Spatial organization of the mammalian genome surveillance machinery in response to DNA strand breaks.
We show that DNA double-strand breaks (DSBs) induce complex subcompartmentalization of genome surveillance regulators. Chromatin marked by gamma-H2AX is occupied by ataxia telangiectasia-mutated (ATM) kinase, Mdc1, and 53BP1. In contrast, repair factors (Rad51, Rad52, BRCA2, and FANCD2), ATM and Rad-3-related (ATR) cascade (ATR, ATR interacting protein, and replication protein A), and the DNA clamp (Rad17 and -9) accumulate in subchromatin microcompartments delineated by single-stranded DNA (ssDNA). BRCA1 and the Mre11-Rad50-Nbs1 complex interact with both of these compartments. Importantly, some core DSB regulators do not form cytologically discernible foci. These are further subclassified to proteins that connect DSBs with the rest of the nucleus (Chk1 and -2), that assemble at unprocessed DSBs (DNA-PK/Ku70), and that exist on chromatin as preassembled complexes but become locally modified after DNA damage (Smc1/Smc3). Finally, checkpoint effectors such as p53 and Cdc25A do not accumulate at DSBs at all. We propose that subclassification of DSB regulators according to their residence sites provides a useful framework for understanding their involvement in diverse processes of genome surveillance.
Bekker-Jensen
Simon
S
Institute of Cancer Biology and Centre for Genotoxic Stress Research, Danish Cancer Society, DK-2100 Copenhagen, Denmark.
Lukas
Claudia
C
Kitagawa
Risa
R
Melander
Fredrik
F
Kastan
Michael B
MB
Bartek
Jiri
J
Lukas
Jiri
J
D000818:Animals; D019313:BRCA1 Protein / Q000502:physiology; D018797:Cell Cycle Proteins / Q000502:physiology*; D002460:Cell Line; D002478:Cells, Cultured; D000071877:Checkpoint Kinase 1; D002843:Chromatin / Q000502:physiology; D002868:Chromosomal Proteins, Non-Histone / Q000502:physiology; D004247:DNA / Q000378:metabolism* / Q000528:radiation effects; D004249:DNA Damage; D004260:DNA Repair; D016678:Genome; D006801:Humans; D007834:Lasers; D009687:Nuclear Proteins / Q000502:physiology; D010766:Phosphorylation; D011494:Protein Kinases / Q000502:physiology; D000098550:Structural Maintenance of Chromosome Protein 1
D019313:BRCA1 Protein; D018797:Cell Cycle Proteins; D002843:Chromatin; D002868:Chromosomal Proteins, Non-Histone; C112156:NBN protein, human; D009687:Nuclear Proteins; D000098550:Structural Maintenance of Chromosome Protein 1; D004247:DNA; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1