Biology subjects

Thombare, K.

Publications and source records attributed to Thombare, K..

3 recordsLinked to original sources

bioRxiv2023

METTL3/MYCN cooperation drives m6A modification during trunk neural crest differentiation and represents a therapeutic vulnerability in MYCN-amplified neuroblastoma

Neuroblastoma (NB) is the most common extracranial childhood cancer, caused by the improper differentiation of developing trunk neural crest cells (tNCC) in the sympathetic nervous system. The N6-methyladenosine (m6A) epitranscriptomic modification controls post-transcriptional gene expression but the mechanism by which the m6A methyltransferase complex METTL3/METTL14/WTAP is recruited to specific loci remains to be fully characterized. We explored whether the m6A epitranscriptome could fine-tune gene regulation in migrating/differentiating tNCC. We demonstrate that the m6A modification regulates the expression of HOX genes in tNCC, thereby contributing to their timely differentiation into sympathetic neurons. Furthermore, we show that posterior HOX genes are m6A modified in MYCN-amplified NB with reduced expression. In addition, we provide evidence that sustained overexpression of the MYCN oncogene in tNCC drives METTL3 recruitment to a specific subset of genes including posterior HOX genes creating an undifferentiated state. Moreover, METTL3 depletion/inhibition induces DNA damage and differentiation of MYCN overexpressing cells and increases vulnerability to chemotherapeutic drugs in MYCN-amplified patient-derived xenografts (PDX) cells, suggesting METTL3 inhibition could be a potential therapeutic approach for NB.

cancer biology↗

bioRxiv2022

m6A modification of TERRA RNA is required for telomere maintenance and is a therapeutic target for ALT positive Neuroblastoma

Telomerase-negative tumors maintain telomere length by alternative lengthening of telomeres (ALT), but the underlying mechanism behind ALT remains poorly understood. A proportion of aggressive neuroblastoma (NB), particularly relapsed tumors, are positive for ALT (ALT+), suggesting that a better dissection of the ALT mechanism could lead to novel therapeutic opportunities. TERRA, a long non-coding RNA (lncRNA) derived from telomere ends, localizes to telomeres in a R-loop-dependent manner and plays a crucial role in telomere maintenance. Here we present evidence that RNA modification at the N6 position of internal adenosine (m6A) in TERRA by the methyltransferase METTL3 is essential for telomere maintenance in ALT+ cells, and the loss of TERRA m6A/METTL3 results in telomere damage. We observed that m6A modification is abundant in R-loop enriched TERRA, and the m6A-mediated recruitment of hnRNPA2B1 to TERRA is critical for R-loop formation. Our findings suggest that m6A drives telomere targeting of TERRA via R-loops, and this m6A-mediated R-loop formation could be a widespread mechanism employed by other chromatin-interacting lncRNAs. Furthermore, treatment of ALT+ NB cells with a METTL3 inhibitor resulted in compromised telomere targeting of TERRA and accumulation of DNA damage at telomeres, indicating that METTL3 inhibition may represent a therapeutic approach for ALT+ NB.

cancer biology↗

bioRxiv2022

Global loss of cellular m6A RNA methylation following infection with different SARS-CoV-2 variants

Host-viral interactions during SARS-CoV-2 infection are needed to understand COVID-19 pathogenesis and may help to guide the design of novel antiviral therapeutics. N6-methyladenosine modification (m6A), one of the most abundant cellular RNA modifications, regulates key processes in RNA metabolism during a stress response. Gene expression profiles observed post-infection with different SARS-CoV-2 variants show changes in the expression of genes related to RNA catabolism, including m6A readers and erasers. We found that infection with SARS-CoV-2 variants caused a loss of m6A in cellular RNAs, whereas m6A was detected abundantly in viral RNA. METTL3, the m6A methyltransferase, showed an unusual cytoplasmic localization post-infection. The B.1.351 variant had a less pronounced effect on METTL3 localization and loss of m6A than the B.1 and B.1.1.7 variants. We also observed a loss of m6A upon SARS-CoV-2 infection in air/liquid interface cultures of human airway epithelia, confirming that m6A loss is characteristic of SARS-CoV-2 infected cells. Further, transcripts with m6A modification were preferentially down-regulated post-infection. Inhibition of the export protein XPO1 resulted in the restoration of METTL3 localization, recovery of m6A on cellular RNA, and increased mRNA expression. Stress granule formation, which was compromised by SARS-CoV-2 infection, was restored by XPO1 inhibition and accompanied by a reduced viral infection in vitro. Together, our study elucidates how SARS-CoV-2 inhibits the stress response and perturbs cellular gene expression in an m6A-dependent manner.

molecular biology↗