Structure of BRCA1-BRCT/Abraxas Complex Reveals Phosphorylation-Dependent BRCT Dimerization at DNA Damage Sites.


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Authors
Wu, Qian 
Paul, Atanu 
Su, Dan 
Mehmood, Shahid 
Foo, Tzeh Keong 
Abstract

BRCA1 accumulation at DNA damage sites is an important step for its function in the DNA damage response and in DNA repair. BRCA1-BRCT domains bind to proteins containing the phosphorylated serine-proline-x-phenylalanine (pSPxF) motif including Abraxas, Bach1/FancJ, and CtIP. In this study, we demonstrate that ionizing radiation (IR)-induces ATM-dependent phosphorylation of serine 404 (S404) next to the pSPxF motif. Crystal structures of BRCT/Abraxas show that phosphorylation of S404 is important for extensive interactions through the N-terminal sequence outside the pSPxF motif and leads to formation of a stable dimer. Mutation of S404 leads to deficiency in BRCA1 accumulation at DNA damage sites and cellular sensitivity to IR. In addition, two germline mutations of BRCA1 are found to disrupt the dimer interface and dimer formation. Thus, we demonstrate a mechanism involving IR-induced phosphorylation and dimerization of the BRCT/Abraxas complex for regulating Abraxas-mediated recruitment of BRCA1 in response to IR.

Description
Keywords
Amino Acid Sequence, Ataxia Telangiectasia Mutated Proteins, BRCA1 Protein, Bone Neoplasms, Carrier Proteins, Cell Line, Tumor, Cell Nucleus, Crystallography, X-Ray, DNA Damage, Germ-Line Mutation, Humans, Models, Molecular, Molecular Sequence Data, Osteosarcoma, Phosphorylation, Protein Binding, Protein Interaction Domains and Motifs, Protein Multimerization, RNA Interference, Serine, Transfection
Journal Title
Mol Cell
Conference Name
Journal ISSN
1097-2765
1097-4164
Volume Title
61
Publisher
Elsevier BV
Sponsorship
Wellcome Trust (093167/Z/10/Z)
We thank beamline scientists at Diamond Light Source for help during data collection of crystal and SAXS. The crystallization experiments were performed in the Crystallographic X-ray facility at the Department of Biochemistry, University of Cambridge. We are grateful to the Facility Manager, Dr. Dimitri Chirgadze, for his assistance in using these facilities and advice during crystal structure determination. We also thank Dr. Yanfen Hu (University of Texas Health Science Center at San Antonia) for the pFlag-BRCA1 plasmid and Dr. Angela Pacitto (University of Cambridge) for reading the manuscript. We thank Dr. Adriana Paulucci-Holthanuzen (Department of Genetics-MD Anderson Microscopy Core Facility) for assistance with images and analysis. Q.W., T.O. and T.L.B. are funded by the Wellcome Trust (Grant 093167/Z/10/Z). A.P. is an awardee of the Schissler Foundation Fellowship, the Center for Cancer Epigenetics Scholarship and the Andrew Sowell-Wade Huggins Scholarship. This work is supported by the National Institutes of Health grant (CA155025 to B.W) with funds from the University of Texas MD Anderson Cancer Center (IRG, Center for Cancer Epigenetics, Center for Genetics and Genomics Pilot Award). S.M. is funded by the Medical Research Council (grant 98101 to C.V.R.) and C.V.R. is a Royal Society Research Professor. T.K.F and B.X. are supported by National Institutes of Health grant (R01CA138804 to B.X).