The progression of human immunodeficiency virus (HIV) infection from asymptomatic infection to the development of AIDS is the result of a complex series of interactions between the virus and the infected host. Within an infected cell, both trans-acting viral gene products and cellular transcriptional factors participate in the regulation of viral gene expression and virus production, exerting their effects through cis-acting regulatory sequences present in the HIV genome. The goals of this project are to understand the roles that these regulatory sequences play in viral replication and in the activation or suppression of the expression of integrated HIV proviruses. Studies of the role of regulatory sequences in productive HIV infection will be based on mutagenesis of regulatory elements in the long terminal repeat (LTR) of an infectious molecular clone of HIV. These studies shall focus on the role of the HIV NF-kB and Spl binding sites in HIV replication in human T lymphocytes and monocytic cells; the function of other LTR sites will also be studied and the effects of the substitution of heterologous promoter/enhancer elements will be analyzed. Studies of the role of HIV regulatory sequences in chronically infected cells will be undertaken using cell lines containing HIV proviruses, whose expression can be induced. Techniques designed to study in vivo protein- DNA interactions will be employed in the study of these cells and will include DNAse hypersensitivity studies and in vivo footprinting assays. Regions of DNA-protein interactions associated with the suppression or activation of HIV production will be identified. Chronically infected cell lines containing HIV proviruses with LTR deletions will also be constructed to allow further study of the role of regulatory elements in expression of integrated HIV proviruses. Additional studies will be aimed at identifying previously unrecognized sites of DNA-protein interactions in the HIV genome and at identifying previously cellular factors associated with the high level of HIV gene expression seen in certain T cell lines. The long term goal of these studies is to develop a detailed understanding of the cis-acting elements in the HIV genome that are responsible for the activation or suppression of HIV gene expression and replication. Such an understanding may aid in the development of therapeutic strategies for HIV infection that would be aimed at blocking HIV gene expression, either by interfering with known activators of HIV gene expression or by identifying cellular inhibitors of viral expression.
