Catalytic and noncatalytic roles of the CtIP endonuclease in double-strand break end resection.
The carboxy-terminal binding protein (CtBP)-interacting protein (CtIP) is known to function in 5' strand resection during homologous recombination, similar to the budding yeast Sae2 protein, but its role in this process is unclear. Here, we characterize recombinant human CtIP and find that it exhibits 5' flap endonuclease activity on branched DNA structures, independent of the MRN complex. Phosphorylation of CtIP at known damage-dependent sites and other sites is essential for its catalytic activity, although the S327 and T847 phosphorylation sites are dispensable. A catalytic mutant of CtIP that is deficient in endonuclease activity exhibits wild-type levels of homologous recombination at restriction enzyme-generated breaks but is deficient in processing topoisomerase adducts and radiation-induced breaks in human cells, suggesting that the nuclease activity of CtIP is specifically required for the removal of DNA adducts at sites of DNA breaks.
The Howard Hughes Medical Institute and Department of Molecular Biosciences, Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, TX 78712, USA.
Nodar
N
Makharashvili
Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110, USA.
Anthony T
AT
Tubbs
The Howard Hughes Medical Institute and Department of Molecular Biosciences, Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, TX 78712, USA.
Soo-Hyun
SH
Yang
Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA.
Hailong
H
Wang
Radiation Biology and DNA Repair, Darmstadt University of Technology, 64287 Darmstadt, Germany.
Olivia
O
Barton
The Howard Hughes Medical Institute and Department of Molecular Biosciences, Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, TX 78712, USA.
Yi
Y
Zhou
The Howard Hughes Medical Institute and Department of Molecular Biosciences, Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, TX 78712, USA.
Rajashree A
RA
Deshpande
The Howard Hughes Medical Institute and Department of Molecular Biosciences, Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, TX 78712, USA.
Ji-Hoon
JH
Lee
Radiation Biology and DNA Repair, Darmstadt University of Technology, 64287 Darmstadt, Germany.
Markus
M
Lobrich
Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110, USA.
Barry P
BP
Sleckman
Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA.
Xiaohua
X
Wu
The Howard Hughes Medical Institute and Department of Molecular Biosciences, Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, TX 78712, USA. Electronic address: tpaull@utexas.edu.
Tanya T
TT
Paull
D001665:Binding Sites / Q000235:genetics; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002384:Catalysis; D002460:Cell Line; D002470:Cell Survival / Q000235:genetics; D004247:DNA / Q000235:genetics; D053903:DNA Breaks, Double-Stranded; D059766:DNA End-Joining Repair / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics; D004706:Endodeoxyribonucleases; D004720:Endonucleases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000235:genetics; D011499:Protein Processing, Post-Translational / Q000235:genetics; D011839:Radiation, Ionizing; D011995:Recombination, Genetic; D059767:Recombinational DNA Repair / Q000235:genetics*
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D004247:DNA; D004706:Endodeoxyribonucleases; D004720:Endonucleases; C410741:RBBP8 protein, human