Replication and Evolution of Primate Lentiviruses - PROJECT SUMMARY Nup358 is a major component of the nuclear pore complex, and forms the long filaments that extend from the nuclear pore rim into the cytoplasm. Nup358 comprises four RAN-binding domains (RBD1-4) separated by several unique domains. HIV-1 capsid (CA) binds Nup358 via its C-terminal cyclophilin-homology domain (CHD). Our comparative analysis of diverse primate lentiviruses and cognate Nup358 homologs from reservoir and spillover hosts revealed a previously unknown effect of the fourth RAN-binding domain (RBD4) on specificity of CA interactions with the CHD. However, this effect is consistent with an earlier literature reporting an effect of the RBD4 on interactions with cellular substrates. Preliminary results show that RBD4-dependent interactions with the CHD affect nuclear import and infectivity. The most striking example involves HIV-2, which had to adapt to differences between the sooty mangabey reservoir host and human Nup358 during emergence in humans. Relevance of the CA-CHD interaction was further underscored by the discovery that it is more conserved among primate lentiviruses than previously reported. The exceptions were viruses of the SIVmac lineage: these lost the CA-CHD interaction during adaptation to macaque hosts, most likely as an evolutionary trade-off that allowed these viruses to evade restriction by macaque TRIM5a and TRIMCyp. The lack of this otherwise conserved interaction with Nup358 may be a critical difference between HIV-1 infection in humans and the SIVmac/SHIV animal models widely used in preclinical HIV/AIDS research. We will use a comparative approach to explore the molecular level details of the conserved interactions between various lentivirus capsids and the RBD4-CHD didomain of the relevant Nup358 homologs, and to determine the basis for the RBD4-dependent effects on CA-CHD binding specificity. The major innovation of this proposal is to leverage naturally occurring variations in 1) lentiviruses CAs and 2) primate Nup358 homologs, which allows us to precisely manipulate virus-host interactions in cell culture, while avoiding the pleiotropic complications associated with the more drastic approaches used to date (e.g. deletions in Nup358 and whole gene knockdowns or knockouts). Our goal is to produce a working model of the interactions between lentiviruses and Nup358, including both conserved and virus-specific elements, and to understand how these interactions contribute to nuclear docking, nuclear transport, integration site selection, and evasion of innate immune responses. This will also help us to understand how the RBD4 determines specificity of CHD-CA interactions. Finally, our results may expose potential differences between HIV-1 and HIV-2 infections in humans and experimental infection of macaques with SIVmac and SHIVs.
