Search Results (3,909)

Genome-wide studies have identified significant allelic associations between genetic variants in or near the IKZF1 gene and multiple autoimmune disorders. IKZF1, encoding the transcription factor IKAROS, produces at least 10 distinct transcripts. To explore the impact of alternative splicing of IKZF1 on the function of mature T cells and the risk of autoimmunity, we generated a panel of human T-cell clones with truncating mutations in IKZF1 exons 4, 6, or both. Differences in gene expression, chromatin accessibility, and protein abundance among clones were assessed by RNA-seq, ATAC-seq, and immunoblotting. Clones with single targeting events clustered separately from double-targeted clones on multiple parameters, but overall, clone responses were highly heterogeneous. Perturbation of IKZF1 splicing resulted in significant differences in expression and chromatin accessibility of other autoimmunity-associated genes and elicited compensatory expression changes in other IKAROS family members. Our results suggest that even modest alterations of IKZF1 splicing can have significant effects on gene expression and function in mature T cells, potentially contributing to autoimmunity in susceptible individuals. Full article

The neurofibromin 1 (NF1) splice-site mutation c.61-2A>G (rs1131691100) is a rare, pathogenic, autosomal dominant variant that disrupts NF1 tumor-suppressor function, causing neurofibromatosis type 1 (NF1). Its pathogenic mechanism is poorly understood, and the potential for personalized therapeutic genome editing remains unknown due to the absence of a standard framework for investigating splicing disorders. Here, we performed a comprehensive multi-omics analysis of a de novo c.61-2A>G case from South Korea, integrating short- and long-read whole genome sequencing, whole transcriptome sequencing, and methylation profiling. We confirm that c.61-2A>G abolishes the canonical splice acceptor site, activating a cryptic splice acceptor 16 nucleotides downstream in exon 2. This splicing shift generates a 16-nucleotide deletion, causing a frameshift and premature stop codon that truncates the protein’s N-terminal region. Long-read sequencing further reveals that the mutation creates a novel CpG dinucleotide, which is methylated in the majority of reads. Finally, we assessed therapeutic correction strategies, revealing that CRISPR-Cas9 prime editing is the only viable approach for in vivo correction. This study provides the first comprehensive multi-omics characterization of the NF1 c.61-2A>G mutation and establishes a minimal framework for precision therapeutic development in silico in monogenic splicing disorders. Full article

Glioblastoma (GBM) is the most aggressive primary brain tumor, marked by molecular heterogeneity and poor clinical prognosis. Lysyl oxidase-like 3 (LOXL3) is frequently upregulated in GBM, but its mechanistic contribution remains insufficiently defined. Here, we investigated the functional role of LOXL3 in GBM using CRISPR-Cas9-mediated LOXL3 knockdown in two genetically distinct GBM cell lines: U87MG (wild-type TP53) and U251 (mutant TP53). Reduced LOXL3 expression markedly reduced α-tubulin acetylation, particularly in U87MG cells, and downregulated genes involved in cell cycle progression and proliferation. Both cell lines exhibited mitotic defects, including delayed cell cycle progression and spindle abnormalities; however, cell fate diverged according to TP53 status. U87MG cells, sustained spindle checkpoint activation triggered a p53-dependent spindle checkpoint response culminating in apoptosis, while U251 cells underwent mitotic slippage and senescence. Transcriptomic analyses confirmed differential regulation of apoptosis versus senescence pathways in accordance with TP53 functionality. Additionally, reduced LOXL3 expression markedly impaired adhesion and migration in U87MG cells, whereas U251 cells were minimally affected, consistent with more pronounced microtubule destabilization. Collectively, these findings identify that LOXL3 is a key regulator of microtubule homeostasis, mitotic fidelity, adhesion, and invasive behavior in GBM. Targeting LOXL3 may therefore provide a therapeutic opportunity for genotype-informed intervention in GBM. Full article

Premelanosomal protein (Pmel, also known as Pmel17) is the major component of melanosomal fibrils and plays a key role in melanin polymerization, making it an important factor in melanogenesis. We investigated how the absence of Pmel affects the properties of B16F10 melanoma cells. Pmel-knockout B16F10 cells were generated using CRISPR/Cas9-mediated genome editing. A viability assay revealed no significant differences between wild-type (WT) and Pmel-knockout (KO) sublines; however, melanosome maturation was impaired. In Pmel KO cells, the cell cycle was disrupted, and higher levels of reactive oxygen species (ROS) were observed compared with WT cells. Moreover, the migration capacity and tube formation of melanoma cells were increased. Tumors derived from Pmel KO cells exhibited unchanged growth kinetics but reduced melanin content, along with enhanced vascularization and oxygenation. Thus, knockout of the Pmel17 gene in melanoma cells alters pigmentation, vascularization, and oxygenation of tumors. These parameters are crucial for both tumor progression and therapeutic response. Full article

