MITOGEN-ACTIVATED PROTEIN-KINASE PHOSPHORYLATES AND REGULATES THE HIV-1 VIF PROTEIN

The human immunodeficiency virus type 1 (HIV-1) Vif protein plays a cr itical role in virus replication and infectivity. Here we show that Vi f is phosphorylated and regulated by p44/42 mitogen-activated protein kinase (MAPK). Vif phosphorylation by MAPK was demonstrated in vitro a s well as in vivo and was shown to occur on serine and threonine resid ues. Two-dimensional tryptic phosphopeptide mapping indicated that Vif is phosphorylated by MAPK on the same sites in vitro and in vivo. Rad ioactive peptide sequencing identified two phosphorylation sites, Thr( 96) and Ser(165). These phosphorylation sites do not correspond to the known optimum consensus sequences for phosphorylation by MAPK (PX(S/T )P) nor to the minimum consensus sequence ((S/ T)P), indicating that M APK can phosphorylate proteins at sites other than those containing th e PX(SPT)P or (S/T)P motifs. Synthetic Vif peptides corresponding to t he local sequences of the phosphorylation sites were not phosphorylate d by MAPK, suggesting that recognition of these sites by MAPK is likel y to require structural determinants outside the phosphorylation site. Mutations of the Thr96 site, which is conserved among Vif sequences f rom HIV-1, HIV-2, and SIV, resulted in significant loss of Vif activit y and inhibition of HIV-1 replication. These results suggest that MAPK plays a direct role in regulating HIV-1 replication and infectivity b y phosphorylating Vif and identify a novel mechanism for activation of HIV-1 replication by mitogens and other extracellular stimuli.