Biology subjects

Venkat, S.

Publications and source records attributed to Venkat, S..

6 recordsLinked to original sources

bioRxiv2026

Disconnect Between Succinate, Succinyl-CoA, and Histone Succinylation in Cells and Gastrointestinal Stromal Tumors with Succinate Dehydrogenase Deficiency

Background: Succinate dehydrogenase (SDH) is a heterotetrameric enzyme in both the tricarboxylic acid (TCA) cycle and the electron transport chain. SDH-deficiency has been shown to drive the oncogenesis of cancer types including gastrointestinal stromal tumors (GIST), paraganglioma/pheochromocytoma, and clear cell renal cell carcinoma. Previous work on SDH loss has shown increases in succinate, decreases in fumarate, as well as increase in the succinate to -ketoglutarate ratio, and correlated these changes to metabolite signaling and epigenetic dysregulation. Alterations to other TCA intermediates, including succinyl-CoA, the acyl-donor for succinylation, and the resulting cellular effects has not been fully explored. Methods: We used liquid chromatography high resolution mass spectrometry to quantify TCA cycle metabolites across the NCI-60 panel of cell lines, three SDH-competent and SDH-deficient model systems, and human tumor samples. We measured histone post-translational modifications in SHD-deficient and SDH-competent models and examined the correlation and potential pharmacological targeting of described desuccinylase enzymes by protein and cell assays. Results: We found an increase in succinyl-CoA in multiple SDH-deficient models, but no correlation between succinate to fumarate ratio and succinyl-CoA concentrations in the NCI-60. Interestingly, we found that while there was an increase in succinyl-CoA in the SDH-deficient cells, there was no corresponding increase in histone succinylation. We also observed a decrease in lysine malonylation and glutarylation, two other anionic lysine modifications. Cell viability and clonogenic assays revealed very weak or no selective effect of desuccinylase targeting in SDH-deficient models. Similarly, using SDH-deficient versus SDH-competent GIST patient samples, we examined both protein expression and metabolite abundance at the bulk tumor level and found no significant increase in succinyl-CoA or desuccinylase expression, despite the increase in succinate to fumarate ratio. Conclusions: We found that SDH-deficiency led to altered central carbon metabolism including increases to succinate and succinyl-CoA. Despite data that suggested the upregulation of desuccinylases may mediate a disconnect between succinyl-CoA concentration and histone succinylation, we found weak or no reductions in cell viability in response to pharmacological inhibition of desuccinylases. Additionally, no correlation of SDH loss with desuccinylase expression and succinyl-CoA in bulk tumor samples was observed.

cancer biology↗

bioRxiv2026

Developmentally programmed nuclear pore complex replacement enables oocyte specification

Oocytes endow embryos with molecular machinery essential for development, but not all maternal components are inherited indiscriminately. In Drosophila, surveillance pathways eliminate defective mitochondria and aberrant RNAs from the maternal pool. Whether stable nuclear structures, like nuclear pore complexes (NPCs), are similarly curated remains unknown. Here, we uncover a developmentally programmed NPC turnover pathway that renews NPCs during oocyte specification. NPC levels decline through a combination of passive dilution, driven by deferred nucleoporin expression, and active degradation mediated by the ESCRT-III/Vps4 pathway. This clearance is counterbalanced by subsequent de novo NPC synthesis. Failure to turn over NPCs results in aberrantly persistent germ cell gene expression and defective oocyte specification. These findings establish NPC renewal as a critical step in oocyte identity establishment and maternal provisioning.

developmental biology↗

bioRxiv2022

Development of a high-throughput barrier function assay using primary human gut organoids

Disruptions in the gut epithelial barrier can lead to the development of chronic indications such as inflammatory bowel disease (IBD). Historically, barrier function has been assessed in cancer cell lines, which do not contain all human intestinal cell types, leading to poor translatability. To bridge this gap, we adapted human primary gut organoids grown as monolayers to assess barrier function. In this work we describe and characterize a novel 96-well human gut organoid-derived monolayer system that enables medium/high-throughput quantitative assessment of candidate therapeutics. Normal human intestine differentiation patterns and barrier function were characterized and confirmed to recapitulate key aspects of in vivo biology. Next, cellular response to TNF- (a central driver of IBD) was determined using a diverse cadre of readouts.These outputs included transcription factor phosphorylation, target gene expression, cytokine/chemokine production, and barrier function. Additionally, we showed that TNF- pathway antagonists rescued damage caused by TNF- in a dose-dependent manner, indicating that this system is suitable for quantitative assessment of barrier modulating factors. Taken together, we have established a robust primary cell-based 96-well system capable of interrogating questions around mucosal response. This system is well suited to provide pivotal functional data to support translational target and drug discovery efforts.

immunology↗

bioRxiv2022

CPSF3 inhibition blocks pancreatic cancer cell proliferation through disruption of core histone processing

