Characterization of the role of Drosophila melanogaster Pol32 in Homologous Recombination and DNA Repair.
- Double strand breaks (DSBs) in DNA represent one of the most toxic and dangerous classes of DNA lesions. DSB repair (DSBR) occurs through two canonical pathways, homologous recombination (HR) or nonhomologous end joining (NHEJ). In Drosophila melanogaster, DSBR of P-element induced breaks is believed to occur through the HR pathway known as synthesis dependent strand annealing (SDSA). A variety of... read moreproteins have been implicated in SDSA, but the polymerases involved in initiating and extending repair products have not been fully characterized. The Pol32 subunit of polymerase Î´ in S. cerevisiae has been found to be necessary for break induced replication and various DNA repair pathways. We hypothesize that the pol32 Drosophila ortholog may also play a role in processive synthesis during HR repair. Two deletion mutations in pol32 utilized in this study were generated by an imprecise P-element excision (L2 and L30). Mutagen sensitivity assays and a site specific DSBR assay were employed to study the mutants. The L2 null mutation was found to be sensitive to a range of mutagens, including MMS and IR, indicating a role for pol32 in base excision repair, nucleotide excision repair, replication restart, and HR. Analysis of the repair assay revealed a significant defect in pol32 mutants for HR repair and processive synthesis. These results implicate Pol32 as an important player in DSBR and in the processive synthesis of polymerase δ.read less