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The zinc-finger domains of PARP1 cooperate to recognize DNA strand breaks

A Corrigendum to this article was published on 05 August 2015

This article has been updated

Abstract

Poly(ADP-ribose) polymerase 1 (PARP1) is a primary DNA damage sensor whose (ADP-ribose) polymerase activity is acutely regulated by interaction with DNA breaks. Upon activation at sites of DNA damage, PARP1 modifies itself and other proteins by covalent addition of long, branched polymers of ADP-ribose, which in turn recruit downstream DNA repair and chromatin remodeling factors. PARP1 recognizes DNA damage through its N-terminal DNA-binding domain (DBD), which consists of a tandem repeat of an unusual zinc-finger (ZnF) domain. We have determined the crystal structure of the human PARP1-DBD bound to a DNA break. Along with functional analysis of PARP1 recruitment to sites of DNA damage in vivo, the structure reveals a dimeric assembly whereby ZnF1 and ZnF2 domains from separate PARP1 molecules form a strand-break recognition module that helps activate PARP1 by facilitating its dimerization and consequent trans-automodification.

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Figure 1: PARP1-DNA interactions.
Figure 2: DNA-end binding by the ZnF1-ZnF2 complex.
Figure 3: DNA break detection by PARP1 ZnF2.
Figure 4: DNA damage focus formation by PARP1 and its mobility.
Figure 5: Intermolecular dimerization of PARP1 Znf1 and ZnF2 domain.
Figure 6: A mechanism for DNA damage–dependent trans-automodification by PARP1.

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  • 10 July 2015

    In the version of this article initially published, the image in the bottom row of Figure 4a (full-length PARP1-EGFP mutant R138E) was mistakenly replaced with a duplicate of the bottom-row image in Figure 4c (full-length PARP1-EGFP mutant M43D F44D) during preparation of the accepted version of the manuscript, after peer review and editorial evaluation had taken place. The error has been corrected in the HTML and PDF versions of the article.

References

  1. Pion, E. et al. DNA-induced dimerization of poly(ADP-ribose) polymerase-1 triggers its activation. Biochemistry 44, 14670–14681 (2005).

    Article  CAS  PubMed  Google Scholar 

  2. Kirsten, E., Kun, E., Mendeleyev, J. & Ordahl, C.P. Activity assays for poly-ADP ribose polymerase. Methods Mol. Biol. 287, 137–149 (2004).

    CAS  PubMed  Google Scholar 

  3. Huang, K. et al. Analysis of nucleotide sequence-dependent DNA binding of poly(ADP-ribose) polymerase in a purified system. Biochemistry 43, 217–223 (2004).

    Article  CAS  PubMed  Google Scholar 

  4. Shall, S. & de Murcia, G. Poly(ADP-ribose) polymerase-1: what have we learned from the deficient mouse model? Mutat. Res. DNA Repair 460, 1–15 (2000).

    Article  CAS  PubMed  Google Scholar 

  5. Amé, J.C., Spenlehauer, C. & de Murcia, G. The PARP superfamily. Bioessays 26, 882–893 (2004).

    Article  PubMed  Google Scholar 

  6. D'Amours, D., Desnoyers, S., D'Silva, I. & Poirier, G.G. Poly(ADP-ribosyl)ation reactions in the regulation of nuclear functions. Biochem. J. 342, 249–268 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Altmeyer, M., Messner, S., Hassa, P.O., Fey, M. & Hottiger, M.O. Molecular mechanism of poly(ADP-ribosyl)ation by PARP1 and identification of lysine residues as ADP-ribose acceptor sites. Nucleic Acids Res. 37, 3723–3738 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Fontan-Lozano, A. et al. Histone H1 poly[ADP]-ribosylation regulates the chromatin alterations required for learning consolidation. J. Neurosci. 30, 13305–13313 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Messner, S. et al. PARP1 ADP-ribosylates lysine residues of the core histone tails. Nucleic Acids Res. 38, 6350–6362 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Messner, S. & Hottiger, M.O. Histone ADP-ribosylation in DNA repair, replication and transcription. Trends Cell Biol. 21, 534–542 (2011).

