The HIV-1 matrix protein (MAp17) plays an essential role in the viral life cycle. In addition, this structural protein also has extracellular functions manifested in vitro mainly by enhanced T-cell proliferation, HIV-1 replication and proinflammatory cytokine production. We have found that the MAp 17 exerts its biological activities via binding as yet to an unidentified cell surface receptor expressed on peripheral blood-derived mononuclear cells. Mouse antibodies generated to the receptor-binding region (RBR) of the MAp 17 blocked binding to the receptor and abrogated the extracellular effects. Moreover, asymptomatic HIV-1 infected individuals, in contrast to AIDS patients, frequently recognized a potent B cell epitope located at the N-terminus of MAp17, which overlaps with the RBR. Based on these observations, we hypothesize that HIV-1 positive asymptomatic individuals with slower progression towards AIDS generate antibody responses to the RBR of the MAp 17, and these antibodies block MAp 17 binding to its receptor. To address the role of these blocking antibodies in the pathogenesis of AIDS a sensitive serological assay will be developed. The assay will be based on measurements of biotin-conjugated MAp 17 binding to cells using flow cytometry. The establishment of the methodology will proceed as follows: (1) development of a binding assay suitable for assessment of sera from immunized animals to block the specific binding of the MAp 17 to its receptor. Important steps in this stage of the binding assay development are: (a) identification of a permanent cell line suitable for MAp17binding; (b) optimization of blocking the MAp 17 interaction with its receptor using antibodies to the MAp 17; (c) assessment of antibodies that recognize the functional/B-cell epitope region, termed also functional RBR, in MAp17pep scan analyses; and (d) testing of these antibodies to neutralize MAp 17 capacity to induce proinflammatory cytokine production; (2) evaluation of applicability of the binding assay for detecting antibodies with capacity to block MAp 17 binding to its receptor in sera from individuals positive for HIV-1p 17/p24. The binding assay for assessment of human sera will include the following important steps: (a) using human sera, we will reassess the optimal conditions for blocking of MAp 17/receptor interactions that were determined in the testing of sera from immunized animals; (b) testing the capacity of sera to recognize the functional RBR in the MAp 17 pep scan analyses; and (c) evaluate RBR positive human sera to neutralize capacity of MAp 17 to induce the proinflammatory cytokine production. The binding assay development using sera from MAp 17 immunized animals will be performed in the first year and its applicability for testing HIV-1 (MAp17) positive human sera will be performed in the second year of the project.
| Popovic, Mikulas; Tenner-Racz, Klara; Pelser, Colleen et al. (2005) Persistence of HIV-1 structural proteins and glycoproteins in lymph nodes of patients under highly active antiretroviral therapy. Proc Natl Acad Sci U S A 102:14807-12 |
