The disappointing results of the recent Merck vaccine clinical phase IIb trials, which utilized adenovirus-based HIV-1 candidates capable of generating an anti-HIV CTL response (HIV vaccine failure prompts Merck to halt trial. Nature 449, 390 (2007), www.hvtn.org/science/1107.html) highlights the importance of understanding the mechanisms through which HIV successfully evades the anti-HIV CTL response. In addition, the inability of current therapies to cure HIV disease underlines the importance of taking new approaches at targeting HIV. Existing drugs effectively prevent new infections, however none of the currently available anti- HIV therapies aid in the eradication of pre-existing infected cells. Thus, it is more clear than ever before, that we need to fully understand the viral mechanisms for immune escape to eradicate infected cells, cure disease and prevent new infection. To this end, the HIV Nef protein is an important and under-developed drug target. HIV-1 Nef has been shown to protect infected primary T cells from CTL recognition and killing in vitro by downmodulating MHC-I protein [7]. There is also substantial in vivo evidence that MHC-I downmodulation is critically important in SIV-infected monkeys for HIV disease pathogenesis [8, 9]. Over the past two funding periods of this grant, we have generated substantial evidence indicating that Nef-mediated MHC-I downmodulation occurs in the following manner: Nef binds to immature forms of MHC-I by targeting hypophosphorylated MHC-I cytoplasmic tail domains in the ER/early Golgi. Binding prevents phosphorylation of the MHC-I cytoplasmic tail and disrupts MHC-I transport from the trans-Golgi network (TGN) to the cell surface. MHC-I is then targeted for degradation in lysosomes. Targeting of MHC-I into the endo-lysosomal pathway from the TGN requires the activity of two cellular co-factors, AP-1 and 2-COP. These cellular trafficking proteins bind the Nef-MHC-I complex at distinct sub-cellular locations and promote targeting into the endosomal and lysosomal pathways respectively. Moreover, we now have a detailed understanding of which amino acids are necessary to form the Nef-MHC-I-AP-1 three-way complex. In the up-coming funding period, we will continue to uncover the mechanism by which Nef functions by determining how Nef is able to bypass the normal cellular regulation of these molecules. In addition, we will begin to apply our assay systems to other Nef targets to compare and contrast the manner in which Nef affects other cellular pathways. Finally, we will demonstrate to what extent these Nef activities are generalized across HIV subtypes.

Public Health Relevance

The recently reported failure of the Merck vaccine trial, which utilized antigens capable of generating an anti-HIV immune response, highlights the importance of understanding the mechanisms through which HIV successfully evades the immune response. In addition, the inability of current therapies to cure HIV disease underlines the importance of taking new tacks at targeting HIV. Existing drugs effectively prevent new infections, however none of the currently available anti-HIV therapies aid in the eradication of pre-existing infected cells. This grant proposal aims to more fully understand the viral mechanisms for immune escape and viral persistence in order to eradicate infected cells, cure disease and prevent new infection.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI046998-14
Application #
8444385
Study Section
Special Emphasis Panel (ZRG1-AARR-H (02))
Program Officer
Stansell, Elizabeth H
Project Start
1999-12-01
Project End
2014-03-31
Budget Start
2013-04-01
Budget End
2014-03-31
Support Year
14
Fiscal Year
2013
Total Cost
$355,851
Indirect Cost
$125,527
Name
University of Michigan Ann Arbor
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
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Kulpa, Deanna A; Del Cid, Natasha; Peterson, Kirsten A et al. (2013) Adaptor protein 1 promotes cross-presentation through the same tyrosine signal in major histocompatibility complex class I as that targeted by HIV-1. J Virol 87:8085-98
Wonderlich, Elizabeth R; Leonard, Jolie A; Kulpa, Deanna A et al. (2011) ADP ribosylation factor 1 activity is required to recruit AP-1 to the major histocompatibility complex class I (MHC-I) cytoplasmic tail and disrupt MHC-I trafficking in HIV-1-infected primary T cells. J Virol 85:12216-26
Wonderlich, Elizabeth R; Leonard, Jolie A; Collins, Kathleen L (2011) HIV immune evasion disruption of antigen presentation by the HIV Nef protein. Adv Virus Res 80:103-27
Schaefer, Malinda R; Williams, Maya; Kulpa, Deanna A et al. (2008) A novel trafficking signal within the HLA-C cytoplasmic tail allows regulated expression upon differentiation of macrophages. J Immunol 180:7804-17
Schaefer, Malinda R; Wonderlich, Elizabeth R; Roeth, Jeremiah F et al. (2008) HIV-1 Nef targets MHC-I and CD4 for degradation via a final common beta-COP-dependent pathway in T cells. PLoS Pathog 4:e1000131
Wonderlich, Elizabeth R; Williams, Maya; Collins, Kathleen L (2008) The tyrosine binding pocket in the adaptor protein 1 (AP-1) mu1 subunit is necessary for Nef to recruit AP-1 to the major histocompatibility complex class I cytoplasmic tail. J Biol Chem 283:3011-22
Roeth, Jeremiah F; Collins, Kathleen L (2006) Human immunodeficiency virus type 1 Nef: adapting to intracellular trafficking pathways. Microbiol Mol Biol Rev 70:548-63
Kasper, Matthew R; Roeth, Jeremiah F; Williams, Maya et al. (2005) HIV-1 Nef disrupts antigen presentation early in the secretory pathway. J Biol Chem 280:12840-8
Williams, Maya; Roeth, Jeremiah F; Kasper, Matthew R et al. (2005) Human immunodeficiency virus type 1 Nef domains required for disruption of major histocompatibility complex class I trafficking are also necessary for coprecipitation of Nef with HLA-A2. J Virol 79:632-6

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