Biology subjects

Singh, S. V.

Publications and source records attributed to Singh, S. V..

3 recordsLinked to original sources

bioRxiv2023

Medulloblastoma-associated DDX3X mutants are oncogenic having a defect in translation-promoting activity but functional in stress granule formation and interferon signaling

DDX3X, a DEAD box-containing RNA helicase, is known to play diverse roles in RNA metabolism, stress response, innate immunity, and cancer. Medulloblastoma is the single most common malignant brain tumor in children. DDX3X is recurrently mutated in the WNT and SHH subgroups of medulloblastoma. CRISPR-Cas9 mediated DDX3X knockout was successful in the HEK293FT cells but generated only non-truncating indels in the medulloblastoma cells suggesting DDX3X is necessary for the viability of the cells. Downregulation of DDX3X expression using shRNA also brought about a considerable reduction in proliferation, clonogenic potential, and anchorage-independent growth of the medulloblastoma cells. Thus, DDX3X expression was found to be essential for the survival, growth, and malignant potential of the medulloblastoma cells consistent with the non-truncating nature of medulloblastoma-associated DDX3X mutations. The medulloblastoma-associated DDX3X mutants were found to be defective in their ability to drive the translation of mRNAs with complex 5-UTR that is dependent on the ATP-dependent helicase activity of DDX3X. These helicase defective DDX3X mutants could restore the expression of interferon signaling genes and malignant potential lost upon DDX3X knockdown in medulloblastoma cells. Their N-terminal domain is intact and was found to be functional in stress granule formation. DDX3X mutants upregulated expression of malignancy-related genes suggesting tumor suppressive role for the helicase activity in the medulloblastoma pathogenesis. Inhibitors of the N-terminal domain of DDX3X which is essential for the viability of medulloblastoma cells could have therapeutic potential in the treatment of WNT and SHH subgroup medulloblastomas.

cancer biology↗

bioRxiv2020

Novel mechanistic targets of Forkhead box Q1 transcription factor in human breast cancer cells

The transcription factor forkhead box Q1 (FoxQ1), which is overexpressed in different solid tumors, has emerged as a key player in the pathogenesis of breast cancer by regulating epithelial-mesenchymal transition, maintenance of cancer-stem like cells, and metastasis. However, the mechanism underlying oncogenic function of FoxQ1 is still not fully understood. In this study, we compared the RNA-seq data from FoxQ1 overexpressing SUM159 cells with that of empty vector-transfected control (EV) cells to identify novel mechanistic targets of this transcription factor. Consistent with published results in basal-like subtype, immunohistochemistry revealed upregulation of FoxQ1 protein in luminal-type human breast cancer tissue microarrays when compared to normal mammary tissues. Many previously reported transcriptional targets of FoxQ1 (e.g., E-cadherin, N-cadherin, fibronectin 1, etc.) were verified from the RNA-Seq analysis. FoxQ1 overexpression resulted in downregulation of genes associated with cell cycle checkpoints, M phase, and cellular response to stress/external stimuli as evidenced from the Reactome pathway analysis. Consequently, FoxQ1 overexpression resulted in S, G2M and mitotic arrest in basal-like SUM159 and HMLE cells, but not in luminal-type MCF-7 cells. There were differences in expression of cell cycle-associated proteins between FoxQ1 overexpressing SUM159 and MCF-7 cells. Finally, we show for the first time that FoxQ1 is a direct transcriptional regulator of interleukin (IL)-1, IL-8, and vascular endothelial growth factor in breast cancer cells. Chromatin immunoprecipitation revealed FoxQ1 occupancy at the promoters of IL-1, IL-8, and VEGF. In conclusion, the present study reports novel mechanistic targets of FoxQ1 in human breast cancer cells.

cancer biology↗

bioRxiv2018

Inhibitor of sarco/endoplasmic reticulum calcium-ATPase impairs paramyxovirus replication

Sarco/endoplasmic reticulum calcium-ATPase (SERCA) is a membrane bound cytosolic enzyme that is known to regulate the uptake of calcium into the sarco/endoplasmic reticulum. Herein, we demonstrate for the first time that SERCA can also regulate paramyxovirus [Peste des petits ruminants virus (PPRV) and Newcastle disease virus (NDV)] replication. Treatment of Vero cells with SERCA specific inhibitor (Thapsigargin) at a concentration that is nontoxic to the cells significantly reduced virus replication. Conversely, overexpression of SERCA rescued the inhibitory effect of Thapsigargin on virus replication. PPRV/NDV infection induced SERCA expression in Vero cells which could be blocked by Thapsigargin. With the help of time-of-addition and virus step-specific assays, it was observed that Thapsigargin specifically inhibits viral entry and subcellular localization of the viral proteins. Furthermore, NDV, but not PPRV acquired a significant resistance to Thapsigargin on long-term passage (P=70) in Vero cells. To the best of our knowledge, this is the first report describing virus supportive role of SERCA and a rare report suggesting that viruses may acquire resistance even in the presence of an inhibitor that targets a cellular factor. This study will contribute in understanding paramyxovirus replication and development of antiviral therapeutics using SERCA (host factor) as a candidate drug target.

microbiology↗