Spatiotemporal dynamics of homologous recombination repair at single collapsed replication forks

Nat Commun. 2018 Sep 24;9(1):3882. doi: 10.1038/s41467-018-06435-3.

Abstract

Homologous recombination (HR) is a crucial pathway for the repair of DNA double-strand breaks. BRCA1/2 breast cancer proteins are key players in HR via their mediation of RAD51 nucleofilament formation and function; however, their individual roles and crosstalk in vivo are unknown. Here we use super-resolution (SR) imaging to map the spatiotemporal kinetics of HR proteins, revealing the interdependent relationships that govern the dynamic interplay and progression of repair events. We show that initial single-stranded DNA/RAD51 nucleofilament formation is mediated by RAD52 or, in the absence of RAD52, by BRCA2. In contrast, only BRCA2 can orchestrate later RAD51 recombinase activity during homology search and resolution. Furthermore, we establish that upstream BRCA1 activity is critical for BRCA2 function. Our analyses reveal the underlying epistatic landscape of RAD51 functional dependence on RAD52, BRCA1, and BRCA2 during HR and explain the phenotypic similarity of diseases associated with mutations in these proteins.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • BRCA1 Protein / genetics
  • BRCA1 Protein / metabolism
  • BRCA2 Protein / genetics
  • BRCA2 Protein / metabolism
  • Cell Line, Tumor
  • DNA Breaks, Double-Stranded*
  • DNA, Single-Stranded / genetics
  • DNA, Single-Stranded / metabolism*
  • Gene Knockdown Techniques
  • Humans
  • Image Processing, Computer-Assisted
  • Microscopy, Fluorescence / methods
  • RNA, Small Interfering / metabolism
  • Rad51 Recombinase / metabolism*
  • Rad52 DNA Repair and Recombination Protein / genetics
  • Rad52 DNA Repair and Recombination Protein / metabolism
  • Recombinational DNA Repair*
  • Single Molecule Imaging / methods

Substances

  • BRCA1 Protein
  • BRCA1 protein, human
  • BRCA2 Protein
  • BRCA2 protein, human
  • DNA, Single-Stranded
  • RAD52 protein, human
  • RNA, Small Interfering
  • Rad52 DNA Repair and Recombination Protein
  • RAD51 protein, human
  • Rad51 Recombinase