The pFOXC linear mitochondrial retroplasmids have features that make them possible evolutionary precursors of a broad range of genetic elements that replicate via reverse transcription. The 1.9 kb plasmid DNAs have a unique structure that includes a hairpin, terminal telomere-like repeats and a 5'- linked protein. The plasmids encode a reverse transcriptase (RT) that has been shown to exhibit unique enzymatic properties. The pFOXC-RT is deeply rooted in RT phylogeny which supports the hypothesis that the plasmids are ancestral to other retroelements, as well as the enzyme complex telomerase. Preliminary studies indicate that reverse transcription is protein-primed, which suggests the plasmids also show a mechanistic relationship to hepadnaviruses, a viral family that includes the important human pathogen hepatitis B. Proposed studies seek a better understanding of the mechanism of protein-priming and its relationship to the maintenance of the terminal telomere-like repeats. Using a well established in vitro reverse transcription system, models for repeat addition will be tested. We also propose to express the RT in a heterologous system to facilitate long-term biochemical characterization of the RT and explore possible practical applications. To more fully understand the relationship to other retroelements, we intend to conduct a comparative analysis of related linear mitochondrial retroplasmids. Our studies promise to provide valuable information concerning the mechanisms of reverse transcription and the evolution of retroelements. In addition, the research plan will provide numerous opportunities for undergraduate researchers, and students may take part in internships that prepares them for careers in biotechnology-related fields. ? ? ?
