title: ATM damage response and XLF repair factor are functionally redundant in joining DNA breaks.
abstract: Classical non-homologous DNA end-joining (NHEJ) is a major mammalian DNA double-strand-break (DSB) repair pathway. Deficiencies for classical NHEJ factors, such as XRCC4, abrogate lymphocyte development, owing to a strict requirement for classical NHEJ to join V(D)J recombination DSB intermediates. The XRCC4-like factor (XLF; also called NHEJ1) is mutated in certain immunodeficient human patients and has been implicated in classical NHEJ; however, XLF-deficient mice have relatively normal lymphocyte development and their lymphocytes support normal V(D)J recombination. The ataxia telangiectasia-mutated protein (ATM) detects DSBs and activates DSB responses by phosphorylating substrates including histone H2AX. However, ATM deficiency causes only modest V(D)J recombination and lymphocyte developmental defects, and H2AX deficiency does not have a measurable impact on these processes. Here we show that XLF, ATM and H2AX all have fundamental roles in processing and joining DNA ends during V(D)J recombination, but that these roles have been masked by unanticipated functional redundancies. Thus, combined deficiency of ATM and XLF nearly blocks mouse lymphocyte development due to an inability to process and join chromosomal V(D)J recombination DSB intermediates. Combined XLF and ATM deficiency also severely impairs classical NHEJ, but not alternative end-joining, during IgH class switch recombination. Redundant ATM and XLF functions in classical NHEJ are mediated by ATM kinase activity and are not required for extra-chromosomal V(D)J recombination, indicating a role for chromatin-associated ATM substrates. Correspondingly, conditional H2AX inactivation in XLF-deficient pro-B lines leads to V(D)J recombination defects associated with marked degradation of unjoined V(D)J ends, revealing that H2AX has a role in this process.
authors.affiliation: Howard Hughes Medical Institute, The Children's Hospital, the Immune Disease Institute and the Harvard Medical School, Boston, Massachusetts 02115, USA.
authors.forename: Shan
authors.initials: S
authors.lastname: Zha
authors.forename: Chunguang
authors.initials: C
authors.lastname: Guo
authors.forename: Cristian
authors.initials: C
authors.lastname: Boboila
authors.forename: Valentyn
authors.initials: V
authors.lastname: Oksenych
authors.forename: Hwei-Ling
authors.initials: HL
authors.lastname: Cheng
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhang
authors.forename: Duane R
authors.initials: DR
authors.lastname: Wesemann
authors.forename: Grace
authors.initials: G
authors.lastname: Yuen
authors.forename: Harin
authors.initials: H
authors.lastname: Patel
authors.forename: Peter H
authors.initials: PH
authors.lastname: Goff
authors.forename: Richard L
authors.initials: RL
authors.lastname: Dubois
authors.forename: Frederick W
authors.initials: FW
authors.lastname: Alt
mesh_terms: D000818:Animals; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D002461:Cell Line, Transformed; D002843:Chromatin / Q000378:metabolism; D033481:Chromosomes, Mammalian / Q000235:genetics / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D004622:Embryo, Mammalian / Q000196:embryology / Q000378:metabolism; D015322:Gene Rearrangement, B-Lymphocyte / Q000235:genetics; D006657:Histones / Q000378:metabolism*; D051379:Mice; D054448:Precursor Cells, B-Lymphoid / Q000166:cytology / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D011995:Recombination, Genetic; D025521:Tumor Suppressor Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*
chemical_list: D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; D006657:Histones; D017346:Protein Serine-Threonine Kinases; D025521:Tumor Suppressor Proteins; C519508:XLF protein, mouse; C576324:ATM protein, human; C576325:Atm protein, mouse; C475961:gamma-H2AX protein, mouse