p53-derived host restriction of HIV-1 replication by protein kinase R-mediated tat phosphorylation and inactivation

Tumor suppressor p53 has been suggested to be a host restriction factor against HIV-1 replication, but the detailed molecular mechanism has remained elusive for decades. Here, we demonstrate that p53-mediated HIV-1 suppression is attributed to double- stranded RNA (dsRNA)-dependent protein kinase (PKR)-mediated HIV-1 trans-activator (Tat) phosphorylation and inactivation. p53 silencing significantly enhanced HIV-1 replication in infected cells. Ectopic expression of p53 suppressed Tat activity, which was rescued by PKR silencing. In addition, ectopic expression of PKR abolished Tat activity in p53^-/- and eIF2α^CA cells. Finally, we found that HIV-1 infection activates p53, followed by the induction and activation of PKR. PKR directly interacted with HIV-1 Tat and phosphorylates the first exon of Tat exclusively at five Ser/Thr residues (T23, T40, S46, S62, and S68), which inhibits Tat-mediated provirus transcription in three critical steps: (i) phosphorylation near the arginine-rich motif (ARM) inhibits Tat translocation into the nucleus, (ii) accumulation of Tat phosphorylation abolishes Tat-Tat-responsive region (TAR) binding, and (iii) Tat phosphorylation at T23 and/or T40 obliterates the Tat-cyclin T1 interaction. These five Ser/Thr sites on Tat were highly conserved in HIV-1 strains prevalent in Europe and the United States. Taken together, our findings indicate that p53-derived host restriction of HIV-1 replication is likely attributable, at least in part, to a noncanonical p53/PKR/Tat phosphorylation and inactivation pathway in HIV-1 infection and AIDS pathogenesis.