targeting in all groups; however, the reduction was more dramatic in HiFi Cas9 and LZ3 Cas9 injected embryos. No embryonic toxicity was identified in embryo injected with Cas9 variants and more embryos reached blastocyst stage when injected with either eSpCas9 or HiFiCas9 mRNA. In vivo competency of embryos receiving eSpCas9 was examined by embryo transfer and fetuses recovered from a pregnant sow presented 100% on-target editing efficiency without any detectable off-target events. In summary, among the Cas9 variants examined, eSpCas9 presented the highest specificity with no detectable off-target events and supported the development of gene-edited fetuses. Our findings indicate that the use of Cas9 variants can advance the field of gene editing in livestock models.},
}

Animals
*Gene Editing/veterinary/methods
Swine/embryology/genetics
*CRISPR-Cas Systems
*CRISPR-Associated Protein 9/genetics/metabolism
*Embryo, Mammalian
Female
RNA, Guide, CRISPR-Cas Systems/genetics

@article {pmid41644661,
year = {2026},
author = {Davydova, S and Liu, J and Kandul, NP and Antoshechkin, I and Mann, J and Braswell, WE and Akbari, OS and Meccariello, A},
title = {Generating cisgenic sexing strains in insect pests.},
journal = {Communications biology},
volume = {9},
number = {1},
pages = {},
pmid = {41644661},
issn = {2399-3642},
support = {101059523//EC | Horizon 2020 Framework Programme (EU Framework Programme for Research and Innovation H2020)/ ; AP23PPQS&T00C108//United States Department of Agriculture | Agricultural Research Service (USDA Agricultural Research Service)/ ; },
mesh = {Animals ; Male ; Female ; *Ceratitis capitata/genetics ; CRISPR-Cas Systems ; *Pest Control, Biological/methods ; Gene Editing ; Introns ; },
abstract = {Insect pest population control via sterile insect technique markedly benefits from separation by sex prior to release. To simplify this process, traditional genetics has been deployed to develop genetic sexing strains (GSSs) for several disease vectors and agricultural pests of vast economic significance, although very few are applied in the field due to associated fitness costs and instability. In this study, we generated a method to engineer cisgenic GSS (CGSS) in insects. We use CRISPR/Cas9-mediated homology-directed repair to seamlessly translocate a sex-specific alternatively spliced intron into a dominant phenotypic gene generating a genetically stable strain that enables sex-sorting by eye. To achieve this feat, we use Ceratitis capitata as our model and relied on the sex-specifically spliced intron of its endogenous transformer gene, which we seamlessly inserted a copy into the pupal colouration white pupae gene. This minimal modification resulted in the generation of a homozygous strain we term IMPERIAL that was genetically and phenotypically stable where all female pupae are brown while male pupae are white with overall good fitness. By minimally editing the genome, our novel CGSS approach can be applied to other pests that may aid more efficient and economically suitable pest control.},
}

Animals
Male
Female
*Ceratitis capitata/genetics
CRISPR-Cas Systems
*Pest Control, Biological/methods
Gene Editing
Introns

@article {pmid41454745,
year = {2026},
author = {Mehnath, S},
title = {Engineering stimuli-responsive nanocarriers for CRISPR/Cas9 genome editing: next-generation cancer therapeutics.},
journal = {The Journal of pharmacy and pharmacology},
volume = {78},
number = {3},
pages = {},
doi = {10.1093/jpp/rgaf127},
pmid = {41454745},
issn = {2042-7158},
mesh = {Humans ; *CRISPR-Cas Systems/genetics ; *Gene Editing/methods ; *Neoplasms/genetics/therapy ; Animals ; *Nanoparticles ; Drug Delivery Systems/methods ; Drug Carriers ; Genetic Therapy/methods ; },
abstract = {OBJECTIVES: To highlight recent developments in CRISPR/Cas9 genome-editing strategies for cancer therapy and to evaluate how nanocarrier-based delivery systems enable controlled, spatiotemporal manipulation of genetic information to overcome off-target effects, cytotoxicity, and limitations in clinical translation.