    Article  CAS  PubMed  Google Scholar 

  11. Mathis, G. & Althaus, F.R. Release of core DNA from nucleosomal core particles following (ADP-ribose)N-modification in vitro. Biochem. Biophys. Res. Commun. 143, 1049–1054 (1987).

    Article  CAS  PubMed  Google Scholar 

  12. Althaus, F.R. Poly (ADP-ribose) and chromatin organization in DNA excision repair. Br. J. Cancer 56, 176–176 (1987).

    Google Scholar 

  13. Karras, G.I. et al. The macro domain is an ADP-ribose binding module. EMBO J. 24, 1911–1920 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Timinszky, G. et al. A macrodomain-containing histone rearranges chromatin upon sensing PARP1 activation. Nat. Struct. Mol. Biol. 16, 923–929 (2009).

    Article  CAS  PubMed  Google Scholar 

  15. Ahel, D. et al. Poly(ADP-ribose)-dependent regulation of DNA repair by the chromatin remodeling enzyme ALC1. Science 325, 1240–1243 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Gottschalk, A.J. et al. Poly(ADP-ribosyl)ation directs recruitment and activation of an ATP-dependent chromatin remodeler. Proc. Natl. Acad. Sci. USA 106, 13770–13774 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Ahel, I. et al. Poly(ADP-ribose)-binding zinc finger motifs in DNA repair/checkpoint proteins. Nature 451, 81–85 (2008).

    Article  CAS  PubMed  Google Scholar 

  18. Bryant, H.E. et al. Specific killing of BRCA2-deficient tumours with inhibitors of poly(ADP-ribose) polymerase. Nature 434, 913–917 (2005).

    Article  CAS  PubMed  Google Scholar 

  19. Farmer, H. et al. Targeting the DNA repair defect in BRCA mutant cells as a therapeutic strategy. Nature 434, 917–921 (2005).

    Article  CAS  PubMed  Google Scholar 

  20. Lilyestrom, W., van der Woerd, M.J., Clark, N. & Luger, K. Structural and biophysical studies of human PARP-1 in complex with damaged DNA. J. Mol. Biol. 395, 983–994 (2010).

    Article  CAS  PubMed  Google Scholar 

  21. Eustermann, S. et al. The DNA-binding domain of human PARP-1 interacts with DNA single-strand breaks as a monomer through its second zinc finger. J. Mol. Biol. 407, 149–170 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Langelier, M.F., Planck, J.L., Roy, S. & Pascal, J.M. Crystal structures of poly(ADP-ribose) polymerase-1 (PARP-1) zinc fingers bound to DNA: structural and functional insights into DNA-dependent PARP-1 activity. J. Biol. Chem. 286, 10690–10701 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Gradwohl, G. et al. The second zinc-finger domain of poly(ADP-ribose) polymerase determines specificity for single-stranded breaks in DNA. Proc. Natl. Acad. Sci. USA 87, 2990–2994 (1990).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Panzeter, P.L. & Althaus, F.R. DNA strand break-mediated partitioning of poly(ADP-ribose) polymerase function. Biochemistry 33, 9600–9605 (1994).

    Article  CAS  PubMed  Google Scholar 

  25. Kim, J.W., Kim, K., Kang, K. & Joe, C.O. Inhibition of homodimerization of poly(ADP-ribose) polymerase by its C-terminal cleavage products produced during apoptosis. J. Biol. Chem. 275, 8121–8125 (2000).

    Article  CAS  PubMed  Google Scholar 

  26. Mendoza-Alvarez, H. & Alvarez-Gonzalez, R. Poly(ADP-ribose) polymerase is a catalytic dimer and the automodification reaction is intermolecular. J. Biol. Chem. 268, 22575–22580 (1993).

    CAS  PubMed  Google Scholar 

  27. Bauer, P.I., Buki, K.G., Hakam, A. & Kun, E. Macromolecular association of ADP-ribosyltransferase and its correlation with enzymic activity. Biochem. J. 270, 17–26 (1990).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Pion, E. et al. Poly(ADP-ribose) polymerase-1 dimerizes at a 5′ recessed DNA end in vitro: a fluorescence study. Biochemistry 42, 12409–12417 (2003).

