The long term goal of our research is to elucidate the mechanisms of retroviral replication and recombination: (1) to increase our basic understanding of retrovirus biology, (ii) to aid in the development of alternative strategies to produce vector virus for applications (i.e. gene therapy), and (iii) to lead to new antiretroviral therapies for treatment of patients infected with retroviral pathogens (i.e. human immunodeficiency virus or human T-cell leukemia virus). Retroviruses are unique among all viruses because retrovirus virions contain two complete copies of the viral genetic material. The dimer RNA is reverse transcribed into DNA by a virally-encoded enzyme, reverse transcriptase (RT). The widely accepted model of reverse transcription requires that RT switch templates twice during synthesis of retroviral DNA from viral RNA, which is unusual for a polymerase. These required template switches result in formation of the long terminal repeats characteristic of retroviral DNA. Additional template switches occur frequently resulting in deletions, deletions with insertions and nonhomologous and homologous recombination. Homologous recombination is probably a major mechanism by which retroviruses recover from potentially lethal damage to the genetic material in the virion and is also thought to be important since it may contribute to the high rate of retroviral variation. Thus, template switching during retroviral DNA synthesis is indispensable for the retrovirus. However, little is known about the mechanism of retroviral template switching. This lack of information is particularly great for the more complex retroviruses, specifically human immunodeficiency virus. The goals of the proposed experiments are to extend our understanding of the template switches that must occur during reverse transcription, and the additional template switches that result in recombination. To accomplish these goals, we will use a well-studied and extremely manipulable system to characterize: (1) lesions that should be lethal in the absence of recombination, (2) the effect of cis-acting mutations on inter- and intramolecular template switches, (3) the effect of trans- acting factors on inter- and intramolecular template switches, and (4) the template switches utilized by more complex retroviruses.