CD4+ T cells and macrophages are the primary targets for primate lentivirus infection. Replication of HIV and SIV within macrophages may play a fundamental role in virus transmission, in the sequestration of virus in tissues, and in maintenance of viral burden. Macrophages are terminally differentiated and non-dividing cells. Although onco- retroviruses such as MLV require host-cell mitosis for nuclear localization of viral DNA and provirus establishment, lentiviruses such as HIV have evolved a specific mechanism which allows provirus establishment in non-dividing cells. This property of HIV is governed by the nucleophilic virion proteins gag matrix (MA) and vpr, which are essential for nuclear localization of viral DNA in non-dividing cells and for infection of macrophages. In addition to its nuclear import function, gag MA is myristoylated; a modification which is necessary for membrane targeting of gag precursors during virus assembly. Thus, there exists a mechanism which allows the opposing membrane targeting and nuclear targeting functions of gag MA to operate independently during virus assembly and virus infection. We have now determined that phosphorylation of gag MA prior to infection is necessary to overcome membrane attachment of gag and allow the nuclear import functions of gag MA to operate. Inhibition of gag MA phosphorylation prevents its nuclear import and attenuates virus infectivity. The requirement for gag MA phosphorylation in virus infection is underscored by our finding that a cellular serine/threonine kinase is packaged within virions of HIV-1. These results reveal a novel level of regulation of primate lentivirus infectivity. The object of this continuation is to: 1. Identify residues in gag MA which are phosphorylated during HIV-1 infection and construct molecular clones of HIV-1 and SIV containing mutations at these specific residues. 2. Define the role of gag MA phosphorylation in its nuclear import function and characterize infectivity of wild type and non-phosphorylated gag MA its variants of HIV and SIV in dividing and non-dividing cell systems. 3. Characterize the serine/threonine kinase that is packaged within virions of HIV and examine effects of tyrosine and serine/threonine kinase inhibitors on HIV infectivity. 4. Examine pathogenic potential of SIVagm gag MA nuclear import mutants in pig-tailed macaques.
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