    Article  CAS  PubMed  Google Scholar 

  29. Langelier, M.F., Ruhl, D.D., Planck, J.L., Kraus, W.L. & Pascal, J.M. The Zn3 domain of human poly(ADP-ribose) polymerase-1 (PARP-1) functions in both DNA-dependent poly(ADP-ribose) synthesis activity and chromatin compaction. J. Biol. Chem. 285, 18877–18887 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. de Murcia, G. & Menissier de Murcia, J. Poly(ADP-ribose) polymerase: a molecular nick-sensor. Trends Biochem. Sci. 19, 172–176 (1994).

    Article  CAS  PubMed  Google Scholar 

  31. D'Silva, I. et al. Relative affinities of poly(ADP-ribose) polymerase and DNA- dependent protein kinase for DNA strand interruptions. Biochim. Biophys. Acta 1430, 119–126 (1999).

    Article  CAS  PubMed  Google Scholar 

  32. Potaman, V.N., Shlyakhtenko, L.S., Oussatcheva, E.A., Lyubchenko, Y.L. & Soldatenkov, V.A. Specific binding of poly(ADP-ribose) polymerase-1 to cruciform hairpins. J. Mol. Biol. 348, 609–615 (2005).

    Article  CAS  PubMed  Google Scholar 

  33. Lonskaya, I. et al. Regulation of poly(ADP-ribose) polymerase-1 by DNA structure-specific binding. J. Biol. Chem. 280, 17076–17083 (2005).

    Article  CAS  PubMed  Google Scholar 

  34. Bryant, H.E. et al. PARP is activated at stalled forks to mediate Mre11-dependent replication restart and recombination. EMBO J. 28, 2601–2615 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Ikejima, M. et al. The zinc fingers of human poly(ADP-ribose) polymerase are differentially required for the recognition of DNA breaks and nicks and the consequent enzyme activation. Other structures recognize intact DNA. J. Biol. Chem. 265, 21907–21913 (1990).

    CAS  PubMed  Google Scholar 

  36. Kim, M.Y., Mauro, S., Gevry, N., Lis, J.T. & Kraus, W.L. NAD+-dependent modulation of chromatin structure and transcription by nucleosome binding properties of PARP-1. Cell 119, 803–814 (2004).

    Article  CAS  PubMed  Google Scholar 

  37. Tulin, A. & Spradling, A. Chromatin loosening by poly(ADP)-ribose polymerase (PARP) at Drosophila puff loci. Science 299, 560–562 (2003).

    Article  CAS  PubMed  Google Scholar 

  38. Ju, B.G. et al. Activating the PARP-1 sensor component of the groucho/ TLE1 corepressor complex mediates a CaMKinase IIδ-dependent neurogenic gene activation pathway. Cell 119, 815–829 (2004).

    Article  CAS  PubMed  Google Scholar 

  39. David, K.K., Andrabi, S.A., Dawson, T.M. & Dawson, V.L. Parthanatos, a messenger of death. Front. Biosci. 14, 1116–1128 (2009).

    Article  CAS  Google Scholar 

  40. Kotova, E., Jarnik, M. & Tulin, A.V. Uncoupling of the transactivation and transrepression functions of PARP1 protein. Proc. Natl. Acad. Sci. USA 107, 6406–6411 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Audebert, M., Salles, B. & Calsou, P. Involvement of poly(ADP-ribose) polymerase-1 and XRCC1/DNA ligase III in an alternative route for DNA double-strand breaks rejoining. J. Biol. Chem. 279, 55117–55126 (2004).

    Article  CAS  PubMed  Google Scholar 

  42. Audebert, M., Salles, B., Weinfeld, M. & Calsou, P. Involvement of polynucleotide kinase in a poly(ADP-ribose) polymerase-1-dependent DNA double-strand breaks rejoining pathway. J. Mol. Biol. 356, 257–265 (2006).

    Article  CAS  PubMed  Google Scholar 

  43. Audebert, M., Salles, B. & Calsou, P. Effect of double-strand break DNA sequence on the PARP-1 NHEJ pathway. Biochem. Biophys. Res. Commun. 369, 982–988 (2008).

