The last few years have witnessed a paradigm shift from the view that HIV-1 genetic recombination is a somewhat rare error mechanism to the recognition that it is frequent and is probably an integral part of viral DNA synthesis. This proposal describes a basic molecular genetics study with the long-term objectives of better understanding HIV genetic recombination mechanisms and the factors that modulate it. In the proposed work, a series of modified HIV-1 genomes will be used in single replication cycle tissue culture-based assays to test basic properties of recombination and how properties of RNA templates like those that the virus encounters in vivo affect the frequency and outcomes of recombinogenic template switching.
The specific aims are 1) To develop new assays for studying recombination related properties during HIV-1 replication in cultured cells, including testing predictions of the model that some HIV variability may result from transductive recombination. 2) To test the effects of different physical contexts of potential recombination substrates, such as the nature of dimer linkages, on the substrates abilities to contribute to reverse transcription products, and 3) To examine properties of template switch complexes including the determinants of transfer point, and whether or not mismatch excision and/or extension is apparent among template switch products. The proposed work will advance basic understanding of HIV-1 reverse transcription properties. It is important that this work be performed with HIV-1 because these properties may differ from analogous processes for simple retroviruses. Gamma- and other simple retroviruses' reverse transcription properties during replication have been better studied than have those of HIV. Although it has been assumed that simple retroviruses' properties represent those of HIV, recent advances provide examples of significant differences. Reverse transcription remains the foremost target of antiretroviral drugs, and a more thorough understanding of basic features of the process during HIV replication will be useful to the design and implementation of AIDS antiviral strategies. ? ? ?