KEY FINDINGS: CRISPR/Cas9 has emerged as a simple and programmable tool for correcting cancer-associated mutations and regulating adaptive immune responses; however, challenges such as off-target effects, unintended mutations in healthy cells, and cytotoxicity hinder its clinical application. Nanocarriers address these limitations through refined spatiotemporal delivery of Cas9 nuclease and sgRNA using internal and external stimuli-responsive functional groups. These systems improve cancer-cell specificity by engineering guide RNAs, prevent premature clearance, enhance systemic circulation and intracellular delivery, enable nuclear targeting, and regulate Cas9 activity. Stimuli such as light, heat, ultrasound, magnetic fields, pH, redox conditions, glutathione, and oxygen play key roles in controlled activation and release.

SUMMARY: This review critically evaluates the structural design of nanocarriers, advanced spatiotemporal regulation strategies, and safety and efficacy concerns in CRISPR/Cas9-based cancer therapeutics. It discusses the role of cell-specific promoters, small-molecule stimulation, and stimuli-responsive delivery systems in improving genome-editing precision and therapeutic outcomes. The review also outlines future opportunities for exploiting CRISPR/Cas9 in advanced biomedical applications to enhance the effectiveness of next-generation cancer therapy.},
}

Humans
*CRISPR-Cas Systems/genetics
*Gene Editing/methods
*Neoplasms/genetics/therapy
Animals
*Nanoparticles
Drug Delivery Systems/methods
Drug Carriers
Genetic Therapy/methods

@article {pmid41235827,
year = {2026},
author = {Chen, W and Zhang, X and Fan, R and Li, X and Guan, F and Wan, G and Kong, W and Qi, X and Pan, S and Shi, S and Su, Y and Gao, S and Huang, W and Xian, X and Liu, J and Wang, Y and Ma, Y},
title = {Generating golden Syrian hamsters with conditional alleles via zygote microinjection of CRISPR/Cas9.},
journal = {Animal models and experimental medicine},
volume = {9},
number = {2},
pages = {308-318},
doi = {10.1002/ame2.70107},
pmid = {41235827},
issn = {2576-2095},
support = {2060204//State Key Laboratory Special Fund/ ; Peking University, 202411//Open Research Project in State Key Laboratory of Vascular Homeostasis and Remodeling/ ; 2023-PT180-01//The Non-profit Central Research Institute Fund of the Chinese Academy of Medical Sciences/ ; HH24KYZX0007//Haihe Laboratory of Cell Ecosystem Innovation Fund/ ; 2021-I2M-1-024//CAMS Innovation Fund for Medical Sciences/ ; 2022-I2M-1-020//CAMS Innovation Fund for Medical Sciences/ ; 2023-I2M-2-001//CAMS Innovation Fund for Medical Sciences/ ; 2021YFF0702802//the National Key Research and Development Program of China from the Ministry of Science and Technology/ ; },
mesh = {Animals ; *CRISPR-Cas Systems/genetics ; *Mesocricetus/genetics ; *Microinjections ; Cricetinae ; *Zygote ; Alleles ; Gene Knockout Techniques ; Female ; Male ; Animals, Genetically Modified ; },
abstract = {BACKGROUND: The golden Syrian hamster is a valuable animal model for studying carcinogenesis, metabolic disorders, cardiovascular diseases, and viral infections due to its biological and pathological similarities to humans. However, the development of genetically engineered hamsters has lagged behind that of mice and rats, largely because of an embryonic development block at the two-cell stage in vitro. Although CRISPR/Cas9-mediated gene knockout has been achieved in hamsters, precise DNA fragment insertion or conditional knockout (cKO) models have not previously been reported, likely due to technical limitations in embryo manipulation and insufficient efficiency of homology-directed repair (HDR).

METHODS: In this study, we generated conditional alleles of the ApoF gene in golden Syrian hamsters. A two-cut strategy was applied using Cas9 protein, two sgRNAs, and a single donor plasmid containing exon 2 flanked by loxP sites and two ~0.8 kb homology arms. A mixture of Cas9 protein, sgRNAs, and the donor plasmid was microinjected into the pronuclei of one-cell stage hamster embryos.

RESULTS: The efficiency of CRISPR/Cas9-mediated loxP knock-in reached up to 27%, and the genetically modified floxed alleles were successfully transmitted through the germline. The functionality of the inserted loxP sites was validated by in vivo Cre-mediated recombination following local administration of AAV vectors, including AAV-cTnT-Cre in the heart and AAV-CMV-Cre in the brain.