Ascorbic acid (AsA) is a key antioxidant and nutrient in plants, regulating reactive oxygen species (ROS) levels and maintaining cellular redox homeostasis. The AsA recycling pathway sustains AsA pools by restoring its oxidized forms, ensuring intracellular balance. Among the enzymes involved, monodehydroascorbate reductase (MDHAR) is important for the regeneration of AsA from monodehydroascorbate. In this study, we analyzed the four MDHAR paralogs in Lactuca sativa using CRISPR/Cas9 to determine whether disruption of individual MDHAR genes could alter AsA levels in lettuce leaves. Unexpectedly, none of the knockouts caused long-term changes in leaf AsA content. Transcriptomic analyses at 14 and 28 days showed minimal effects on AsA recycling or biosynthesis genes, except MDHAR genes. However, several other genes indirectly implicated in AsA regulation displayed differential expression in all mutants compared to the wild type, suggesting the presence of a complex regulatory network. In particular, genes encoding transcription factors (TFs), such as mTERF15, COL9, UPBEAT1, NAC28, and NAC42, were differentially regulated in all MDHAR mutants compared to the wild type at 28 days. These findings indicate that, although AsA content remains unchanged, MDHAR single knockouts alter expression of other genes through which the plants may indirectly compensate to maintain redox homeostasis. Full article

Fungal diseases continue to limit global crop production and drive major economic losses. Conventional diagnostic and control approaches depend on time-consuming culture-based methods and broad-spectrum chemicals, which offer limited precision. Advances in molecular identification have changed this landscape. PCR, qPCR, LAMP, sequencing and portable platforms enable rapid and species-level detection directly from plant tissue. These tools feed into RNA-based control strategies, where knowledge of pathogen genomes and sRNA exchange enables targeted suppression of essential fungal genes. Host-induced and spray-induced gene silencing provide selective control without the long-term environmental costs associated with chemical use. CRISPR/Cas9 based tools now refine both diagnostics and resistance development, and bioinformatics improves target gene selection. Rising integration of artificial intelligence indicates a future in which disease detection, prediction and management connect in near real time. The major challenge lies in limited field validation and the narrow range of fungal species with complete molecular datasets, yet coordinated multi-site trials and expansion of annotated genomic resources can enable wider implementation. The combined use of molecular diagnostics and RNA-based strategies marks a shift from disease reaction to disease prevention and moves crop protection towards a precise, sustainable and responsive management system. This review synthesizes the information related to current molecular identification tools and RNA-based management strategies, and evaluates how their integration supports precise and sustainable approaches for fungal disease control under diverse environmental settings. Full article

The Maternal Embryonic Leucine Zipper Kinase (MELK) gene is a member of the Snf1/AMPK serine/threonine kinase family. MELK has recently attracted considerable interest in cancer biology due to its aberrant overexpression in various malignancies, including glioma, breast, lung, colorectal, gastric, and hematological cancers. It has been shown that higher MELK levels are often correlated with unfavorable prognosis, aggressive tumor manifestations, resistance to treatment, and stem-like tumor morphologies. In this review we aim to summarize the current understanding of MELK biology, including its functions in cell cycle regulation, apoptosis, oncogenic signaling pathways, and tumor stemness. We also discuss the therapeutic potential, limitations, and controversy of MELK inhibitors, and implications in cancer diagnosis and treatment. MELK may not be a universal driver oncogene; nonetheless, it is consistently linked to aggressive disease, underscoring its potential as a prognostic biomarker and a candidate for therapeutic co-targeting in combination treatments. Full article

