This grant proposes to identify determinants on HIV-l encoded proteins recognized by T cells, characterize these determinants, modify them to increase their immunogenicity, and make them into universally recognized epitopes. Cell-mediated immune responses will be an important component of immunity to HIV-l infection and this project will focus on the determinants recognized by T cells. The initial studies will examine determinants on the nef protein, a negative regulator of viral transcription. This protein could be an important target for the immune system since it is produced in infected cells, such as macrophages, which harbor the virus in a quiescent state. To identify the determinants recognized by nef specific cytotoxic T lymphocytes (CTL) as well as helper T cells (Th), we will generate cell lines expressing the nef protein. These cells will be used to prime mice in vivo as well as be used as target cells in CTL assays. To detect Th cell responses, we also will produce soluble nef protein. Bulk and cloned CTL and Th cell lines will be produced from the in vivo primed mice of three different strains and the determinants being recognized will be ascertained using synthetic peptides. Various algorithms will be used to predict the T cell epitopes, and then tested to determine which are recognized by T cells. The identified determinants will then be tested in other mouse strains to find a determinant which is recognized by many different mouse strains. After the determinants have been identified, we will then choose the best 2 or 3, and characterize them in detail using synthetic peptides. The purpose of the detailed analysis is to identify the critical residues involved in the formation of the determinant. The HIV-1 encoded proteins have tremendous genetic variability, which makes it difficult to find an epitope which is present on all HIV-l isolates. T cell epitopes have only 5-6 critical residues, and 5- 10 non-critical residues. By determining which residues are critical for the formation of each epitope, we then can identify the epitope in which these critical residues are conserved in all HIV-l isolates. Further studies will involve modifying these epitopes so that they will be universally recognized by mouse strains, as well as modifications which will enhance the immunogenicity of the determinants. From these studies we will be able to identify and enhance T cell epitopes on HIV-l encoded proteins which could be used as part of a HIV-1 subunit vaccine.
| Graziano, R F; Allen, P M (1992) Enhancing the immunogenicity of a permissive binding T cell epitope derived from the simian immunodeficiency virus-encoded negative regulatory factor. J Immunol 149:556-61 |
| Chen, J S; Lorenz, R G; Goldberg, J et al. (1991) Identification and characterization of a T cell-inducing epitope of bovine ribonuclease that can be restricted by multiple class II molecules. J Immunol 147:3672-8 |
