Hiv-1 Infection

HIV-1 Gag protein[Protein]

miR-17[miRNA]

NA[mRNA]

Hek293 Cells Line

Human immunodeficiency virus 1(HIV-1)

Proc Natl Acad Sci U S A 2014 Jul 1 111, E2676-83 doi:10.1073/pnas.1408037111 PMID:24938790

MicroRNA binding to the HIV-1 Gag protein inhibits Gag assembly and virus production

Western blot;qRT-PCR;luciferase reporter assays;RNA immunoprecipitation

Recently, miRNAs have been shown to also interact with proteins outside RISCs, impacting cellular processes through mechanisms not involving gene silencing. Here, we define a previously unappreciated activity of miRNAs in inhibiting RNA-protein interactions that in the context of HIV-1 biology blocks HIV virus budding and reduces virus infectivity. This occurs by miRNA binding to the nucleocapsid domain of the Gag protein, the main structural component of HIV-1 virions. The resulting miRNA-Gag complexes interfere with viral-RNA-mediated Gag assembly and viral budding at the plasma membrane, with imperfectly assembled Gag complexes endocytosed and delivered to lysosomes. The blockade of virus production by miRNA is reversed by adding the miRNA's target mRNA and stimulated by depleting Argonaute-2, suggesting that when miRNAs are not mediating gene silencing, they can block HIV-1 production through disruption of Gag assembly on membranes. This prompted us to examine whether Gag could bind to endogenous miRNAs,not just exogenous miRNAs.Supporting this hypothesis, Gag-miRNA immunoprecipitation experiments revealed that in addition to miRNA-146a,other host miRNA species coimmunoprecipitated wit hGag,including miR-17,miR-19,and miR-16,consistent with previous work showing that Gagcannonspecifically bind to nucleicacids(20). Overall, our findings have significant implications for understanding how cells modulate HIV-1 infection by miRNA expression and raise the possibility that miRNAs can function to disrupt RNA-mediated protein assembly processes in other cellular contexts.