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PMID: 15929725 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, U.S. Gov't, P.H.S.

Preferential localization of hyperphosphorylated replication protein A to double-strand break repair and checkpoint complexes upon DNA damage.

The Biochemical journal ·Vol. 391 ·No. Pt 3 ·2005-11-01 ·Pages 473-80

Wu X, Yang Z, Liu Y, Zou Y

Abstract

RPA (replication protein A) is an essential factor for DNA DSB (double-strand break) repair and cell cycle checkpoint activation. The 32 kDa subunit of RPA undergoes hyperphosphorylation in response to cellular genotoxic insults. However, the potential involvement of hyperphosphorylated RPA in DSB repair and checkpoint activation remains unclear. Using co-immunoprecipitation assays, we showed that cellular interaction of RPA with two DSB repair factors, Rad51 and Rad52, was predominantly mediated by the hyperphosphorylated species of RPA in cells after UV and camptothecin treatment. Moreover, Rad51 and Rad52 displayed higher affinity for the hyperphosphorylated RPA than native RPA in an in vitro binding assay. Checkpoint kinase ATR (ataxia telangiectasia mutated and Rad3-related) also interacted more efficiently with the hyperphosphorylated RPA than with native RPA following DNA damage. Consistently, immunofluorescence microscopy demonstrated that the hyperphosphorylated RPA was able to co-localize with Rad52 and ATR to form significant nuclear foci in cells. Our results suggest that hyperphosphorylated RPA is preferentially localized to DSB repair and the DNA damage checkpoint complexes in response to DNA damage.

MeSH Terms
Adenocarcinoma/genetics,metabolism Cell Line, Tumor DNA Damage/genetics,radiation effects DNA Repair Gene Expression Regulation Humans Lung Neoplasms/genetics,metabolism Phosphorylation Protein Binding Protein Transport Rad51 Recombinase/metabolism Rad52 DNA Repair and Recombination Protein/metabolism Replication Protein A/metabolism Ultraviolet Rays
Chemicals
Rad52 DNA Repair and Recombination Protein Replication Protein A Rad51 Recombinase
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Wu Xiaoming
Department of Biochemistry and Molecular Biology, James H. Quillen College of Medicine, East Tennessee State University, Johnson City, TN 37614, USA.
Yang Zhengguan
Liu Yiyong
Zou Yue
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Article Info
Journal
The Biochemical journal
Abbr.
Biochem J
ISSN
1470-8728
DOI
10.1042/BJ20050379
Published
2005-11-01
Pages
473-80
Language
English
Region
England
NLM ID
2984726R
PMCID
PMC1276948
Subset
IM
Grants
NCI NIH HHS · R01 CA086927 · United States
NCI NIH HHS · R56 CA086927 · United States
NCI NIH HHS · CA86927 · United States
External Links
PubMed source DOI: 10.1042/BJ20050379 PMC Full Text View journal details
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