CONCLUSIONS: To our knowledge, this work represents the first successful establishment of a conditional knockout model in the golden Syrian hamster, providing a valuable tool for mechanistic studies of gene function and disease modeling.},
}

Animals
*CRISPR-Cas Systems/genetics
*Mesocricetus/genetics
*Microinjections
Cricetinae
*Zygote
Alleles
Gene Knockout Techniques
Female
Male
Animals, Genetically Modified

@article {pmid41191919,
year = {2026},
author = {Liu, C and Xu, F and Wu, Y and Li, J and Ni, M and Xia, S and Chen, L and Zhao, H and Yu, M and Zhou, Y and Zhang, M and Li, J and Wu, X and Huang, Y and Zhu, T and Chen, X},
title = {Genome-wide CRISPR-Cas9 screening identifies CLK1 inhibition as a strategy to restore PARP inhibitor sensitivity via ERCC1 isoform switching.},
journal = {Protein & cell},
volume = {17},
number = {3},
pages = {248-262},
doi = {10.1093/procel/pwaf091},
pmid = {41191919},
issn = {1674-8018},
support = {82403208//National Natural Science Foundation of China/ ; 82303660//National Natural Science Foundation of China/ ; 82272898//National Natural Science Foundation of China/ ; 82272682//National Natural Science Foundation of China/ ; 82503636//National Natural Science Foundation of China/ ; SACA-AX202215//Shanghai Anti-Cancer Association/ ; 22YF1408100//Science and Technology Commission of Shanghai Municipality/ ; 16411950200//Science and Technology Commission of Shanghai Municipality/ ; KW1711//Science and Technology Commission of Shanghai Municipality/ ; 17411963000//Science and Technology Commission of Shanghai Municipality/ ; 22YF1409100//Science and Technology Commission of Shanghai Municipality/ ; CORP-252-1//Chinese Anti-Cancer Association-Hengrui PARP Inhibitor Cancer Research Fund/ ; 2025ZD0545600//Noncommunicable Chronic Diseases-National Science and Technology Major/ ; LGF21H160008//Social Development Project of Public Welfare Technology Research in Zhejiang Province/ ; },
mesh = {Humans ; *CRISPR-Cas Systems ; Female ; *Protein-Tyrosine Kinases/genetics/metabolism ; *Endonucleases/metabolism/genetics ; *Poly(ADP-ribose) Polymerase Inhibitors/pharmacology ; *Ovarian Neoplasms/drug therapy/genetics/pathology/metabolism ; Animals ; *DNA-Binding Proteins/metabolism/genetics ; Cell Line, Tumor ; *Protein Serine-Threonine Kinases/antagonists & inhibitors/genetics/metabolism ; Mice ; Phthalazines/pharmacology ; Piperazines/pharmacology ; Drug Resistance, Neoplasm/drug effects ; *Carcinoma, Ovarian Epithelial/drug therapy/genetics/pathology/metabolism ; Protein Isoforms/metabolism/genetics ; Apoptosis/drug effects ; },
abstract = {Epithelial ovarian cancer (EOC) is an aggressive malignancy with limited therapeutic options. Poly(ADP-ribose) polymerase inhibitors (PARPi) have shown remarkable efficacy, especially in BRCA-mutant patients, and are approved as maintenance therapy to prevent recurrence after initial response to chemotherapy. However, the development of PARPi resistance poses a major clinical challenge. This study utilized a whole-genome CRISPR-Cas9 genetic screening to identify genes associated with PARPi sensitivity upon knockout. Based on the screening and validated through further experiments, we confirmed that CLK1 knockdown is synthetically lethal with PARPi in ovarian cancer. The combination of the PARPi Olaparib and CLK1 inhibitor TG003 exhibited potent anti-proliferative effects both in vitro and in vivo. Mechanistically, CLK1 inhibition downregulated the functional ERCC1-202 isoform, resulting in enhanced DNA damage and apoptosis. Our findings reveal a novel mechanism underlying PARPi sensitivity and suggest that targeting CLK1 in combination with PARPi may represent a promising therapeutic strategy for PARPi-resistant ovarian cancer.},
}

