Cooperation of the Cockayne Syndrome Group B Protein and Poly(ADP-Ribose) Polymerase 1 in the Response to Oxidative Stress

Abstract

Cockayne syndrome (CS) is a rare genetic disorder characterized as a segmental premature-aging syndrome. The CS group B (CSB) protein has previously been implicated in transcription-coupled repair, transcriptional elongation, and restoration of RNA synthesis after DNA damage. Recently, evidence for a role of CSB in base excision repair of oxidative DNA lesions has accumulated. In our search to understand the molecular function of CSB in this process, we identify a physical and functional interaction between CSB and poly(ADP-ribose) polymerase-1 (PARP-1). PARP-1 is a nuclear enzyme that protects the integrity of the genome by responding to oxidative DNA damage and facilitating DNA repair. PARP-1 binds to single-strand DNA breaks which activate the catalytic ability of PARP-1 to add polymers of ADP-ribose to various proteins. We find that CSB is present at sites of activated PARP-1 after oxidative stress, identify CSB as a new substrate of PARP-1, and demonstrate that poly(ADP-ribosyl)ation of CSB inhibits its DNA-dependent ATPase activity. Furthermore, we find that CSB-deficient cell lines are hypersensitive to inhibition of PARP. Our results implicate CSB in the PARP-1 poly(ADP-ribosyl)ation response after oxidative stress and thus suggest a novel role of CSB in the cellular response to oxidative damage.

ACKNOWLEDGMENTS

We thank Gilbert de Murcia for providing purified PARP-1, Jean-Marc Egly for providing CSB antibodies, and Wim Vermeulen for providing the pSLME6-dtCSB vector. Ulla Birk Henriksen is thanked for supreme technical assistance. David M. Wilson III is thanked for critical reading of the manuscript. Wen-Hsing Cheng is thanked for help with the immunofluorescence studies.

This work was supported by the Danish Medical Research Council (grant no. 22-03-0253). T.T. was supported by the Danish Medical Research Council (grant no. 22-02-0104) and the Knud Højgaard Foundation. M.C. was supported by the Carlsberg Foundation.