The HIV-1 envelope proteins are recognized as primary immune targets during infection and have been a major focus of AIDS vaccine development to elicit enduring broadly neutralizing antibody responses. These efforts have largely focused on the gp120 and gp41 ectodomains, based on the assumption that the gp41 C-terminal tail (CTT) was not exposed and important for envelope immunogenicity. Recent studies from our lab and others, however, clearly indicate that the CTT is a major determinant of HIV-1 envelope structure, function, and antigenicity and that the CTT can be a potent target for neutralizing antibodies directed to relatively conserved protein segments. Based on these observations, we hypothesize that the topology of the HIV gp41 CTT relative to the lipid bilayer is dynamic, and that CTT sequences are, at least transiently, located on the exterior of the viral or cellular lipid bilayer. We postulate further that CTT sequences are important determinants of Env antigenic and immunogenic properties, both by their association with other Env domains and as distinct immunologic targets, including neutralizing antibodies directed to conserved CTT sequences. We propose in this application to test this hypothesis in two specific aims: (i), to produce a definitive topological map of the CTT sequences relative to viral or cellular membranes and the influence of Gag association or fusion process on this topology, and (ii) to assess the role of the CTT as a determinant of Env immunogenicity and antigenicity, especially as related to neutralizing antibodies. The structural studies will utilize high-resolution membrane protein topology mapping techniques and antibody reactivity assays to define CTT structure in the context of virus- and cell-associated Env and in the context of novel chimeric proteins containing the HIV-1 CTT fused to the ectodomain of influenza virus HA envelope protein. The immunologic studies will employ experimental immunization of rabbits using established adenovirus vector or VLP vaccine procedures with codon optimized HIV-1 full length of truncated Envs or the engineered HA-CTT constructs to isolate CTT immunogenicity in the context of viral or cellular membranes. It is anticipated that the results of these studies will for the first time define the topology of the HIV-1 CTT and characterize its role as an immunogenic determinant and its potential as an immunogen to elicit broadly neutralizing antibodies to conserved sequences.

Public Health Relevance

HIV-1 envelope proteins are recognized as primary immune targets during infection and are a focal point of AIDS vaccine development. These efforts have largely focused on the gp120 and gp41 ectodomains, based on the assumption that the gp41 C-terminal tail (CTT) was not exposed and important for envelope immunogenicity. Recent studies, however, clearly indicate that the CTT is a major determinant of HIV-1 envelope structure, function, and antigenicity and that the CTT can be a potent target for neutralizing antibodies directed to relatively conserved protein segments. This proposal evaluates the structure of this relatively uncharacterized segment of the HIV-1 envelope and examines the role of these envelope sequences as determinants of viral neutralization properties and for its potential as a vaccine to enhance the production of neutralizing antibodies.

Agency
National Institute of Health (NIH)
Type
Research Project (R01)
Project #
5R01AI087533-05
Application #
8638885
Study Section
HIV/AIDS Vaccines Study Study Section (VACC)
Program Officer
Miller, Nancy R
Project Start
Project End
Budget Start
Budget End
Support Year
5
Fiscal Year
2014
Total Cost
Indirect Cost
Name
University of Pittsburgh
Department
Genetics
Type
Schools of Medicine
DUNS #
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
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Kuhlmann, Anne-Sophie; Steckbeck, Jonathan D; Sturgeon, Timothy J et al. (2014) Unique functional properties of conserved arginine residues in the lentivirus lytic peptide domains of the C-terminal tail of HIV-1 gp41. J Biol Chem 289:7630-40
Boscia, Alexander L; Akabori, Kiyotaka; Benamram, Zachary et al. (2013) Membrane structure correlates to function of LLP2 on the cytoplasmic tail of HIV-1 gp41 protein. Biophys J 105:657-66
Steckbeck, Jonathan D; Sun, Chengqun; Sturgeon, Timothy J et al. (2013) Detailed topology mapping reveals substantial exposure of the "cytoplasmic" C-terminal tail (CTT) sequences in HIV-1 Env proteins at the cell surface. PLoS One 8:e65220
Steckbeck, Jonathan D; Kuhlmann, Anne-Sophie; Montelaro, Ronald C (2013) C-terminal tail of human immunodeficiency virus gp41: functionally rich and structurally enigmatic. J Gen Virol 94:1-19
Steckbeck, Jonathan D; Craigo, Jodi K; Barnes, Christopher O et al. (2011) Highly conserved structural properties of the C-terminal tail of HIV-1 gp41 protein despite substantial sequence variation among diverse clades: implications for functions in viral replication. J Biol Chem 286:27156-66