Humans
*CRISPR-Cas Systems
Female
*Protein-Tyrosine Kinases/genetics/metabolism
*Endonucleases/metabolism/genetics
*Poly(ADP-ribose) Polymerase Inhibitors/pharmacology
*Ovarian Neoplasms/drug therapy/genetics/pathology/metabolism
Animals
*DNA-Binding Proteins/metabolism/genetics
Cell Line, Tumor
*Protein Serine-Threonine Kinases/antagonists & inhibitors/genetics/metabolism
Mice
Phthalazines/pharmacology
Piperazines/pharmacology
Drug Resistance, Neoplasm/drug effects
*Carcinoma, Ovarian Epithelial/drug therapy/genetics/pathology/metabolism
Protein Isoforms/metabolism/genetics
Apoptosis/drug effects

@article {pmid41806414,
year = {2026},
author = {Kim, H and Kim, D and Han, H and Lee, C and Roh, YH and Han, TS and Lim, EK and Park, J and Ahn, JK and Kang, T and Jung, J and Lee, CY},
title = {On-site microRNA detection with 'off-the-shelf' glucose meter empowered by chimeric probe connecting CRISPR/Cas13a activation to kinases-driven glucose phosphorylation.},
journal = {Biosensors & bioelectronics},
volume = {304},
number = {},
pages = {118568},
doi = {10.1016/j.bios.2026.118568},
pmid = {41806414},
issn = {1873-4235},
abstract = {MicroRNAs (miRNAs) are promising biomarkers for cancer diagnosis due to their stability in body fluids and disease-specific expression profiles. However, current detection methods suffer from limitations including cumbersome workflows, heavy instrumentation for signal readout, or vulnerability in minimizing instrumentation. To address these challenges, we describe a novel point-of-care miRNA detection platform executable with "off-the-shelf", personal glucose meter (PGM), termed 'KEY-FACT (Kinases Ensemble-driven glucose phosphorYlation upon Fuel-Aided CRISPR acTivation)'. Upon recognition of target miRNA, a fuel-assisted toehold-mediated strand displacement reactions liberate guide RNAs (gRNAs) to activate Cas13a to cleave a chimeric reporter probe, producing 2',3'-cyclic adenosine monophosphates (cAMP). Subsequent dephosphorylation and kinases ensemble-mediated phosphorylation/dephosphorylation cycles lead cAMP to consume a large amount of glucose. A user can immediately measure resulting glucose level change with PGM on the spot. This strategy allows sensitive, prompt detection of miR-135b, a gastric cancer (GC) biomarker, with a limit of detection (LOD) of 1.4 pM within 2 h. KEY-FACT is specific to the target miRNA and is applicable to body fluids such as human serum with dilution (95.2% < recovery rates <104.3%, coefficients of variation ≤13%). Owing to its simple probe design, KEY-FACT was readily expanded to detect another GC biomarker, miR-21, with comparable sensitivity (LOD = 1.5 pM). The proposed platform fulfills minimal instrumentation and thus enables cost-effective, field-deployable analysis, paving the way for practical, on-demand miRNA diagnostics.},
}

@article {pmid41806413,
year = {2026},
author = {Jeong, Y and Lee, J and Choi, S and Shin, D and Jang, S and Son, SU and Kang, T and Jung, J and Hwang, J and Lim, EK},
title = {On-site detection of airborne foodborne pathogens using a field-deployable recombinase polymerase amplification and CRISPR/Cas12a cleavage activity assay.},
journal = {Biosensors & bioelectronics},
volume = {304},
number = {},
pages = {118571},
doi = {10.1016/j.bios.2026.118571},
pmid = {41806413},
issn = {1873-4235},
abstract = {With the global increase in single-person households, the demand for meal kits is increasing, leading to the development of large-scale food production systems and complex supply chains. However, under the influence of global warming, these systems can be susceptible to food contamination, particularly by airborne foodborne bacteria. Conventional methods for detecting airborne bacteria involve complex, time-consuming, and labor-intensive processes, which limit their applicability for field use and rapid food hygiene surveillance. In the present study, we developed a field-deployable diagnostic platform by combining recombinase polymerase amplification with CRISPR/Cas12a cleaVage Activity (RCCVA assay) for the rapid and sensitive identification of airborne foodborne bacteria. Airborne bacteria were collected using a self-developed electrostatic air sampler and analyzed using a portable isothermal amplification device. The RCCVA assay was designed to detect four major foodborne pathogens: Staphylococcus aureus, Salmonella enteritidis, Listeria monocytogenes, and Bacillus cereus. The limit of detection was measured as 274.9, 4.5, 9.5, and 28.5 culture-forming units (CFU)/mL, respectively, within 45 min. This platform enables early on-site detection of airborne pathogens within approximately 1 h (for the analytical phase) and shows potential for real-time monitoring in food processing environments, thereby contributing to improved public health and food safety.},
}