Pancreatic ductal adenocarcinoma (PDAC) is a lethal disease with limited effective treatment options. This potentiates the importance of uncovering novel drug targets. We have discovered global dysregulation of the gene regulatory process alternative polyadenylation (APA) in PDAC. APA is a pre-mRNA processing mechanism that generates mRNAs with distinct 3 ends, impacting gene expression and protein function. We revealed that APA dysregulation in PDAC drives oncogenic signatures and predicts poor patient outcome. As APA directs widespread gene expression dysregulation across the PDAC patient population, we hypothesized that inhibition of APA has therapeutic potential. APA is controlled by a complex of proteins, including cleavage and polyadenylation specificity factor 3 (CPSF3). CPSF3 is the endonuclease catalyzing mRNA cleavage, and a potentially druggable target. We now find that CPSF3 is highly expressed and associated with poor prognosis in PDAC patients. CPSF3 knockdown decreases PDAC proliferation and clonogenicity in vitro and tumor growth in vivo. We demonstrate that CPSF3 knockdown induces widespread APA alterations of oncogenes and tumor suppressors, and determine the contribution of one of these events to CPSF3-induced cell proliferation phenotype. Furthermore, we find that PDAC, but not non-transformed pancreatic cells, are sensitive to the CPSF3 small molecule inhibitor JTE-607. Mechanistically, JTE-607 impairs replication-dependent histone processing, disrupting nucleosome assembly and destabilizing chromatin structure. Finally, we determine that JTE-607 attenuates cell proliferation by arresting cells in early S-phase of the cell cycle. Altogether, we identify CPSF3 as a druggable target in PDAC and reveal novel mechanisms by which CPSF3 controls cancer cell growth. SignificanceThis work identifies CPSF3 as a potential drug target in pancreatic ductal adenocarcinoma and reveals new mechanisms by which CPSF3 inhibition attenuates PDAC cell proliferation through modulating alternative polyadenylation and histone processing.

cancer biology↗

bioRxiv2021

Alternative polyadenylation characterizes epithelial and fibroblast phenotypic heterogeneity in pancreatic ductal adenocarcinoma

Human tumors are characterized by extensive intratumoral transcriptional variability within the cancer cell and stromal compartments. This variation drives phenotypic heterogeneity, producing cell states with differential pro- and anti-tumorigenic properties. While bulk RNA sequencing cannot achieve cell type specific transcriptional granularity, single cell sequencing has permitted an unprecedented view of these cell states. Despite this knowledge, we lack an understanding of the mechanistic drivers of this transcriptional and phenotypic heterogeneity. 3 untranslated region alternative polyadenylation (3 UTR-APA) drives gene expression alterations through regulation of 3 UTR length. These 3 UTR alterations modulate mRNA stability, protein expression and protein localization, resulting in cellular phenotypes including differentiation, cell proliferation, and migration. Therefore, we sought to determine whether 3 UTR-APA events could characterize phenotypic heterogeneity of tumor cell states. Here we analyze the largest single cell human pancreatic ductal adenocarcinoma (PDAC) dataset and resolve 3 UTR-APA patterns across PDAC cell states. We find that increased proximal 3 UTR-APA is associated with PDAC progression and characterizes a metastatic ductal epithelial subpopulation and an inflammatory fibroblast population. Furthermore, we find significant 3 UTR shortening events in cell state-specific marker genes associated with increased expression. Therefore, we propose that 3 UTR-APA drives phenotypic heterogeneity in cancer.

cancer biology↗

bioRxiv2019

Alternative polyadenylation drives oncogenic gene expression in pancreatic ductal adenocarcinoma

Alternative polyadenylation (APA) is a gene regulatory process that dictates mRNA 3-UTR length, resulting in changes in mRNA stability and localization. APA is frequently disrupted in cancer and promotes tumorigenesis through altered expression of oncogenes and tumor suppressors. Pan-cancer analyses have revealed common APA events across the tumor landscape; however, little is known about tumor type-specific alterations that may uncover novel events and vulnerabilities. Here we integrate RNA-sequencing data from the Genotype-Tissue Expression (GTEx) project and The Cancer Genome Atlas (TCGA) to comprehensively analyze APA events in 148 pancreatic ductal adenocarcinomas (PDAs). We report widespread, recurrent and functionally relevant 3-UTR alterations associated with gene expression changes of known and newly identified PDA growth-promoting genes and experimentally validate the effects of these APA events on expression. We find enrichment for APA events in genes associated with known PDA pathways, loss of tumor-suppressive miRNA binding sites, and increased heterogeneity in 3-UTR forms of metabolic genes. Survival analyses reveal a subset of 3-UTR alterations that independently characterize a poor prognostic cohort among PDA patients. Finally, we identify and validate the casein kinase CK1 as an APA-regulated therapeutic target in PDA. Knockdown or pharmacological inhibition of CK1 attenuates PDA cell proliferation and clonogenic growth. Our single-cancer analysis reveals APA as an underappreciated driver of pro-tumorigenic gene expression in PDA via the loss of miRNA regulation.

cancer biology↗