12th Conference on Retroviruses and Opportunistic Infections Boston, Massachusetts - February 22-25, 2005

Conf Retrovir Opportunistic Infect 2005 Feb 22-25;12:abstract no. 37LB

Deborah Nguyen, K Wolff, H Yin, J Caldwell, and K Kuhen
Genomics Inst. of the Novartis Res. Fndn., San Diego, CA, USA

BACKGROUND: The HIV Nef accessory factor binds to and interacts with many host signaling molecules, hijacking the pathways to augment HIV replication. Immunoprecipitation of Nef from cells has been shown to precipitate a member of the p21-activated kinase (PAK) family whose activation leads to enhancement of viral transcription from the LTR. Group I PAK are involved in cytoskeletal reorganization, cell morphology and motility, and apoptosis. They have also been shown to activate JNK, ERK, and NFaT pathways in T cells following TCR stimulation. Early evidence suggested that the Nef-associated PAK was either PAK1 or PAK2. While the evidence in support of PAK2 is strong, no studies on the effects of blocking endogenous PAK protein production on HIV infection have been published.

METHODS: We performed a sub-genomic siRNA screen for host proteins involved in HIV infection in HeLaCD4ßgal cells which showed that inhibiting group I PAK kinase expression had a negative impact on infection. We further investigated the role of the group I PAK in HIV infection utilizing validated siRNA in multiple cell types as well as probing the effects of PAK knock-down on Tat-dependent transcription and subsequent virion infectivity.

RESULTS: Contrary to previous studies, we found that inhibition of PAK1 expression using siRNA significantly inhibited HIV while PAK2 depletion had no effect. This pattern was also seen in studies in the Jurkat T-cell line. Mechanistic studies showed an inhibitory effect of PAK1 knock-down on LTR-dependent transcription as well as in other promoter systems, indicating a possible common pathway of transcriptional repression. As seen previously with Nef mutants defective in kinase association, virions produced in cells treated with PAK1 siRNA had diminished infectivity in subsequent cell targets.

CONCLUSIONS: Together, these studies argue that PAK1 rather than PAK2 is the dominant PAK involved in HIV infection, and that PAK1 is involved in multiple stages of HIV infection. Targeting of PAK kinases may represent a novel strategy for development of anti-viral therapeutics.

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