SNM1A (DCLRE1A) is crucial for efficient repair of complex DNA breaks in human cells

DNA double-strand breaks (DSBs), such as those produced by radiation and radiomimetics, are amongst the most toxic forms of cellular damage, in part because they involve extensive oxidative modifications at the break termini. Prior to completion of DSB repair, the chemically modified termini must be removed. Various DNA processing enzymes have been implicated in the processing of these ‘dirty ends’, but molecular knowledge of this process is limited. Here, we demonstrate a role for the metallo-β-lactamase fold 5ʹ-3ʹ exonuclease SNM1A in this vital process. Cells disrupted for SNM1A manifest increased sensitivity to radiation and radiomimetic agents and show defects in DSB damage repair. SNM1A is recruited and is retained at the sites of DSB damage via the concerted action of its three highly conserved PBZ, PIP box and UBZ interaction domains, which mediate interactions with poly-ADP-ribose chains, PCNA and the ubiquitinated form of PCNA, respectively. SNM1A can resect DNA containing oxidative lesions induced by radiation damage at break termini. The combined results reveal a crucial role for SNM1A to digest chemically modified DNA during the repair of DSBs and imply that the catalytic domain of SNM1A is an attractive target for potentiation of radiotherapy.

[1]  R. Greenberg,et al.  Break-induced replication orchestrates resection-dependent template switching , 2023, Nature.

[2]  N. Lowndes,et al.  Immediate-Early, Early, and Late Responses to DNA Double Stranded Breaks , 2022, Frontiers in Genetics.

[3]  S. Ovchinnikov,et al.  ColabFold: making protein folding accessible to all , 2022, Nature Methods.

[4]  O. Gileadi,et al.  A phosphate binding pocket is a key determinant of exo- versus endo-nucleolytic activity in the SNM1 nuclease family , 2021, Nucleic acids research.

[5]  Oriol Vinyals,et al.  Highly accurate protein structure prediction with AlphaFold , 2021, Nature.

[6]  Conrad C. Huang,et al.  UCSF ChimeraX: Structure visualization for researchers, educators, and developers , 2020, Protein science : a publication of the Protein Society.

[7]  W. Xiao,et al.  DNA-damage tolerance through PCNA ubiquitination and sumoylation. , 2020, The Biochemical journal.

[8]  O. Gileadi,et al.  The SNM1A DNA repair nuclease , 2020, DNA repair.

[9]  A. Bielinsky,et al.  Ubiquitinated-PCNA protects replication forks from DNA2-mediated degradation by regulating Okazaki fragment maturation and chromatin assembly , 2020, Nature Communications.

[10]  N. Willis,et al.  DNA double-strand break repair-pathway choice in somatic mammalian cells , 2019, Nature Reviews Molecular Cell Biology.

[11]  A. Shibata,et al.  A historical reflection on our understanding of radiation-induced DNA double strand break repair in somatic mammalian cells; interfacing the past with the present , 2019, International journal of radiation biology.

[12]  A. D’Andrea.  Mechanisms of PARP inhibitor sensitivity and resistance. , 2018, DNA repair.

[13]  Ulrich Rothbauer,et al.  Under the Microscope: Single-Domain Antibodies for Live-Cell Imaging and Super-Resolution Microscopy , 2017, Front. Immunol..

[14]  I. Demuth,et al.  SNM1B/Apollo in the DNA damage response and telomere maintenance , 2017, Oncotarget.

[15]  P. McHugh,et al.  The Chemical Biology of Human Metallo-β-Lactamase Fold Proteins , 2016, Trends in biochemical sciences.

[16]  Junjie Chen.  Faculty Opinions recommendation of DNA repair. Proteomics reveals dynamic assembly of repair complexes during bypass of DNA cross-links. , 2015 .

[17]  Jürgen Cox,et al.  Proteomics reveals dynamic assembly of repair complexes during bypass of DNA cross-links , 2015, Science.

[18]  S. Jackson,et al.  Neddylation Promotes Ubiquitylation and Release of Ku from DNA-Damage Sites , 2015, Cell reports.

