N6-Methyladenosine-binding proteins suppress HIV-1 infectivity and viral production.
The internal N6-methyladenosine (m6A) modification of cellular mRNA regulates post-transcriptional gene expression. The YTH domain family proteins (YTHDF1-3 or Y1-3) bind to m6A-modified cellular mRNAs and modulate their metabolism and processing, thereby affecting cellular protein translation. We previously reported that HIV-1 RNA contains the m6A modification and that Y1-3 proteins inhibit HIV-1 infection by decreasing HIV-1 reverse transcription activity. Here, we investigated the mechanisms of Y1-3-mediated inhibition of HIV-1 infection in target cells and the effect of Y1-3 on viral production levels in virus-producing cells. We found that Y1-3 protein overexpression in HIV-1 target cells decreases viral genomic RNA (gRNA) levels and inhibits both early and late reverse transcription. Purified recombinant Y1-3 proteins preferentially bound to the m6A-modified 5' leader sequence of gRNA compared with its unmodified RNA counterpart, consistent with the strong binding of Y1-3 proteins to HIV-1 gRNA in infected cells. HIV-1 mutants with two altered m6A modification sites in the 5' leader sequence of gRNA exhibited significantly lower infectivity than WT, replication-competent HIV-1, confirming that these sites alter viral infection. HIV-1 produced from cells in which endogenous Y1, Y3, or Y1-3 proteins were knocked down singly or together had increased viral infectivity compared with HIV-1 produced in control cells. Interestingly, we found that Y1-3 proteins and HIV-1 Gag protein formed a complex with RNA in HIV-1-producing cells. Overall, these results indicate that Y1-3 proteins inhibit HIV-1 infection and provide new insights into the mechanisms by which the m6A modification of HIV-1 RNA affects viral replication.
['Lu|Wuxun|W|', 'Tirumuru|Nagaraja|N|', 'St Gelais|Corine|C|', 'Koneru|Pratibha C|PC|', 'Liu|Chang|C|', 'Kvaratskhelia|Mamuka|M|', 'He|Chuan|C|', 'Wu|Li|L|0000-0002-5468-2487']
D000241:Adenosine D015683:Gene Products, gag D015658:HIV Infections D015497:HIV-1 D006367:HeLa Cells D006801:Humans D011485:Protein Binding D012367:RNA, Viral D016601:RNA-Binding Proteins D014771:Virion D053586:Virus Internalization
HIV-1 infection infection N6-methyladenosine reverse transcription HIV-1 production RNA m6A modification RNA modification RNA-binding protein YTH domain family proteins (YTHDF1–3), Y1–3 protein human immunodeficiency virus (HIV) m6A reader protein m6A-binding proteins protein expression