title: Mechanism of microhomology-mediated end-joining promoted by human DNA polymerase θ.
abstract: Microhomology-mediated end-joining (MMEJ) is an error-prone alternative double-strand break-repair pathway that uses sequence microhomology to recombine broken DNA. Although MMEJ has been implicated in cancer development, the mechanism of this pathway is unknown. We demonstrate that purified human DNA polymerase θ (Polθ) performs MMEJ of DNA containing 3' single-strand DNA overhangs with ≥2 bp of homology, including DNA modeled after telomeres, and show that MMEJ is dependent on Polθ in human cells. Our data support a mechanism whereby Polθ facilitates end-joining and microhomology annealing, then uses the opposing overhang as a template in trans to stabilize the DNA synapse. Polθ exhibits a preference for DNA containing a 5'-terminal phosphate, similarly to polymerases involved in nonhomologous end-joining. Finally, we identify a conserved loop domain that is essential for MMEJ and higher-order structures of Polθ that probably promote DNA synapse formation.
authors: 1] Fels Institute for Cancer Research and Molecular Biology, Temple University School of Medicine, Philadelphia, Pennsylvania, USA. [2] Department of Biochemistry, Temple University School of Medicine, Philadelphia, Pennsylvania, USA.
authors: Tatiana
authors: T
authors: Kent
authors: 1] Fels Institute for Cancer Research and Molecular Biology, Temple University School of Medicine, Philadelphia, Pennsylvania, USA. [2] Department of Biochemistry, Temple University School of Medicine, Philadelphia, Pennsylvania, USA.
authors: Gurushankar
authors: G
authors: Chandramouly
authors: 1] Fels Institute for Cancer Research and Molecular Biology, Temple University School of Medicine, Philadelphia, Pennsylvania, USA. [2] Department of Biochemistry, Temple University School of Medicine, Philadelphia, Pennsylvania, USA.
authors: Shane Michael
authors: SM
authors: McDevitt
authors: 1] Fels Institute for Cancer Research and Molecular Biology, Temple University School of Medicine, Philadelphia, Pennsylvania, USA. [2] Department of Biochemistry, Temple University School of Medicine, Philadelphia, Pennsylvania, USA.
authors: Ahmet Y
authors: AY
authors: Ozdemir
authors: 1] Fels Institute for Cancer Research and Molecular Biology, Temple University School of Medicine, Philadelphia, Pennsylvania, USA. [2] Department of Biochemistry, Temple University School of Medicine, Philadelphia, Pennsylvania, USA.
authors: Richard T
authors: RT
authors: Pomerantz
mesh_terms: D045744:Cell Line, Tumor; D053903:DNA Breaks, Double-Stranded; D059766:DNA End-Joining Repair / Q000502:physiology*; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry / Q000502:physiology*; D006801:Humans; D008957:Models, Genetic; D008958:Models, Molecular; D000097462:DNA Polymerase theta
chemical_list: D004259:DNA-Directed DNA Polymerase; D000097462:DNA Polymerase theta
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