[19]  J. Doudna,et al.  The new frontier of genome engineering with CRISPR-Cas9 , 2014, Science.

[20]  D. Korzhnev,et al.  NMR structure of the human Rad18 zinc finger in complex with ubiquitin defines a class of UBZ domains in proteins linked to the DNA damage response. , 2014, Biochemistry.

[21]  Michal Zimmermann,et al.  53BP1: pro choice in DNA repair. , 2014, Trends in cell biology.

[22]  R. Scully,et al.  Double strand break repair functions of histone H2AX. , 2013, Mutation research.

[23]  T. Sixma,et al.  Intrinsic Flexibility of Ubiquitin on Proliferating Cell Nuclear Antigen (PCNA) in Translesion Synthesis* , 2012, The Journal of Biological Chemistry.

[24]  S. Boulton,et al.  Playing the end game: DNA double-strand break repair pathway choice. , 2012, Molecular cell.

[25]  Johannes E. Schindelin,et al.  Fiji: an open-source platform for biological-image analysis , 2012, Nature Methods.

[26]  C. Allerston,et al.  Characterization of the Human SNM1A and SNM1B/Apollo DNA Repair Exonucleases , 2012, The Journal of Biological Chemistry.

[27]  N. Maizels,et al.  DNA Repair Factor MRE11/RAD50 Cleaves 3′-Phosphotyrosyl Bonds and Resects DNA to Repair Damage Caused by Topoisomerase 1 Poisons* , 2011, The Journal of Biological Chemistry.

[28]  O. Gileadi,et al.  Human SNM1A and XPF-ERCC1 collaborate to initiate DNA interstrand cross-link repair. , 2011, Genes & development.

[29]  A. D’Andrea,et al.  RAD18-dependent Recruitment of SNM1A to DNA Repair Complexes by a Ubiquitin-binding Zinc Finger* , 2010, The Journal of Biological Chemistry.

[30]  S. West,et al.  Solution structures of the two PBZ domains from human APLF and their interaction with poly(ADP-ribose) , 2010, Nature Structural &Molecular Biology.

[31]  H. Ulrich,et al.  Regulating post-translational modifications of the eukaryotic replication clamp PCNA. , 2009, DNA repair.

[32]  J. Glover,et al.  RAD18 transmits DNA damage signaling to elicit homologous recombination repair , 2009, Nature Cell Biology.

[33]  M. J. Neale,et al.  Distinct requirements for the Rad32(Mre11) nuclease and Ctp1(CtIP) in the removal of covalently bound topoisomerase I and II from DNA. , 2009, Molecular cell.

[34]  Jeremy M. Stark,et al.  Alternative-NHEJ Is a Mechanistically Distinct Pathway of Mammalian Chromosome Break Repair , 2008, PLoS genetics.

[35]  L. Symington,et al.  Unique and overlapping functions of the Exo1, Mre11 and Pso2 nucleases in DNA repair. , 2008, DNA repair.

[36]  Stephen C. West,et al.  Poly(ADP-ribose)-binding zinc finger motifs in DNA repair/checkpoint proteins , 2008, Nature.

[37]  S. Tsuji,et al.  Aprataxin, causative gene product for EAOH/AOA1, repairs DNA single-strand breaks with damaged 3′-phosphate and 3′-phosphoglycolate ends , 2007, Nucleic acids research.

[38]  S. West,et al.  Actions of Aprataxin in Multiple DNA Repair Pathways* , 2007, Journal of Biological Chemistry.

[39]  K. Caldecott,et al.  APLF (C2orf13) Is a Novel Human Protein Involved in the Cellular Response to Chromosomal DNA Strand Breaks , 2007, Molecular and Cellular Biology.

[40]  S. West,et al.  The neurodegenerative disease protein aprataxin resolves abortive DNA ligation intermediates , 2006, Nature.

[41]  Nianxiang Zhang,et al.  Snm1-Deficient Mice Exhibit Accelerated Tumorigenesis and Susceptibility to Infection , 2005, Molecular and Cellular Biology.

[42]  Linda Hicke,et al.  Ubiquitin-binding domains , 2005, Nature Reviews Molecular Cell Biology.

