A novel role of the Dna2 translocase function in DNA break resection.
DNA double-strand break repair by homologous recombination entails nucleolytic resection of the 5' strand at break ends. Dna2, a flap endonuclease with 5'-3' helicase activity, is involved in the resection process. The Dna2 helicase activity has been implicated in Okazaki fragment processing during DNA replication but is thought to be dispensable for DNA end resection. Unexpectedly, we found a requirement for the helicase function of Dna2 in end resection in budding yeast cells lacking exonuclease 1. Biochemical analysis reveals that ATP hydrolysis-fueled translocation of Dna2 on ssDNA facilitates 5' flap cleavage near a single-strand-double strand junction while attenuating 3' flap incision. Accordingly, the ATP hydrolysis-defective dna2-K1080E mutant is less able to generate long products in a reconstituted resection system. Our study thus reveals a previously unrecognized role of the Dna2 translocase activity in DNA break end resection and in the imposition of the 5' strand specificity of end resection.
Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
Adam S
AS
Miller
Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
James M
0000-0002-7706-6134
JM
Daley
Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas 77030, USA.
Nhung Tuyet
NT
Pham
Molecular and Cellular Biochemistry Department, Indiana University, Bloomington, Indiana 47405, USA.
Hengyao
H
Niu
Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
Xiaoyu
X
Xue
Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas 77030, USA.
Grzegorz
G
Ira
Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
Patrick
P
Sung
D024506:5' Flanking Region / Q000235:genetics; D000255:Adenosine Triphosphate / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded; D059766:DNA End-Joining Repair / Q000235:genetics; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair / Q000235:genetics*; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
D029701:Saccharomyces cerevisiae Proteins; D000255:Adenosine Triphosphate; D004265:DNA Helicases; C410222:DNA2 protein, S cerevisiae
DNA repair; double-strand break; end resection; homologous recombination