XRCC1 interacts with the p58 subunit of DNA Pol alpha-primase and may coordinate DNA repair and replication during S phase.
Repair of single-stranded DNA breaks before DNA replication is critical in maintaining genomic stability; however, how cells deal with these lesions during S phase is not clear. Using combined approaches of proteomics and in vitro and in vivo protein-protein interaction, we identified the p58 subunit of DNA Pol alpha-primase as a new binding partner of XRCC1, a key protein of the single strand break repair (SSBR) complex. In vitro experiments reveal that the binding of poly(ADP-ribose) to p58 inhibits primase activity by competition with its DNA binding property. Overexpression of the XRCC1-BRCT1 domain in HeLa cells induces poly(ADP-ribose) synthesis, PARP-1 and XRCC1-BRCT1 poly(ADP-ribosyl)ation and a strong S phase delay in the presence of DNA damage. Addition of recombinant XRCC1-BRCT1 to Xenopus egg extracts slows down DNA synthesis and inhibits the binding of PCNA, but not MCM2 to alkylated chromatin, thus indicating interference with the assembly of functional replication forks. Altogether these results suggest a critical role for XRCC1 in connecting the SSBR machinery with the replication fork to halt DNA synthesis in response to DNA damage.
Lévy
Nicolas
N
FRE 3211, Institut de Recherche de l'Ecole de Biotechnologie de Strasbourg, CNRS/Université de Strasbourg, Ecole Supérieure de Biotechnologie de Strasbourg, Boulevard S. Brant, BP 10413, F-67412, Illkirch Cedex, France.
Oehlmann
Maren
M
Delalande
François
F
Nasheuer
Heinz Peter
HP
Van Dorsselaer
Alain
A
Schreiber
Valérie
V
de Murcia
Gilbert
G
Ménissier-de Murcia
Josiane
J
Maiorano
Domenico
D
Bresson
Anne
A
D000818:Animals; D002843:Chromatin / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis; D004249:DNA Damage; D004256:DNA Polymerase I / Q000378:metabolism; D019915:DNA Primase / Q000737:chemistry / Q000378:metabolism*; D004260:DNA Repair; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D054730:Protein Interaction Domains and Motifs; D016196:S Phase / Q000235:genetics*; D000076105:X-ray Repair Cross Complementing Protein 1; D014982:Xenopus laevis
D002843:Chromatin; D004268:DNA-Binding Proteins; D000076105:X-ray Repair Cross Complementing Protein 1; C000619371:XRCC1 protein, human; D004247:DNA; C499320:PARP1 protein, human; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D019915:DNA Primase; C116932:DNA polymerase alpha-primase; C525356:PRIM2 protein, human; D004256:DNA Polymerase I