Climate change introduces a critical threat to global viticulture, compromising grape yield, quality, and the long-term sustainability of Vitis vinifera cultivation. Addressing these challenges requires innovative strategies to enhance grapevine resilience. The integration of multi-omics data, predictive breeding, and physiological insights into ripening and stress responses is refining our understanding of grapevine adaptation mechanisms. In parallel, recent advances in plant biotechnology have accelerated progress from marker-assisted and genomic selection to targeted genome editing, with CRISPR/Cas systems and other New Genomic Techniques (NGTs) offering advanced precision tools for sustainable improvement. This review synthesizes the major achievements in grapevine genetic improvement over time, tracing the evolution of strategies from traditional breeding to modern genome editing technologies. Overall, we highlight how combining genetics, biotechnology, and physiology is reshaping grapevine breeding towards more sustainable viticulture. The convergence of these disciplines establishes a new integrated framework for developing resilient, climate-adapted grapevines that maintain yield and quality while preserving varietal identity in the face of environmental change. Full article

CRISPR-Cas9 systems have enabled unprecedented advances in genome engineering, particularly in developing treatments for human diseases, like cancer. Despite potential applications, limitations of Cas9 include its relatively large size and strict targeting requirements. Cas12j2, a variant ofCasΦ-2, shows promise for overcoming these limitations. However, its effectiveness in mammalian cells remains relatively unexplored. This study sought to develop an optimized CRISPR-Cas12j2 system for targeted knockout of the E6 oncogene in HPV-associated cancers. A combination of computational tools (ColabFold, CCTop, Cas-OFFinder, HADDOCK2.4, and Amber for Molecular Dynamics) was utilized to investigate the impact of engineered modifications on structural integrity and gRNA binding of Cas12j2 fusion constructs, in potential intracellular conditions. Cas12j2_F2, a Cas12j2 variant designed and evaluated in this study, behaves similarly to the wild-type Cas12j2 structure in terms of RMSD/RMSF profiles, compact Rg values, and minimal electrostatic perturbation. The computationally validated Cas12j2 variant was incorporated into a custom expression vector, co-expressing the engineered construct along with a dual gRNA for packaging into a viral vector for targeted knockout of HPV-associated cancers. This study provides a structural and computational foundation for the rational design of Cas12j2 fusion constructs with enhanced stability and functionality, supporting their potential application for precise genome editing in mammalian cells. Full article

Epithelial ovarian cancer (EOC) represents the most lethal malignancy arising from the female reproductive tract, largely due to the clinical challenge of chemotherapy resistance. Recent studies indicate that ferroptosis—a distinct form of programmed cell death driven by iron accumulation and lipid peroxidation, could potentially exploit a vulnerability in chemoresistant cancer cells. Here, we identify MED12 as a critical regulator of ferroptosis sensitivity in EOC through modulation of the YAP–TEAD1 signaling pathway. Using CRISPR/Cas9-mediated knockout and rescue experiments in EOC cell lines, we demonstrate that MED12 deficiency significantly enhances sensitivity to ferroptosis inducers (RSL3 and Erastin), as evidenced by reduced IC50 values. Transcriptomic and chromatin accessibility analyses reveal that MED12 loss activates YAP signaling through TEAD1 upregulation, increasing chromatin accessibility at YAP–TEAD1 target loci and elevating the expression of downstream effectors CYR61 and CTGF. Pharmacological inhibition of YAP with verteporfin or siRNA-mediated TEAD1 knockdown reverses ferroptosis sensitivity in MED12-deficient cells, confirming pathway specificity. These findings establish MED12 as a modulator of the YAP–TEAD1–ferroptosis axis and suggest that targeting this pathway could overcome chemoresistance in MED12-deficient EOC. Our work provides a mechanistic foundation for exploiting ferroptosis induction as a therapeutic strategy in ovarian cancer. Full article

Despite a thorough understanding of the structure of human papillomavirus (HPV) and its genotypic variations (high-risk and low-risk variants), the mechanisms underlying HPV-induced cervical cancer (CC) pathogenesis and the molecular signatures of drug resistance remain to be fully understood. Accumulating evidence has shown the involvement of kinase targets in the induction of drug resistance in high-risk (HR) HPV-CC. Molecularly, the genome of high-risk HPV is reported to control the expression of host kinases. In particular, Aurora kinases A, B, and C (ARKA, ARKB, and ARKC), phosphotidylinositol–trisphosphate kinase (PI3K)-Akt, and Glycogen synthase kinase3-α/β (GSK3 α/β) promote the transformation of infected cells, and also enhance the resistance of cells to various chemotherapeutic agents such as nelfinavir and cisplatin. However, the precise mechanisms through which HPV activates these kinases are yet to be fully elucidated. Furthermore, there is still ambiguity surrounding whether targeting HPV-induced kinases along with HPV-targeted therapies (such as phytopharmaceuticals and PROTAC/CRISPR-CAS-based systems) synergistically inhibit cervical tumor growth. Given the critical role of kinases in the pathogenesis and treatment of CC, a comprehensive review of current evidence is warranted. This review aims to provide key insights into the mechanisms of HPV-induced CC development, the involvement of kinases in drug resistance induction, and the rationale for combination therapies to improve clinical outcomes. Full article