    Article  CAS  PubMed  Google Scholar 

  44. Cock, J.M., Vanoosthuyse, V. & Gaude, T. Receptor kinase signalling in plants and animals: distinct molecular systems with mechanistic similarities. Curr. Opin. Cell Biol. 14, 230–236 (2002).

    Article  CAS  PubMed  Google Scholar 

  45. Pellegrini, L. Role of heparan sulfate in fibroblast growth factor signalling: a structural view. Curr. Opin. Struct. Biol. 11, 629–634 (2001).

    Article  CAS  PubMed  Google Scholar 

  46. Ali, M.M. et al. Structure of the Ire1 autophosphorylation complex and implications for the unfolded protein response. EMBO J. 30, 894–905 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Leslie, A.G.W. MOSFLM Users Guide (MRC Laboratory of Molecular Biology, Cambridge, UK, 1995).

    Google Scholar 

  48. CCP4. Programs for protein crystallography. Acta Crystallogr. D Biol. Crystallogr. 50, 760–763 (1994).

  49. McCoy, A.J. Solving structures of protein complexes by molecular replacement with Phaser. Acta Crystallogr. D Biol. Crystallogr. 63, 32–41 (2007).

    Article  CAS  PubMed  Google Scholar 

  50. Adams, P.D. et al. Recent developments in the PHENIX software for automated crystallographic structure determination. J. Synchrotron Radiat. 11, 53–55 (2004).

    Article  CAS  PubMed  Google Scholar 

  51. Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D Biol. Crystallogr. 60, 2126–2132 (2004).

    Article  PubMed  Google Scholar 

  52. Chen, V.B. et al. MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr. D Biol. Crystallogr. 66, 12–21 (2010).

    Article  CAS  PubMed  Google Scholar 

  53. Zufferey, R., Nagy, D., Mandel, R.J., Naldini, L. & Trono, D. Multiply attenuated lentiviral vector achieves efficient gene delivery in vivo. Nat. Biotechnol. 15, 871–875 (1997).

    Article  CAS  PubMed  Google Scholar 

  54. Hacham, H. & Ben-Ishai, R. Determination of poly(ADP-ribose) chain length distribution on polyacrylamide gels by silver staining. Anal. Biochem. 184, 83–85 (1990).

    Article  CAS  PubMed  Google Scholar 

  55. Malanga, M., Bachmann, S., Panzeter, P.L., Zweifel, B. & Althaus, F.R. Poly(ADP-ribose) quantification at the femtomole level in mammalian cells. Anal. Biochem. 228, 245–251 (1995).

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We thank M. Roe for assistance with X-ray data collection and K. Caldecott for useful discussion. We are grateful to the Diamond Light Source Ltd., Didcot, UK, for access to synchrotron radiation. We thank the Advanced Light Microscopy Facility (ALMF) at the European Molecular Biology Laboratory (EMBL) and Olympus Europe for supporting EMBL's ALMF. This work was supported by EMBL, the Human Frontiers Science Program and the EU FP6 Marie Curie Research and Training Network “Chromatin Plasticity” to A.G.L. and a Cancer Research UK Programme Grant (C302/A8265) to L.H.P. L.H.P. is supported by a Wellcome Trust Senior Investigator Award.

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Contributions

A.A.E.A. purified the protein, crystallized the complex and collected the X-ray diffraction data; G.T. designed and constructed the PARP1-EGFP constructs and performed the laser DNA-damage experiments; M.K. performed the FRAP experiments; P.O.H. engineered the knockdown PARP1 cell line and the wild-type imaging reporter constructs and performed the in vitro complementation assays; M.H. and R.A.-B. engineered and purified the mutant PARP1 constructs; A.G.L. designed the study and analyzed the data; L.H.P. designed the study, analyzed the data and wrote the paper; A.W.O. designed the study, made the baculovirus constructs, performed the EMSA experiments, designed the purification protocol and solved and refined the crystal structure. All authors discussed the results and commented on the manuscript.

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Correspondence to Andreas G Ladurner, Laurence H Pearl or Antony W Oliver.

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Ali, A., Timinszky, G., Arribas-Bosacoma, R. et al. The zinc-finger domains of PARP1 cooperate to recognize DNA strand breaks. Nat Struct Mol Biol 19, 685–692 (2012). https://doi.org/10.1038/nsmb.2335

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