The HIV/AIDS epidemic is one of the most important public health problems facing this generation. The failure of recent vaccine trials and growing resistance to anti-retroviral drugs underscores the need for novel therapeutic strategies. Design of such therapies will depend on a detailed understanding of the mechanism of action of the HIV-1 gene products. To further that goal, we have undertaken a detailed investigation of the HIV-1 viral protein R (Vpr). We employed cDNA microarray and antibody array analyses using isogenic virus with or without Vpr to determine the effect on host cellular gene expression. Vpr induced differential regulation of 109 cellular genes representing diverse families of signaling molecules. Two gene products, NHE1 and TNF alpha, were further studied for their potential roles in Vpr-mediated apoptosis. NHE1 expression was decreased by 50% at both the protein and mRNA levels in the presence of Vpr. Vpr-mediated NHE1 downregulation correlated with a dose dependent decrease in intracellular pH as well as a decrease in the active form of the pro-survival kinase Akt. The loss of these anti-apoptotic functions of NHE1 is proposed to contribute to the apoptotic role of Vpr. The pro-inflammatory cytokine TNF alpha may also play a part in Vpr-mediated apoptosis. Macrophages infected with vpr-expressing virus secreted 1.1-8.5 fold more soluble TNF alpha in response to LPS stimulation than their counterparts infected with isogenic virus lacking Vpr expression. Fold upregulation of TNF alpha directly correlated with induction of apoptosis in uninfected lymphocytes, implicating TNF alpha regulation by Vpr in bystander cell death. Two polymorphisms in the TNF alpha promoter, positions -238 and -963, were found at a higher prevalence in donors showing the lowest and highest effect of Vpr on the TNF alpha response. These results suggest that host genetic determinants may affect bystander cell death and thus the course of HIV pathogenesis. Together, the results of this study present a molecular basis for changes induced in the host cell by HIV-1 Vpr and elucidate two potential pathways for the design of anti-retroviral therapeutics targeting HIV-1 Vpr.
| Identifer | oai:union.ndltd.org:PITT/oai:PITTETD:etd-12092005-195447 |
| Date | 16 February 2006 |
| Creators | Janket, Michelle L |
| Contributors | Thomas Smithgall, Ph.D., Jeremy Martinson, Ph.D., Todd A. Reinhart, Sc.D., Velpandi Ayyavoo, Ph.D. |
| 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-12092005-195447/ |
| 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. |
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