The oncogene KRAS drives tumor growth by activating pathways such as MAPK and PI3K-AKT in a constitutive manner. Although direct KRAS inhibitors exist, they are often limited in clinical use due to therapeutic resistance and toxicity. Therefore, alternative combinatorial therapeutic strategies are urgently needed. This study examined the knockout of five KRAS-related proteins—galectin-3 (GAL3), phosphodiesterase delta (PDEδ), nucleophosmin (NPM1), IQ motif-containing GTPase-activating protein 1 (IQGAP1), and SHOC2—using CRISPR-Cas9 in adenocarcinoma cell lines harboring the KRAS(G12V) oncogenic mutation, as well as in the noncancerous HEK-293 cell line. These proteins act as critical modulators that regulate KRAS activity, cellular localization, and that of its downstream signaling components. We analyzed the downstream activation of ERK and AKT kinases and evaluated subsequent cancer cell proliferation. Knockout of GAL3 and PDEδ was highly effective, significantly reducing MAPK and PI3K-AKT pathway activity and substantially impairing cell proliferation. SHOC2 knockout selectively and potently disrupted MAPK activation, while NPM1 knockout resulted in the complex, reciprocal modulation of the two major pathways. Notably, knocking out IQGAP1 enhanced PI3K–AKT and mTORC2–AKT signaling without affecting the MAPK pathway. These distinct modulatory roles highlight the non-redundant functions of the accessory proteins. In conclusion, our findings establish GAL3 and PDEδ, two KRAS-associated proteins, as promising combinatorial drug targets. Targeting these modulators provides an effective alternative strategy to overcome resistance mechanisms and enhance the clinical utility of existing KRAS inhibitors. Full article

Cyanobacteria harbor sophisticated molecular defense systems that have evolved over billions of years to protect against viral invasion and foreign genetic elements. These ancient photosynthetic organisms possess a diverse array of restriction-modification (R-M) systems and CRISPR-Cas arrays that present challenges for genetic engineering, but also offer unique opportunities for cancer-targeted biotechnological applications. These systems exist in prokaryotes mainly as defense mechanisms but they are currently used in molecular applications as gene editing tools. Moreover, latest developments in nucleases such as zinc finger nucleases (ZFNs), TALENs (transcription-activator-like effector nucleases) are discussed. A comprehensive genomic analysis of 126 cyanobacterial species found 89% encode multiple R-M systems, averaging 3.2 systems per genome, creating formidable barriers to transformation but also providing molecular machinery that could be harnessed for precise recognition and targeting of cancer cells. This review critically examines the dual nature of these defense systems, their ecological functions, and the emerging strategies to translate their molecular precision into advanced anticancer therapeutics. Hence, the review main objectives are to explore the recent understanding of these mechanisms and to exploit the knowledge gained in opening new avenues for cancer-focused targeted interventions, while acknowledging the significant challenges to translate these systems from laboratory curiosities to practical applications. Full article

Nystatin is a polyene macrolide antibiotic with broad-spectrum antifungal activity and serves as a key therapeutic agent for superficial fungal infections. This review systematically elaborates on its multicomponent chemical nature, its mechanism of action targeting ergosterol, and highlights the potential adverse effects, such as cardiotoxicity, associated with impurities like RT6 (albonoursin). The fundamental analytical techniques for quality control are outlined. Furthermore, the clinical applications and combination therapy strategies of nystatin in treating oral diseases, vaginitis, and otitis externa are summarized in detail. Regarding biosynthesis, the assembly mechanism of nystatin A₁ via the type I polyketide synthase pathway and its subsequent modification processes are thoroughly discussed. Emphasis is placed on the latest advances and potential of gene-editing technologies, particularly CRISPR/Cas9, in the targeted knockout of genes responsible for toxic components and in optimizing production strains to enhance nystatin yield and purity. Finally, this review prospects the future development of nystatin towards improved safety and efficacy through structural optimization, innovative delivery systems, and synthetic biology strategies, aiming to provide a reference for its further research and clinical application. Full article