@article {pmid41805869,
year = {2026},
author = {Moghe, AS and Nandi, SS and Bhonde, RR and Kamyab, SS and Sawant, SA and Karandikar, MN},
title = {Engineering of cell line assembled enteric organoid for enterovirus infection.},
journal = {Archives of virology},
volume = {171},
number = {4},
pages = {},
pmid = {41805869},
issn = {1432-8798},
support = {5/3/8/57/2020-ITR Date:11.03.2021//Indian Council of Medical Research/ ; },
mesh = {Humans ; *Organoids/virology ; Virus Replication ; Cell Line ; *Enterovirus Infections/virology ; *Enterovirus A, Human/physiology ; Epithelial Cells/virology ; *Enterovirus/physiology ; CRISPR-Cas Systems ; Coculture Techniques ; },
abstract = {The non-polio-enteroviruses are ubiquitous pathogens infecting over a billion people in the world. An alarming number of enterovirus-associated acute flaccid paralysis, encephalitis, hand, foot, and mouth disease, conjunctivitis and diarrhoea cases are reported worldwide. Despite their clinical significance, vaccine development has been hindered due to lack of suitable in vitro models for preclinical investigations. The present study was undertaken to develop a cell line assembled organotypic model of human intestine for replication of enteroviruses. An enterovirus specific PSGL1 receptor was introduced in intestinal epithelial HCT-8 cell line employing CRISPR/cas9 gene editing. It was co-cultured with human colon (CCD-18) and endothelial (HUVEC) cell lines with peripheral blood mononuclear cells in hanging drops and rotating wall vessel bioreactor to yield three-dimensional organoids. Histological analysis of the organoids showed presence of columnar epithelium cells with prominent intracytoplasmic mucin, hyperchromatic nuclei and presence of CK, CK20, MUC 2 and Villin markers characteristic of epithelial cells. Infection with Enterovirus A71 (EV-A71) demonstrated significantly higher viral titre in organoids compared to individual cell lines. Collectively, these findings determine, for the first time, a cell line-derived enteric organoid model that supports robust enterovirus replication, offering a cost-effective and physiologically relevant system for virology research and preclinical applications.},
}

Humans
*Organoids/virology
Virus Replication
Cell Line
*Enterovirus Infections/virology
*Enterovirus A, Human/physiology
Epithelial Cells/virology
*Enterovirus/physiology
CRISPR-Cas Systems
Coculture Techniques

@article {pmid41805349,
year = {2026},
author = {Han, F and Xu, Y and Wang, W and Li, Z and Zhang, Z and Du, R and Xu, Q},
title = {Methylobacterium as a Dual-Function Platform: Advances in C1-Based Biomanufacturing and Plant-Associated Applications.},
journal = {Journal of agricultural and food chemistry},
volume = {},
number = {},
pages = {},
doi = {10.1021/acs.jafc.5c08026},
pmid = {41805349},
issn = {1520-5118},
abstract = {One-carbon (C1) substrates are promising feedstocks for microbial bioproduction. Methylobacterium, known for its exceptional C1 utilization capacity, has emerged as a model microbial chassis for sustainable biomanufacturing. In this review, we first outline the C1 assimilation pathways in Methylobacterium and underscore its potential for producing valuable native metabolites. Furthermore, we then survey the genetic tools available for engineering this genus, including plasmid-based methods, transposon mutagenesis, homologous recombination, and CRISPR/Cas systems. Notably, recent advances in metabolic engineering have significantly expanded its biosynthetic scope, enabling the biosynthesis of diverse non-native compounds. Beyond its biomanufacturing potential, Methylobacterium also serves as a versatile plant growth-promoting bacterium, enhancing plant health and productivity through hormone synthesis, nutrient mobilization, stress mitigation, and induced systemic resistance. Collectively, this work highlights the dual potential of Methylobacterium as a sustainable microbial cell factory for biomanufacturing and a beneficial bioinoculant for agriculture.},
}

