Nanopore sequencing data for "Single-molecule epitranscriptomic analysis of full-length HIV-1 RNAs reveals functional roles of site-specific m6As" paper. Single-molecule epitranscriptomic analysis of full-length HIV-1 RNAs reveals functional roles of site-specific m6As

Although the significance of chemical modifications on RNA is acknowledged, the evolutionary benefits and specific roles in HIV-1 replication remain elusive. Most studies have provided only population-averaged values of modifications for fragmented RNAs at low resolution and have relied on indirect analyses of phenotypic effects by perturbing host effectors. Here, we analyzed chemical modifications on HIV-1 RNAs at the full-length, single-RNA level, and nucleotide resolution using novel direct RNA sequencing methods. Our data reveal an unexpectedly simple HIV-1 modification landscape, highlighting three predominant N6-methyladenosine (m6A) modifications near the 3' end. More densely installed in spliced viral mRNAs than in genomic RNAs, these m6As play a crucial role in maintaining normal levels of RNA splicing and translation. HIV-1 generates diverse RNA subspecies with distinct m6A ensembles, and maintaining multiple of these m6As on its RNAs provides additional stability and resilience to HIV-1 replication—an unexplored RNA-level evolutionary strategy.