title: 53BP1-dependent robust localized KAP-1 phosphorylation is essential for heterochromatic DNA double-strand break repair.
abstract: DNA double-strand breaks (DSBs) trigger ATM (ataxia telangiectasia mutated) signalling and elicit genomic rearrangements and chromosomal fragmentation if misrepaired or unrepaired. Although most DSB repair is ATM-independent, approximately 15% of ionizing radiation (IR)-induced breaks persist in the absence of ATM-signalling. 53BP1 (p53-binding protein 1) facilitates ATM-dependent DSB repair but is largely dispensable for ATM activation or checkpoint arrest. ATM promotes DSB repair within heterochromatin by phosphorylating KAP-1 (KRAB-associated protein 1, also known as TIF1beta, TRIM28 or KRIP-1; ref. 2). Here, we show that the ATM signalling mediator proteins MDC1, RNF8, RNF168 and 53BP1 are also required for heterochromatic DSB repair. Although KAP-1 phosphorylation is critical for 53BP1-mediated repair, overall phosphorylated KAP-1 (pKAP-1) levels are only modestly affected by 53BP1 loss. pKAP-1 is transiently pan-nuclear but also forms foci overlapping with gammaH2AX in heterochromatin. Cells that do not form 53BP1 foci, including human RIDDLE (radiosensitivity, immunodeficiency, dysmorphic features and learning difficulties) syndrome cells, fail to form pKAP-1 foci. 53BP1 amplifies Mre11-NBS1 accumulation at late-repairing DSBs, concentrating active ATM and leading to robust, localized pKAP-1. We propose that ionizing-radiation induced foci (IRIF) spatially concentrate ATM activity to promote localized alterations in regions of chromatin otherwise inhibitory to repair.
authors.affiliation: Genome Damage and Stability Centre, University of Sussex, East Sussex, BN1 9RQ, UK.
authors.forename: Angela T
authors.initials: AT
authors.lastname: Noon
authors.forename: Atsushi
authors.initials: A
authors.lastname: Shibata
authors.forename: Nicole
authors.initials: N
authors.lastname: Rief
authors.forename: Markus
authors.initials: M
authors.lastname: Löbrich
authors.forename: Grant S
authors.initials: GS
authors.lastname: Stewart
authors.forename: Penelope A
authors.initials: PA
authors.lastname: Jeggo
authors.forename: Aaron A
authors.initials: AA
authors.lastname: Goodarzi
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000378:metabolism; D002460:Cell Line; D002478:Cells, Cultured; D053903:DNA Breaks, Double-Stranded / Q000528:radiation effects; D004260:DNA Repair / Q000235:genetics / Q000502:physiology*; D045643:DNA Repair Enzymes; D004268:DNA-Binding Proteins / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006570:Heterochromatin / Q000378:metabolism* / Q000528:radiation effects; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000502:physiology*; D000076228:MRE11 Homologue Protein; D051379:Mice; D041681:NIH 3T3 Cells; D009687:Nuclear Proteins / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011839:Radiation, Ionizing; D012097:Repressor Proteins / Q000378:metabolism*; D015534:Trans-Activators / Q000378:metabolism; D000076183:Tripartite Motif-Containing Protein 28; D025521:Tumor Suppressor Proteins / Q000378:metabolism; D000071857:Tumor Suppressor p53-Binding Protein 1; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D006570:Heterochromatin; D047908:Intracellular Signaling Peptides and Proteins; C413163:MDC1 protein, human; C121766:MRE11 protein, human; C512689:Mre11a protein, mouse; C112156:NBN protein, human; C466108:Nijmegen breakage syndrome 1 protein, mouse; D009687:Nuclear Proteins; C486521:RNF8 protein, human; D012097:Repressor Proteins; C498301:TP53BP1 protein, human; D015534:Trans-Activators; D025521:Tumor Suppressor Proteins; D000071857:Tumor Suppressor p53-Binding Protein 1; C535210:RNF168 protein, human; C101116:TRIM28 protein, human; D000076183:Tripartite Motif-Containing Protein 28; D044767:Ubiquitin-Protein Ligases; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C576325:Atm protein, mouse; D017346:Protein Serine-Threonine Kinases; D000076228:MRE11 Homologue Protein; D045643:DNA Repair Enzymes