Retroviral Integration and AIDS Pathogenesis

Our laboratory focuses on retroviral integration using human and other retrovirus systems. Integration results in the permanent incorporation of a double-stranded DNA copy of the retroviral genome into host cell chromosomal DNA. This event contributes directly to the pathogenesis of the acquired immunodeficiency syndrome (AIDS) and certain other human diseases, as well as some animal models of cancer. Integration is catalyzed by the viral integrase enzyme, which is an attractive target for specific antiretroviral therapy because human cells have no closely related enzyme. Several years ago, I participated in the first purification of an enzymatically active retroviral integrase from a bacterial expression system and developed an oligonucleotide-based assay that is used throughout the world by laboratories studying the enzymology of integrases or attempting to identify inhibitors of human retroviral integrases for clinical use.

The overall goal of my laboratory is to understand the mechanism of retroviral integration so as to develop new ways to prevent or treat AIDS. We use a combined biochemical and genetic approach, employing a wide range of molecular biological techniques, to address important questions about the structure and function of integrase. Significantly, we took the novel approach of using the integrase of visna virus, a retrovirus of sheep, as a tool for analyzing the functional organization of the human immunodeficiency virus type 1 (HIV-1) integrase. We also use avian sarcoma viruses in our studies. We routinely clone, express, and purify mutant or chimeric integrase proteins which are analyzed in a variety of biochemical assays to map regions of the enzyme that interact with its various substrates.

Using the above strategy, we produced the first set of functional chimeric lentiviral integrases, altered the prevailing view of the organization of domains of the protein, discovered a previously unrecognized activity of integrase, and identified viral DNA positions that interact critically with this enzyme. Recently, we described the largest collection of natural, patient-derived HIV-1 integrases and used these variant enzymes to identify an amino acid in the central domain of integrase that may interact with human DNA. These results have contributed to the current model of retroviral integration and will enhance efforts to target specific actions or domains of integrase for clinical therapy.