[43]  J. Stubbe,et al.  Bleomycins: towards better therapeutics , 2005, Nature Reviews Cancer.

[44]  J. Lupski,et al.  Deficiency in 3′-phosphoglycolate processing in human cells with a hereditary mutation in tyrosyl-DNA phosphodiesterase (TDP1) , 2005, Nucleic acids research.

[45]  H. Arakawa,et al.  DNA Cross-Link Repair Protein SNM1A Interacts with PIAS1 in Nuclear Focus Formation , 2004, Molecular and Cellular Biology.

[46]  I. Demuth,et al.  Human SNM1B is required for normal cellular response to both DNA interstrand crosslink-inducing agents and ionizing radiation , 2004, Oncogene.

[47]  Osamu Onodera,et al.  Aprataxin, a novel protein that protects against genotoxic stress. , 2004, Human molecular genetics.

[48]  Yunmei Ma,et al.  Functional and biochemical dissection of the structure‐specific nuclease ARTEMIS , 2004, The EMBO journal.

[49]  J. Cadet,et al.  Oxidative damage to DNA: formation, measurement and biochemical features. , 2003, Mutation research.

[50]  M. Jasin.  Homologous repair of DNA damage and tumorigenesis:the BRCA connection , 2002, Oncogene.

[51]  W. Hittelman,et al.  hSnm1 Colocalizes and Physically Associates with 53BP1 before and after DNA Damage , 2002, Molecular and Cellular Biology.

[52]  L. Povirk,et al.  Conversion of Phosphoglycolate to Phosphate Termini on 3′ Overhangs of DNA Double Strand Breaks by the Human Tyrosyl-DNA Phosphodiesterase hTdp1* , 2002, The Journal of Biological Chemistry.

[53]  S. West,et al.  Involvement of human polynucleotide kinase in double‐strand break repair by non‐homologous end joining , 2002, The EMBO journal.

[54]  Paul Modrich,et al.  Human Exonuclease I Is Required for 5′ and 3′ Mismatch Repair* , 2002, The Journal of Biological Chemistry.

[55]  Yunmei Ma,et al.  Hairpin Opening and Overhang Processing by an Artemis/DNA-Dependent Protein Kinase Complex in Nonhomologous End Joining and V(D)J Recombination , 2002, Cell.

[56]  John A Tainer,et al.  DNA double-strand break repair from head to tail. , 2002, Current opinion in structural biology.

[57]  E. Pastwa,et al.  In vitro repair of complex unligatable oxidatively induced DNA double-strand breaks by human cell extracts. , 2001, Nucleic acids research.

[58]  A. Fischer,et al.  Artemis, a Novel DNA Double-Strand Break Repair/V(D)J Recombination Protein, Is Mutated in Human Severe Combined Immune Deficiency , 2001, Cell.

[59]  M. Vasconcelos,et al.  Disruption of Mouse SNM1 Causes Increased Sensitivity to the DNA Interstrand Cross-Linking Agent Mitomycin C , 2000, Molecular and Cellular Biology.

[60]  L. Thompson,et al.  XRCC3 promotes homology-directed repair of DNA damage in mammalian cells. , 1999, Genes & development.

[61]  A. Goldberg,et al.  Proteasome inhibitors: valuable new tools for cell biologists. , 1998, Trends in cell biology.

[62]  John Kuriyan,et al.  Structure of the C-Terminal Region of p21WAF1/CIP1 Complexed with Human PCNA , 1996, Cell.

[63]  L. Povirk,et al.  DNA damage and mutagenesis by radiomimetic DNA-cleaving agents: bleomycin, neocarzinostatin and other enediynes. , 1996, Mutation research.

[64]  A. De Biasio,et al.  Proliferating cell nuclear antigen structure and interactions: too many partners for one dancer? , 2013, Advances in protein chemistry and structural biology.

[65]  S. Botchway,et al.  Laser-induced radiation microbeam technology and simultaneous real-time fluorescence imaging in live cells. , 2012, Methods in enzymology.

[66]  M. Porteus.  Design and testing of zinc finger nucleases for use in mammalian cells. , 2008, Methods in molecular biology.