YTHDF1 and YTHDC1 m6A reader proteins regulate HTLV-1taxandhbzactivity

Human T-cell leukemia virus type 1 (HTLV-1) is a retrovirus responsible for adult T-cell leukemia/lymphoma (ATLL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), a progressive neurodegenerative disease. Regulation of viral gene expression plays a key role in viral persistence and pathogenesis. However, the molecular mechanisms underlying this fine-tuned regulation remain poorly understood. Little is known regarding RNA chemical modifications of HTLV-1 RNA and how these affect viral biology and disease development. Posttranscriptional chemical modification of RNA is common in eukaryotes, with N 6 -methyladenosine (m 6 A) being the most prevalent. In this study, we investigated the role of m 6 A RNA modifications on HTLV-1 gene expression. Using MeRIP-Seq, we mapped sites of m 6 A modification to the 3’ end of the viral genome. We found HTLV-1 RNA, as well as viral oncogene transcripts tax and hbz , contained m 6 A modifications. m 6 A-depletion in HTLV-1-transformed cells decreased sense-derived viral genes ( Tax, Gag, Env ) and increased antisense-derived Hbz expression. Tax and hbz transcripts were bound by reader proteins YTHDF1 and YTHDC1 in a panel of HTLV-1 T-cell lines. Using expression vectors and shRNA-mediated knockdown, we found YTHDF1 had opposing effects on viral gene expression – decreasing sense-derived viral genes and increasing antisense-derived Hbz . Upon further study, the YTHDF1 effects on tax abundance were dependent on tax m 6 A deposition. The nuclear m 6 A reader protein YTHDC1 affected the abundance of both sense- and antisense-derived viral transcripts and specifically enhanced the nuclear export of tax transcript. Collectively, our results demonstrate global m 6 A levels and m 6 A reader proteins YTHDF1 and YTHDC1 regulate HTLV-1 gene expression. Importance Human T-cell leukemia virus type 1 (HTLV-1) persistence and pathogenesis are controlled through tight regulation of viral gene expression. The fate of RNA can be controlled by epigenetic modifications which impact gene expression without altering the DNA sequence. Our study details the impact of N6-methyladenosine (m 6 A) RNA chemical modifications on HTLV-1 gene expression. We found reductions in global m 6 A levels affected viral gene expression, decreasing Tax and other sense-derived viral genes, while increasing the antisense-derived Hbz . Our results suggest the oncogenic viral transcripts, tax and hbz , are m 6 A-modified in cells. We found these viral RNA modifications are interpreted by reader proteins YTHDF1 and YTHDC1, which dictate the fate of the viral RNA. Understanding HTLV-1 RNA chemical modifications offers potential insights into novel therapeutic strategies for HTLV-1-associated diseases.