The p400 complex promotes HIV-1 latency by suppressing 1 viral transcription and altering the host cell state [RNA-seq]

Eradicating HIV-1 is complicated by latently infected CD4+T cells harboring repressed HIV-1 proviruses that can reactivate. Using a pooled shRNAmir screen of all human chromatin regulators we identified previously unappreciated factors that govern HIV-1 latency and reactivation. Depletion of EP400 and DMAP1, core subunits of the multifunctional p400 complex, strongly derepressed HIV-1 transcription in Jurkat and primary CD4+T cells. EP400/DMAP1 co-localize with paused RNA Polymerase II (RNAPII) at the transcriptional start sites (TSS) of protein-coding genes, limiting RNAPII pause release, with HIV-1 elongation particularly affected. TNF-⍺ stimulation robustly promotes co-recruitment of RNAPII, EP400, and DMAP1 across the HIV-1 genome, which is distinct from other genes where this co-recruitment occurs mainly at the TSS. Depletion of EP400/DMAP1 reactivated HIV transcription independently of histone variant exchange and KAT5 activity, and increased gene expression of key T cell factors known to trans-activate HIV-1. Thus, the p400 complex is a host restriction factor that blocks RNAPII transcriptional elongation at the HIV-1 locus and creates a CD4+T cell state unfavorable for HIV-1 transcription. J-Lat 10.6 cells, wild-type Jurkat cells and primary CD4+ T cells were transduced with shRNAmirs (two independent shRNAmirs as replicates) targeting negative control genes CD8B and CD19 or experimental targets EP400 and DMAP1. For J-Lat 10.6 and WT uninfected Jurkats, RNA-sequencing was performed on transduced cells at 9 days post-transduction. For the primary CD4+ T cell experiment: cells were transduced with shRNAmirs and infected or not with HIVGKO. 9 days later transduced cells that were either uninfected, latently infected or productively infected were FACS purified and RNA-seq was performed. Comparisons were made between EP400/DMAP1 depletion and control shRNAmirs.