PrimPol-dependent single-stranded gap formation mediates homologous recombination at bulky DNA adducts

Nat Commun. 2020 Nov 17;11(1):5863. doi: 10.1038/s41467-020-19570-7.

Abstract

Stalled replication forks can be restarted and repaired by RAD51-mediated homologous recombination (HR), but HR can also perform post-replicative repair after bypass of the obstacle. Bulky DNA adducts are important replication-blocking lesions, but it is unknown whether they activate HR at stalled forks or behind ongoing forks. Using mainly BPDE-DNA adducts as model lesions, we show that HR induced by bulky adducts in mammalian cells predominantly occurs at post-replicative gaps formed by the DNA/RNA primase PrimPol. RAD51 recruitment under these conditions does not result from fork stalling, but rather occurs at gaps formed by PrimPol re-priming and resection by MRE11 and EXO1. In contrast, RAD51 loading at double-strand breaks does not require PrimPol. At bulky adducts, PrimPol promotes sister chromatid exchange and genetic recombination. Our data support that HR at bulky adducts in mammalian cells involves post-replicative gap repair and define a role for PrimPol in HR-mediated DNA damage tolerance.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 4-Nitroquinoline-1-oxide / toxicity
  • 7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide / metabolism
  • Benz(a)Anthracenes / administration & dosage
  • Benz(a)Anthracenes / toxicity
  • Cell Line
  • DNA Adducts / genetics*
  • DNA Adducts / metabolism
  • DNA Primase / genetics
  • DNA Primase / metabolism*
  • DNA, Single-Stranded
  • DNA-Directed DNA Polymerase / genetics
  • DNA-Directed DNA Polymerase / metabolism*
  • Homologous Recombination / physiology*
  • Humans
  • Multifunctional Enzymes / genetics
  • Multifunctional Enzymes / metabolism*
  • Quinolones / toxicity
  • Rad51 Recombinase / genetics
  • Rad51 Recombinase / metabolism
  • Single Molecule Imaging
  • Sister Chromatid Exchange

Substances

  • 4-nitroquinolone-1-oxide
  • Benz(a)Anthracenes
  • DNA Adducts
  • DNA, Single-Stranded
  • Multifunctional Enzymes
  • Quinolones
  • benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide-DNA
  • 7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide
  • 4-Nitroquinoline-1-oxide
  • 10,11-dihydrodiol-8,9-epoxide benzanthracene
  • DNA Primase
  • PrimPol protein, human
  • RAD51 protein, human
  • Rad51 Recombinase
  • DNA-Directed DNA Polymerase