Biology subjects

Nagar, P.

Publications and source records attributed to Nagar, P..

6 recordsLinked to original sources

bioRxiv2026

CRISPR-mediated Correction of Oncogenic AS-NMD in Splicing Factor Mutant Cancer

Alternative splicing coupled to nonsense-mediated mRNA decay (AS-NMD) evolved as a master regulator of gene expression. Dysregulated AS-NMD has been identified as the root of many human maladies, from developmental defects to deadly cancer. Poison exons (PEs) are highly conserved alternative exons that contain a premature termination codon and elicit AS-NMD when included in a transcript. Cancer cells often exploit the inclusion of PEs to downregulate tumor suppressors or the exclusion of PEs to upregulate oncoproteins. Therefore, PEs have drawn significant attention as a novel therapeutic avenue for cancer and other diseases. Here, we examine a therapeutic proof-of-concept for manipulating PE-mediated oncogenic AS-NMD using a CRISPR-based approach. Using paired guide RNA, we successfully deleted a PE of a tumor suppressor (EZH2) from the genome of SRSF2-mutated leukemia. This editing resulted in EZH2 mRNAs without a PE, escaped AS-NMD, and restored the protein expression. This subsequently reinstated H3K27 histone methylation and rescued defective chromatin regulation associated with impaired hematopoietic stem cell differentiation. Finally, we showed the preferential advantages of CRISPR over the antisense technology we recently developed targeting the PE of EZH2. Therefore, the CRISPR strategy shows compelling evidence as a therapeutic approach targeting PE in cancer and other human diseases.

cancer biology↗

bioRxiv2026

Modular Architecture of the SAGA Complex Governs Stress Adaptation, Morphogenesis, and Histone Acetylation in Candida albicans

The SAGA complex is a conserved, multifunctional transcriptional co-activator known for its roles in chromatin modification and transcriptional regulation. While SAGA has been extensively characterized in Saccharomyces cerevisiae and metazoans, its modular organization and functional significance in the major human fungal pathogen Candida albicans remain poorly understood. Through bioinformatic analyses, we found that SAGA subunits are conserved in C. albicans. Genetic disruption of the histone acetyltransferase (HAT; GCN5, ADA2), structural (SPT; SPT7, SPT20, TAF12L), and TATA-binding protein interaction (TBP-interaction; SPT3, SPT8) modules leads to impaired growth under oxidative, metal, and antifungal stress conditions and causes severe defects in filamentation. In contrast, deletion of the deubiquitination (DUB) module components UBP8 and SUS1 results in minimal phenotypic consequences. Strikingly, loss of SGF73, a structural component linking the DUB module to the SAGA core, produces pronounced defects in stress conditions and filamentation, phenocopying SPT3 and SPT8 mutants. Consistent with these observations, filamentation-associated genes are significantly upregulated in SGF73, SPT3 and SPT8 mutants. Notably, these mutants also exhibit elevated global levels of histone H3 lysine-9 acetylation (H3K9ac), suggesting a critical role for SGF73-mediated SAGA integrity in coordinating chromatin acetylation with transcriptional programs governing stress responses and filamentation in C. albicans.

molecular biology↗

bioRxiv2026

Targeting EZH2 Oncogenic Splicing: Decoding the Regulatory Network and Antisense Correction

Recurrent mutations in splicing factors (SFs) have been established as crucial drivers of tumorigenesis in several types of blood cancer, and also common in a variety of solid tumors. Mutations change the RNA-binding preferences of SFs, promote global splicing alterations, and often generate erroneous mRNAs that are then degraded by nonsense-mediated mRNA decay (NMD). Consequently, several critical genes linked to hematopoiesis are dysregulated, leading to blood cancer. Although the field has progressed considerably in identifying aberrant genes and affected pathways, effective therapies have not yet emerged in SF-mutated cancers. To address this key gap, we instigated a gene-specific targeted strategy by unlocking the regulatory network. As a proof-of-concept, we scrutinized a tumor suppressor gene EZH2, which is a bona fide target in SRSF2-mutated cancer. We precisely defined splicing cis-elements in EZH2 transcripts and illustrated the dynamic choreography of regulatory proteins in the entire splicing and NMD catalytic pathways. We uncovered a highly coordinated cross-regulation between splicing and NMD promoted by mutant SRSF2 by enhancing the deposition of critical spliceosome- and NMD-associated factors, augmenting mRNA decay to ablate tumor suppression. We then designed antisense oligonucleotides (ASOs) targeting important regulatory sites. Our lead ASO successfully corrects aberrant splicing and NMD, restores the expression and function of EZH2, and partially rescues hematopoietic defects and cellular properties. Our study demonstrates that ASO pharmacology is an actionable strategy for clinical development, challenging the existing paradigms in SF-mutated cancers.

cancer biology↗

bioRxiv2025

Histone acetylation by SAGA Complex but not by NuA4 Complex is Required for filamentation programme in Candida albicans

