The major aim of this project is to demonstrate that superantigens (sAgs) encoded by HIV are involved in the deletion and functional paralysis of CD4+ T cells in HIV-infected patients. In vitro and in vivo models will be used to identify a sAg encoded by HIV. A large panel of human T cell receptor (TCR) beta chain variable regions (Vb's) have been or will be cloned using PCR. These cDNA clones will be introduced by transfection into different T cell lines which lack the TCR Vb. Transfected cells which express the human TCR Vb will be used as immunogens to generate a panel of mAbs specific for human TCR V regions. Generation of these mAbs should be greatly facilitated by the availability of these transfected lines. These mAbs, which will be made available to the whole scientific community should prove a major asset in the analysis and follow-up of the TCR repertoire in HIV infection. These transfected cells will also be used as effector cells in attempting to identify a sAg encoded by HIV. CD4+ class II+ cells susceptible to HIV infection and purified populations of dendritic cells will be used to present HIV to the TCR transfectants. Presence of a sAg in HIV should lead to the activation, as measured by IL-2 production of T cells expressing only a restricted set of Vb regions. Chimeric viruses generated by recombinant DNA techniques or defective HIV particles already available will be used to amp the putative sAg in HIV. A strategy has been devised which will enable the shuffling of polymorphic sequences of HIV, specially in the envelope and nef glycoproteins, between different molecular clones of HIV. To confirm that these mechanisms operate on human peripheral blood lymphocytes, an in vitro system which allows the stimulation of T cells by autologous HIV-infected cells will be used. Expansion of CD4+ and CD8+ cells expressing specific Vbs, similar to those expressed by the T cell hybridomas which respond in vitro, should be observed. Quantitative PCR and flow cytometry with TCR Vb-specific mAbs will be used to monitor the TCR repertoire. Identification of a sAg in HIV will provide a better understanding of the pathophysiology of the virus and will open the way to novel therapeutic strategies.