@article {pmid41805130,
year = {2026},
author = {Yu, W and Yuan, L and Zhou, W and He, L and Huang, X and Yu, J and Deng, J and Zhang, T and Hu, Y and Zhang, Y and Chen, S},
title = {Orn-mediated c-di-GMP regulates the CRISPR-Cas system to confer stress response in Mycobacterium tuberculosis.},
journal = {Nucleic acids research},
volume = {54},
number = {5},
pages = {},
pmid = {41805130},
issn = {1362-4962},
support = {2021YFA1300901//National Key R&D Program of China/ ; 2022YFA1303500//National Key R&D Program of China/ ; GZNL2024A01024//Guangzhou National Laboratory/ ; //National Key Research and Development Program of China/ ; },
mesh = {*Mycobacterium tuberculosis/genetics/metabolism/drug effects ; *CRISPR-Cas Systems/genetics ; *Cyclic GMP/analogs & derivatives/metabolism ; Bacterial Proteins/genetics/metabolism ; Gene Expression Regulation, Bacterial ; Oxidative Stress/genetics ; *Stress, Physiological/genetics ; Promoter Regions, Genetic ; CRISPR-Associated Proteins/genetics/metabolism ; },
abstract = {Mycobacterium tuberculosis (Mtb) possesses a type III-A CRISPR-Cas system and has anti-plasmid immune activity. However, whether this system exerts other additional functions remains to be characterized. Here, we investigated the in vivo roles of the Mtb CRISPR-Cas system. We show that this system is transcriptionally dependent and exhibits limited ability to counteract exogenous nucleic acids, primarily through the Csm6 protein rather than the Cas10 HD domain. We further demonstrate that this system plays a role in mitigating oxidative stress and antibiotic treatment, a function mainly mediated by the Cas10 HD domain. Importantly, through transposon library screening, we identified oligoribonuclease (Orn) as a regulatory protein of the Mtb CRISPR-Cas system. Deletion of the orn gene resulted in elevated c-di-GMP levels. A subsequent biotin-labeled c-di-GMP pull-down assay identified the transcriptional regulator Rv3058. Knockdown of rv3058 significantly increased cas6 promoter activity, and its transcriptional repressor function was directly modulated by c-di-GMP. This regulatory pathway enhances stress defense by activating multiple protective pathways, including DNA repair, cell envelope maintenance, and iron homeostasis regulation. Together, we conclude that the regulation of the CRISPR-Cas system by Orn-mediated c-di-GMP contributes to oxidative and antibiotic stress responses in Mtb.},
}

*Mycobacterium tuberculosis/genetics/metabolism/drug effects
*CRISPR-Cas Systems/genetics
*Cyclic GMP/analogs & derivatives/metabolism
Bacterial Proteins/genetics/metabolism
Gene Expression Regulation, Bacterial
Oxidative Stress/genetics
*Stress, Physiological/genetics
Promoter Regions, Genetic
CRISPR-Associated Proteins/genetics/metabolism

