The nef gene of HIV encodes a small myristylated protein that is required for viral replication and pathogenesis. Nef lacks intrinsic catalytic activity and has been postulated to function through binding to cellular proteins and altering signaling pathways. Nef binds to the macrophage-specific Src family member Hck through its SH3 domain with the highest affinity reported for an SH3-mediated protein-protein interaction. Studies presented in this dissertation characterized the interface constituting this unusually strong interaction by comparing the ability of different Nef alleles to bind and activate Hck in cells. By aid of molecular modeling and mutational analysis, we found the residues in the Nef hydrophobic pocket critically contributing to high affinity interaction with Hck. This is the first study to show that the HIV-1 Nef hydrophobic pocket is critical for SH3-mediated Hck activation in vivo and identified the pocket as a rational target for drug design to selectively disrupt Nef-Hck signaling in HIV-infected cells. We also investigated the role of HIV Nef in survival signaling in myeloid cells, which is a more relevant cell type for HIV infection. Recently, Nef has been demonstrated to be an important factor in promoting the survival of HIV-infected T cells. Since macrophages serve as HIV viral reservoirs, suppression of apoptosis of infected macrophages could have more impact on persistent virus infection in the host. In this study, we demonstrate that Nef protects the macrophage precursor cell line, TF-1 from apoptosis by upregulating the anti-apoptotic gene, Bcl-XL. In addition, the survival signal and Bcl-XL induction by Nef require Erk MAPK activation. This study provides the first evidence that Nef generates anti-apoptosis signals in cells of the myelomonocytic lineage and adds important evidence to the hypothesis that Nef may contribute to the establishment and maintenance of an HIV reservoir by conferring a survival advantage on HIV-infected macrophages
| Identifer | oai:union.ndltd.org:PITT/oai:PITTETD:etd-10252004-162624 |
| Date | 28 October 2004 |
| Creators | Choi, Hyun-Jung |
| Contributors | Thomas E. Smithgall, PhD, Martin C. Schmidt, PhD, Edward Prochownik, MD, PhD, Toshiaki Kodama, DvM, PhD, Todd Reinhart, ScD, Baskaran Rajasekaran, PhD |
| Publisher | University of Pittsburgh |
| Source Sets | University of Pittsburgh |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.library.pitt.edu/ETD/available/etd-10252004-162624/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to University of Pittsburgh or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
Page generated in 0.0028 seconds