title: The Rap80-BRCC36 de-ubiquitinating enzyme complex antagonizes RNF8-Ubc13-dependent ubiquitination events at DNA double strand breaks.
abstract: DNA double strand breaks (DSBs) initiate reversible cellular checkpoint and repair activities. Whereas many of the activating events at DSBs have recently been elucidated, the mechanisms used to terminate responses at these sites are largely undefined. Here we report a pathway required to reverse RNF8-Ubc13 dependent ubiquitination events on chromatin flanking DSBs. Inhibition of the Rap80-BRCC36 de-ubiquitinating enzyme complex partially restored DSB-associated ubiquitin levels following RNF8 knockdown or proteasome inhibition. Similarly, BRCC36 knockdown or expression of a BRCC36 de-ubiquitinating enzyme-inactive mutant rescued both 53BP1 recruitment to DSBs and ionizing radiation-induced gammaH2AX ubiquitination following RNF8 depletion, and mitigated ionizing radiation sensitivity resulting from RNF8 deficiency. Thus, concomitant and opposing RNF8-Ubc13 ubiquitin ligase and Rap80-BRCC36 ubiquitin hydrolysis activities are responsible for determining steady-state ubiquitin levels at DNA DSBs. These findings reveal a Rap80-BRCC36 dependent pathway that is required for appropriate DSB recruitment and repair responses.
authors.affiliation: Department of Cancer Biology, Abramson Family Cancer Research Institute, University of Pennsylvania School of Medicine, Philadelphia, PA 19104-6160, USA.
authors.forename: Genze
authors.initials: G
authors.lastname: Shao
authors.forename: Dana R
authors.initials: DR
authors.lastname: Lilli
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Patterson-Fortin
authors.forename: Kara A
authors.initials: KA
authors.lastname: Coleman
authors.forename: Devon E
authors.initials: DE
authors.lastname: Morrissey
authors.forename: Roger A
authors.initials: RA
authors.lastname: Greenberg
mesh_terms: D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D053903:DNA Breaks, Double-Stranded; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D000072017:Deubiquitinating Enzymes; D006367:HeLa Cells; D056488:Histone Chaperones; D006657:Histones / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000235:genetics; D044763:Ubiquitin-Conjugating Enzymes / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases; D054875:Ubiquitination
chemical_list: D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C468783:H2AX protein, human; D056488:Histone Chaperones; D006657:Histones; D008565:Membrane Proteins; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; C486521:RNF8 protein, human; C466234:UIMC1 protein, human; C471607:UBE2N protein, human; D044763:Ubiquitin-Conjugating Enzymes; D044767:Ubiquitin-Protein Ligases; C518017:BRCC3 protein, human; D000072017:Deubiquitinating Enzymes; D046988:Proteasome Endopeptidase Complex