@article {pmid41804827,
year = {2026},
author = {Parada, F and Cabedo-Díaz, P and Cerda, A and Osorio-Navarro, C and Toledo, JA and Villalobos-González, L and Handford, M and Pimentel, P},
title = {CRISPR/dCas9-Mediated BRL3 Activation Enhances Growth and Metabolic Resilience Under Osmotic Stress in Nicotiana tabacum.},
journal = {Physiologia plantarum},
volume = {178},
number = {2},
pages = {e70816},
doi = {10.1111/ppl.70816},
pmid = {41804827},
issn = {1399-3054},
support = {3240290//ANID-FONDECYT Postdoctoral Project/ ; 3210631//ANID-FONDECYT Postdoctoral Project/ ; 1231417//ANID-FONDECYT Regular Project/ ; RF23F0002//ANID Fortalecimiento de Centros Regionales Project/ ; NCN2024_047//ANID-Millennium Science Initiative Program/ ; },
mesh = {*Nicotiana/genetics/growth & development/metabolism/physiology ; Osmotic Pressure/physiology ; Gene Expression Regulation, Plant ; *Plant Proteins/metabolism/genetics ; *CRISPR-Cas Systems/genetics ; Plants, Genetically Modified ; Promoter Regions, Genetic/genetics ; Brassinosteroids/metabolism ; Plant Leaves/genetics ; },
abstract = {Brassinosteroids (BRs) are crucial plant hormones that influence growth and stress adaptation. However, the specific function of the BR receptor BRL3 under osmotic stress remains largely unexplored outside Arabidopsis thaliana. In this study, we used a CRISPR/dCas9-based transcriptional activation (CRISPRa) system to upregulate the Nicotiana tabacum BRASSINOSTEROID INSENSITIVE-LIKE 3 receptor (NtBRL3) and assessed its impact on osmotic stress tolerance. Synthetic activation vectors were constructed using Loop Assembly, featuring dCas9-6TAL-VP128 modules driven by either a constitutive (CaMV35S) or ABA-inducible (SlAREB) promoter, paired with dual sgRNAs targeting the NtBRL3 promoter. Transient Agrobacterium-mediated transformation followed by PEG treatment was used to impose osmotic stress. RT-qPCR confirmed a 3- to 4-fold activation of NtBRL3 transcripts in CRISPRa-infiltrated leaves. The stress-inducible SlAREB promoter produced the strongest improvements, yielding nearly four-fold higher leaf biomass and a five-fold increase in root biomass relative to PEG-stressed controls. Both constructs reduced malondialdehyde (MDA) accumulation, indicating diminished oxidative damage, and modulated osmoprotectant balance, including reduced root proline and increased total soluble solids, particularly under SlAREB-driven activation. Histological segmentation revealed promoter-dependent anatomical remodeling, with NtBRL3-activated plants exhibiting a higher frequency of enlarged leaf cells and expanded tissue domains, consistent with brassinosteroid-mediated structural plasticity. Collectively, these findings demonstrate that CRISPR/dCas9-mediated transcriptional activation of NtBRL3 enhances osmotic stress resilience in tobacco through coordinated biomass recovery, oxidative stress mitigation, osmolyte homeostasis, and tissue remodeling. This transient, non-integrative CRISPRa approach provides a robust synthetic biology framework for dissecting BR signaling and engineering stress-tolerant crops.},
}

*Nicotiana/genetics/growth & development/metabolism/physiology
Osmotic Pressure/physiology
Gene Expression Regulation, Plant
*Plant Proteins/metabolism/genetics
*CRISPR-Cas Systems/genetics
Plants, Genetically Modified
Promoter Regions, Genetic/genetics
Brassinosteroids/metabolism
Plant Leaves/genetics

@article {pmid41803497,
year = {2026},
author = {Perez Taboada, V and Wu, Y and Cassidy, R and Medvedev, KE and Loeff, L and Nemudraia, A and Nemudryi, A},
title = {Bacterial Schlafen proteins mediate phage defence.},
journal = {Nature microbiology},
volume = {},
number = {},
pages = {},
pmid = {41803497},
issn = {2058-5276},
support = {R00AI171893//U.S. Department of Health & Human Services | NIH | National Institute of Allergy and Infectious Diseases (NIAID)/ ; 1T32GM156737-01//U.S. Department of Health & Human Services | NIH | National Institute of General Medical Sciences (NIGMS)/ ; },
abstract = {Human Schlafen proteins restrict viral replication by cleaving tRNA, thereby suppressing protein synthesis. Although the ribonuclease domain of Schlafen proteins is conserved across all domains of life, its function in prokaryotes has remained unclear. Here we demonstrate that prokaryotic Schlafen nucleases are widespread antiviral effectors that protect bacteria from bacteriophages and are fused to a diverse array of phage-sensing domains. We expressed seven Enterobacterales Schlafen systems in Escherichia coli, identifying two that confer defence against coliphages. We focused on a system where Schlafen nuclease is fused to a previously unknown immunoglobulin-like sensor domain and demonstrated that it recognizes tail assembly chaperones of T5-like phages. Upon activation, the Schlafen nuclease cleaves both E. coli and phage-encoded tRNAs and restricts T5 phage by reducing its burst size. Our findings redefine Schlafens as an ancient, mechanistically conserved family of immune effectors, revealing the deep evolutionary origin of tRNA-targeting antiviral immunity in humans.},
}