Candida albicans, a major human fungal pathogen undergoes filamentation from yeast to hyphal state under filamentation-inducing conditions. Gcn5 and Esa1 are key histone H3 and H4 acetyltransferases, respectively, encoded by the budding yeast and other eukaryotes. While Gcn5, a subunit of the SAGA complex, and Esa1, a subunit of the NuA4 complex are critical for C. albicans virulence and hyphal induction, how the relative HAT activities impinge on hyphal gene expression during filamentation is less understood. We found that hyphal gene promoters are hyperacetylated at H3K9 and H4 upon filamentation. By creating point mutations in the HAT domain of Gcn5 and Esa1, we investigated the relative requirement of the SAGA and NuA4 HAT activities for filamentation response. We show that Gcn5 HAT activity is essential for hyperacetylation of H3K9 and H4 at promoters and across hyphal gene ORFs. Surprisingly, the Esa1 HAT domain mutation did not impair H4 acetylation at hyphal genes suggesting that Gcn5 HAT activity is sufficient for H4 (and H3K9) acetylation. Paradoxically, the Esa1 HAT mutant formed filaments constitutively and showed elevated H3K9ac and H4ac at promoters under inducing conditions. Furthermore, we show that the basic helix-loop-helix transcriptional regulator Efg1 is essential for Gcn5-mediated hyperacetylation and RNA pol II recruitment to promoters. Thus, our results indicate that the SAGA-mediated H3K9 and H4 acetylation is sufficient and essential for induction of C. albicans filamentation.

molecular biology↗

bioRxiv2024

Beyond the Sin3/HDAC Complex: FAM60A emerges as a regulator of RNA Splicing

FAM60A, traditionally linked to chromatin remodeling within the Sin3/HDAC complex, has emerged as a critical regulator in RNA splicing. Employing an integrative approach that combines immunological assays, CRISPR/Cas9 technology, comprehensive genomics, proteomics, and advanced cross-linking mass spectrometry, complemented by sophisticated 3D molecular modeling, our study challenges and extends the existing understanding of FAM60As functional dynamics. Contravening previous perceptions, our findings elucidate that FAM60A does not interact directly with SIN3A, rather establishes direct interactions with SAP30 and HDAC1, redefining its relationship with the Sin3/HDAC complex. These interactions, deciphered through detailed 3D structural analysis supported by cross-linking constraints, signify a complex architectural role of FAM60A within chromatin remodeling processes. Moreover, our research unveils FAM60As pivotal role in RNA processing, particularly in splicing regulation. Through extensive molecular interactions with a diverse array of mRNA-binding proteins and principal spliceosome components, FAM60A emerges as a key regulator of RNA splicing. This expanded role delineates its influence on gene expression regulation, spotlighting its capacity to modulate critical cellular processes. In sum, this study unveils FAM60As key role in gene regulation and RNA splicing, and suggests new paths for cellular and therapeutic research.

molecular biology↗

bioRxiv2023

Sida cordifolia, a medicinal plant is efficacious in models of Huntingtons disease, by reducing ER stress

Background and aimHuntingtons Disease is a severe neurodegenerative disorder caused by misfolded mutant huntingtin proteins with expanded stretches of polyglutamines aggregating and destroying cells in the nervous system. Sida cordifolia and Acorus calamus are medicinal plants used in traditional Ayurvedic medicine to treat neurological disorders. Here, we tested the effectiveness of extracts of both medicinal plants in decreasing aggregation of mutant huntingtin protein in models of Huntingtons Disease and explored the mode of action. Experimental procedureWe used two models, the nematode Caenorhabditis elegans and a transgenic mouse neuroblastoma cell line, both expressing mutant huntingtin proteins with elongated polyglutamines. We assessed the effect of Sida cordifolia and Acorus calamus on mutant huntingtin protein aggregation in both models, and additionally used the cell line for mechanistic studies to identify cellular pathways underlying the effects of treatment. Results and conclusionHere, we show that an extract of Sida cordifolia inhibits aggregation of mutant huntingtin proteins. In the C. elegans model, the extract prolonged life span and improved motility of the nematode by reducing aggregation of the mutant huntingtin protein. Acorus calamus did not exhibit these effects. In the transgenic mouse neuroblastoma cell line, the extract decreased aggregation of the mutant huntingtin protein by suppressing key pathways in the ER stress response caused by the mutant protein. Our results highlight the potential therapeutic value of Sida cordifolia and its promise as a source for novel medications. HighlightsO_LISida cordifolia extract reduces aggregates in HD model of transgenic worms C_LIO_LIReduction in aggregates leads to improved motility and longevity C_LIO_LISida cordifolia extract reduces ER stress in cells expressing mHTT protein C_LIO_LIFirst report on the pharmacology of Sida cordifolia in neurodegeneration C_LI

pharmacology and toxicology↗