HIV-1 and SIV Nef utilize different mechanisms for kinase activation: A novel mechanism for the modulation of PAK/SNTK activation via the acidic domains of HIV-1 and SIV Nef

The Nef protein of the human immunodeficiency viruses (HIV-1 and HIV-2) and simian immunodeficiency virus (SIV) enhances viral replication through its pleiotropic effects, such as downregulation of CD4 and MHC I cell surface receptors from the plasma membrane, enhancement of virion infectivity, and modulation of cellular signaling pathways. In particular, Nef activates a p21-activated kinase (PAK), an interaction that is correlated with induction of high viral loads and disease progression to simian AIDS in rhesus macaques. Nef-mediated PAK activation is a highly conserved function between HIV-1 and SIV. This interaction requires an intact central core domain of Nef. Although this core domain is the most conserved region among Nef isolates, the HIV-1 and SIV Nef cores are not interchangeable. Our findings show that the central core of HIV-1 Nef requires an interaction with the N-terminus, whereas the core domain of SIV Nef utilizes an interaction with the C-terminus. In addition, we have mapped key residues within the core domain that are critical determinants of PAK activation to a contiguous L116WIYHTQ sequence in HIV-1 SF2 Nef and the corresponding I144YLEKEE motif in SIV mac239 Nef. Furthermore, phosphorylation of a serine residue at position 161 within the core of SIVmac239 Nef may play a role in regulating PAK activation. These different requirements for PAK activation suggest that HIV-1 and SIV Nef either utilize different PAK isoforms or they are activated by a common PAK through different adaptor molecules. Recently, our laboratory has also discovered a novel kinase activity that associates with the N-terminus of SIVmac239 Nef. This kinase is designated the SIV Nef N-terminal kinase, or SNTK. Although the identity of SNTK remains unknown, its activation levels are inversely correlated with those of PAK. In addition, we have identified a novel mechanism for the dual modulation of SNTK and PAK activation levels. In particular, the acidic domain of HIV-1 Nef functions to significantly attenuate SNTK activation with a concomitant augmentation of PAK activity. The further elucidation of the mechanism whereby Nef modulates kinase activation levels may reveal the pathway by which Nef enhances viral fitness within the host cell.