Roles of DNA repair pathways in Sleeping Beauty transposition in vertebrate cells

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Several factors determine which DNA repair pathways are used to heal the damage caused by movement of a transposable element, including the transposon itself, the structure of the lesion, the host organism, the status of the cell cycle, and whether transposition occurs in the soma or in the germline. The contribution of DNA repair to DNA transposition in vertebrate cells has not been previously addressed, due to the lack of active transposons in these species. Our research laboratory is using the Sleeping Beauty (SB) transposable element as a tool to probe transposon-host cell interactions in vertebrates. Our previous work has established that nonhomologous end joining and homologous recombination both contribute to the repair of SB-induced double-strand DNA breaks in mammalian somatic cells. The proposed project is aimed at surveying different cellular factors involved in DNA replication, repair and damage signaling for their potential roles in SB transposition. We propose to study the contribution of these factors in establishing a successful transposition event, and differential regulation of these processes through the cell cycle. Transposition-inflicted DNA damage differs form radiation-, retroviral integration-, restriction enzyme-, and V(D)J recombination-generated lesions. The proposed research might therefore provide important clues about differential regulation of transposition and other DNA recombination mechanisms, and extend our understanding of the principal molecular processes involved in cellular responses to DNA damage in mammalian cells.