@article {pmid41803127,
year = {2026},
author = {Zhou, R and Liu, Y and Zhang, Q and Yin, Z and Tong, J and Zhang, C and Zhang, L and Li, X and Zhao, Y and Zhang, S and Liu, Z and Chen, W and Ji, N and Zhang, H and Li, Z and Yin, H and Zuo, S and Wei, Y},
title = {Structural and mechanistic insights into the dual-nuclease defense protein Upx as an anti-phage system.},
journal = {Nature communications},
volume = {},
number = {},
pages = {},
doi = {10.1038/s41467-026-70435-x},
pmid = {41803127},
issn = {2041-1723},
support = {2022YFC3400400//National Science Foundation of China | Key Programme/ ; 23ZYCGSY00750//Tianjin Science and Technology Committee (Tianjin Municipal Science and Technology Commission)/ ; },
abstract = {Nucleic acid degradation is a common strategy for prokaryotic anti-phage systems, as exemplified by the CRISPR-Cas system. The PD-(D/E)-XK nucleases constitute a widely distributed family in these defenses. Notably, most members exhibit a single nuclease domain, while variants containing dual nuclease domains within a single polypeptide remain underexplored, and their molecular mechanisms largely obscure. Here, we biochemically and functionally study a single-protein system containing an uncharacterized PD-(D/E)-XK defense protein (Upx). As revealed by single-particle electron cryo-microscopy (cryo-EM) structure, the C-terminal domain (CTD) harboring the conserved PD-(D/E)XK catalytic core is buttressed by the N-terminal domain (NTD) and the middle domain (MD). Functional assays demonstrate that the nucleic acid binding capability of the CTD is enhanced by the MD. The NTD also displays a noncanonical, basal exonuclease activity that is auto-inhibited by MD. IP-MS experiments identify Upx-interacting phage proteins, and substrate profiling defines its physiological preferences, collectively pointing to its potential physiological targets. Notably, the phage protein gp16 was found to relieve MD-mediated inhibition of the NTD, suggesting a virus-triggered mechanism for activating Upx's dual nuclease activity. Together, these findings establish Upx as a single-protein dual-nuclease anti-phage system, expanding our understanding of bacterial immunity and informing antiviral strategy development.},
}

@article {pmid41802999,
year = {2026},
author = {Hao, M and Zhou, M and Pan, F and Liu, T and Li, Y and Su, N and Ashfaq, A and Song, M and Wang, H and Wang, W and Liu, J and Li, C and Fu, L and He, P and Hu, Q and Mei, D and Cheng, H},
title = {Efficient CRISPR/Cas-SF01 genome editing tools with high editing efficiency in allotetraploid oilseed rape.},
journal = {Journal of integrative plant biology},
volume = {},
number = {},
pages = {},
doi = {10.1111/jipb.70221},
pmid = {41802999},
issn = {1744-7909},
support = {2025BEA003//the Major Program (JD) of Hubei Province/ ; CAAS-CSNCB-202303//Innovation Program of Chinese Academy of Agricultural Sciences/ ; 2025AFB468//Hubei Provincial Natural Science Foundation of China/ ; CARS-12//Earmarked Fund for China Agriculture Research System/ ; },
abstract = {CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)-Cas9 has been widely utilized for plant genome editing, but the protospacer adjacent motif (PAM) requirement limits its editing scope. CRISPR/Cas12i3 belongs to the type-VI Cas system that has gained extensive attention due to its smaller size and less restricted canonical TTN PAM sequence. In this study, we explored the newly developed Cas-SF01 system (Cas12i3 variant) for genome editing in oilseed rape. We established an efficient protoplast transformation system in oilseed rape to compare editing efficiency between Cas-SF01 and Cas9. Cas-SF01 shows cleavage activities at the tested 5'-TTN-3' PAM sites with editing outcomes sharing considerable similarities with the CRISPR-Cas9 system in protoplast. Cas-SF01 also induces high efficiency mutagenesis for multiple target sites in stable transformed oilseed rape lines, generating mutants with multilocular silique and male sterile phenotypes. Furthermore, Cas-SF01-derived cytosine base editors (CBEs) were developed to produce targeted C-to-T base edits. Compared to SpCas9, Cas-SF01 has an expanded PAM range and effectively recognizes TTN PAMs, which has substantially broadened the scope of editable sites within the rapeseed genome. No mutations were identified at the putative off-target sites among the edited plants. This study developed a robust, first-of-its-kind Cas12 system in the allotetraploid Brassica napus, expanding the scope of editing and enriching genome-editing toolkits for biological research and genetic improvement.},
}