Pathway utilization in response to a site-specific DNA double-strand break in fission yeast

EMBO J. 2003 Mar 17;22(6):1419-30. doi: 10.1093/emboj/cdg119.

Abstract

We have examined the genetic requirements for efficient repair of a site-specific DNA double-strand break (DSB) in Schizosaccharomyces pombe. Tech nology was developed in which a unique DSB could be generated in a non-essential minichromosome, Ch(16), using the Saccharomyces cerevisiae HO-endonuclease and its target site, MATa. DSB repair in this context was predominantly through interchromosomal gene conversion. We found that the homologous recombination (HR) genes rhp51(+), rad22A(+), rad32(+) and the nucleotide excision repair gene rad16(+) were required for efficient interchromosomal gene conversion. Further, DSB-induced cell cycle delay and efficient HR required the DNA integrity checkpoint gene rad3(+). Rhp55 was required for interchromosomal gene conversion; however, an alternative DSB repair mechanism was used in an rhp55Delta background involving ku70(+) and rhp51(+). Surprisingly, DSB-induced minichromosome loss was significantly reduced in ku70Delta and lig4Delta non-homologous end joining (NHEJ) mutant backgrounds compared with wild type. Furthermore, roles for Ku70 and Lig4 were identified in suppressing DSB-induced chromosomal rearrangements associated with gene conversion. These findings are consistent with both competitive and cooperative interactions between components of the HR and NHEJ pathways.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromosomes, Fungal
  • DNA Damage*
  • DNA Repair / genetics*
  • DNA, Fungal / genetics*
  • DNA, Fungal / metabolism
  • Deoxyribonucleases, Type II Site-Specific / genetics
  • Deoxyribonucleases, Type II Site-Specific / metabolism*
  • Fungal Proteins / genetics
  • Fungal Proteins / metabolism*
  • Gamma Rays
  • Gene Conversion
  • Genes, Fungal
  • Models, