HIV-1 m6A methylome investigation by m6A-SAC-seq [CD4+ T cells]

N6-methyladenosine (m6A) is the most prevalent cellular mRNA modification and plays an important role in RNA stability, localization, and gene expression. There are several papers reported that HIV-1 infection upregulates cellular RNA m6A in CD4+ T cells, and this upregulation occurs without changes to the expression levels of the cellular enzymes which add or remove m6A. However, the mechanism of HIV-1 inducing cellular mRNA m6A upregulation is unknown. Here we show that HIV-1 infection upregulates m6A of cellular mRNA and promotes METTL3/METTL14 interaction. We found that HIV-1 infection resulted in increased cellular mRNA m6A level in activated primary CD4+ T cells detected by m6A dot blot and m6A ELISA. We found that HIV-1 infection of CD4+ T cells promotes METTL3/METTL14 interaction compared to mock control using co-immunoprecipitation (co-IP). Our ongoing studies will determine the transcriptomic, epitranscriptomic, and proteomic profiles of CD4+ T cells infected with HIV-1 for 4 days. These studies will facilitate the elucidation of the mechanisms by which HIV-1 infection leads to increased levels of cellular m6A and methyltransferase writers comlex. Overall design: Primary activated CD4+ T cells were either infected with Mock or HIV-1NL4-3 for 96 hours and total RNA was isolated from three individual healthy donors. Poly-A RNA was enriched using oligodT beads and used for m6A-SAC-seq and RNA-sequencing.