Integration of the retroviral genome with host-cell DNA is an essential step in retrovirus replication. The integration reaction is catalyzed by a virus-encoded integrase (IN) which, after reverse transcription of genomic viral RNA, remains associated with the viral cDNA in a high molecular weight nucleoprotein pre-integration complex. Thus, targeting of the viral pre-integration complex to host-cell DNA is dependent upon transport of this complex to the nucleus of the host cell. Our studies provide evidence that nuclear import of the pre-integration complex of HIV-1 is a rapid and active process which is independent of the cell cycle. This active transport process is mediated by the nucleophilic properties of the gag matrix protein (MA p17) which contains a nuclear localization sequence (NLS) at its N terminus, and which is associated with the pre-integration complex of HIV-1. In support of this tenet we provide evidence that peptide analogues containing the NLS of HIV-1 MA p17 prevent establishment of proviral HIV-1 DNA. In addition, HIV-1 variants bearing MA p17 NLS mutations are replication defective. Specifically, we propose to: A.Characterize viral and host cell components of the pre-integration complex of HIV-1 and their role in nuclear import of HIV-1 pre-integration complexes. I. Analyze nucleophilic properties of viral pre-integration complex components. II. Examine role of cellular proteins in nuclear import of HIV-1 pre- integration complexes. III. Analyze replicative and nuclear targeting properties of HIV-1 variants bearing MA p17 NLS mutations. B. Evaluate inhibitory properties of NLS peptide analogues on HIV-1 pre- integration complex transport and virus replication. I. Analyze inhibitory properties of MA p17 NLS peptide analogues on HIV-1 and host cell NLS function in a microinjection assay. II. Evaluate antiviral and cytocidal properties of NLS peptide analogues. It is expected that these studies will provide fundamental insight into critical early events in the life cycle of HIV-1 and provide novel targets for therapy aimed at preventing nuclear localization of HIV-1 pre- integration complexes and establishment of the integrated provirus.
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