Deficient nonhomologous end-joining activity in cell-free extracts from BRCA1-null fibroblasts

Qing Zhong, Thomas G. Boyer, Phang Lang Chen, Wen Hwa Lee

Research output: Contribution to journalArticle

109 Citations (Scopus)

Abstract

BRCA1 ensures genomic stability, at least in part, through a functional role in DNA damage repair. BRCA1 interacts with the Rad50/Mre11/Nbs1 complex that occupies a central role in DNA double-strand break repair mediated by homologous recombination and nonhomologous end joining (NHEJ). NHEJ can be catalyzed by mammalian whole cell extract in a reaction dependent upon DNA ligase IV, Xrcc4, Ku70, Ku80, and DNA-PKcs. Here, we show that under identical cell-free reaction conditions, the addition of antibodies specific for BRCA1 and Rad 50 but not Rad51, inhibits end-joining activity. Cell extracts derived from Brca1-deficient mouse embryonic fibroblasts exhibit reduced end-joining activity independent of the endogenous protein amounts of DNA ligase IV, Ku80, and Ku70. The Brca1-dependent NHEJ activity predominates at the lower concentrations of Mg2+ (0.5 mM); elevated Mg2+ or Mn2+ concentrations (10 mM) dramatically increase overall end-joining activity and abrogates the requirement for Brca1, Xrcc4, and Ku70. The addition of partially purified BRCA1, in association with Rad50/Mre11/Nbs1 complex, complements the NHEJ deficiency of Brca1-null fibroblast extracts. These results suggest a role for Brca1 in NHEJ and in the maintenance of genome integrity.

Original languageEnglish (US)
Pages (from-to)3966-3970
Number of pages5
JournalCancer Research
Volume62
Issue number14
StatePublished - Jul 15 2002

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Cell Extracts
Fibroblasts
Recombinational DNA Repair
Double-Stranded DNA Breaks
Genomic Instability
DNA Repair
DNA Damage
Maintenance
Genome
Antibodies
DNA
Proteins
DNA Ligase ATP

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

Cite this

Deficient nonhomologous end-joining activity in cell-free extracts from BRCA1-null fibroblasts. / Zhong, Qing; Boyer, Thomas G.; Chen, Phang Lang; Lee, Wen Hwa.

In: Cancer Research, Vol. 62, No. 14, 15.07.2002, p. 3966-3970.

Research output: Contribution to journalArticle

Zhong, Qing ; Boyer, Thomas G. ; Chen, Phang Lang ; Lee, Wen Hwa. / Deficient nonhomologous end-joining activity in cell-free extracts from BRCA1-null fibroblasts. In: Cancer Research. 2002 ; Vol. 62, No. 14. pp. 3966-3970.
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