The interaction between HIV-1 Nef and adaptor protein-2 reduces Nef-mediated CD4⁺ T cell apoptosis

Acquired Immune Deficiency Syndrome is characterized by a decline in CD4⁺ T cells. Here, we elucidated the mechanism underlying apoptosis in Human Immunodeficiency Virus-1 (HIV-1) infection by examining host apoptotic pathways hijacked by the HIV-1 Nef protein in the CD4⁺ T-cell line Sup-T1. Using a panel of Nef mutants unable to bind specific host proteins we uncovered that Nef generates pro- and anti-apoptotic signals. Apoptosis increased upon mutating the motifs involved in the interaction of Nef:AP-1 (Nef_M20A or Nef_{EEEE62-65AAAA}) or Nef:AP-2 (Nef_LL164/165AA), implying these interactions limit Nef-mediated apoptosis. In contrast, disrupting the Nef:PAK2 interaction motifs (Nef_H89A or Nef_F191A) reduced apoptosis. To validate further, apoptosis was measured after short-hairpin RNA knock-down of AP-1, AP-2 and PAK2. AP-2α depletion enhanced apoptosis, demonstrating that disrupting the Nef:AP-2α interaction limits Nef-mediated apoptosis. Collectively, we describe a mechanism by which HIV-1 regulates cell survival and demonstrate the consequence of interfering with Nef:host protein interactions.