Deep sequencing of HIV-1 reverse transcripts reveals the multifaceted anti-viral functions of APOBEC3G

Following cell entry, the RNA genome of HIV-1 is reverse transcribed into double-stranded DNA that ultimately integrates into the host cell genome to establish the provirus. These early phases of infection are notably vulnerable to suppression by a collection of cellular anti-viral effectors, called restriction or resistance factors. The host anti-viral protein APOBEC3G (A3G) antagonizes the early steps of HIV-1 infection through the combined effects of inhibiting viral cDNA production, and cytidine-to-uridine driven hypermutation of this cDNA. In seeking to address the underlying molecular mechanism for inhibited cDNA synthesis, we developed a deep sequencing strategy to characterize nascent reverse transcription products and their precise 3'-termini in HIV-1 infected T cells. Our results demonstrate site- and sequence-independent interference with reverse transcription, which requires A3G’s specific interaction with reverse transcriptase (RT) itself. This approach also established, contrary to current ideas, that cellular uracil base excision repair (UBER) enzymes target and cleave A3G-edited uridine-containing viral cDNA. Together, these findings yield further insights into the regulatory interplay between RT, A3G and the cellular DNA repair machinery, and identify the suppression of HIV-1 RT by a directly interacting host protein as a new cell-mediated anti-viral mechanism.