title: Direct Binding to Replication Protein A (RPA)-coated Single-stranded DNA Allows Recruitment of the ATR Activator TopBP1 to Sites of DNA Damage.
abstract: A critical event for the ability of cells to tolerate DNA damage and replication stress is activation of the ATR kinase. ATR activation is dependent on the BRCT (BRCA1 C terminus) repeat-containing protein TopBP1. Previous work has shown that recruitment of TopBP1 to sites of DNA damage and stalled replication forks is necessary for downstream events in ATR activation; however, the mechanism for this recruitment was not known. Here, we use protein binding assays and functional studies in Xenopus egg extracts to show that TopBP1 makes a direct interaction, via its BRCT2 domain, with RPA-coated single-stranded DNA. We identify a point mutant that abrogates this interaction and show that this mutant fails to accumulate at sites of DNA damage and that the mutant cannot activate ATR. These data thus supply a mechanism for how the critical ATR activator, TopBP1, senses DNA damage and stalled replication forks to initiate assembly of checkpoint signaling complexes.
authors.affiliation: From the Molecular and Computational Biology Section, Department of Biological Sciences, University of Southern California, Los Angeles, California 90089 and.
authors.forename: Julyana
authors.initials: J
authors.lastname: Acevedo
authors.affiliation: the Department of Biological Sciences, University of North Carolina, Charlotte, North Carolina 28223.
authors.forename: Shan
authors.initials: S
authors.lastname: Yan
authors.affiliation: From the Molecular and Computational Biology Section, Department of Biological Sciences, University of Southern California, Los Angeles, California 90089 and mattm@usc.edu.
authors.forename: W Matthew
authors.initials: WM
authors.lastname: Michael
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry*; D004249:DNA Damage; D004277:DNA, Single-Stranded / Q000737:chemistry*; D004268:DNA-Binding Proteins; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051716:Replication Protein A / Q000737:chemistry*; D014982:Xenopus laevis
chemical_list: D002352:Carrier Proteins; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D051716:Replication Protein A; C509993:TopBP1 protein, Xenopus
keywords: ATR kinase; BRCT domain; DNA damage response; DNA repair; DNA replication; RPA; TopBP1; cell cycle; checkpoint control; ssDNA