N6-methyladenosine modification of HCMV IE1 transcript promotes the repressive state of viral genome to achieve latent infection.

Human cytomegalovirus (HCMV) is a prevalent pathogen that chronically infects the majority of human population. Among the many features that allow such widespread HCMV infection, one is its ability to maintain a transcriptionally dormant immune-evasive state called latency by suppressing its own major immediate early promoter (MIEP) via epigenetic alterations. In this study, we show a mechanism of MIEP regulation in which the major immediate early (MIE) gene product, immediate early 1 (IE1) transcript, downregulates its own promoter activity in an m6A modification-dependent manner. We found that the loss of the m6A writer, METTL3, in host cells impedes latency establishment in these cells. Through transcriptome-wide m6A profiling of latently infected monocytes, we identified that the major immediate early gene product IE1 transcript is m6A-modified during latent infection. Using IE1-specific m6A-abolished mutants, we found that m6A modification of the IE1 transcript was necessary for the efficient repression of MIEP, and these mutant viruses exhibited a significant defect in establishing latency and progressed toward lytic-like infection in the human monocytic cell line (THP-1) and primary CD14+ monocytes. Our findings demonstrate that HCMV exploits the host m6A machinery to suppress its own lytic program to establish latency and uncover an unexpected role of immediate early gene messenger RNA (mRNA) in regulating its own expression.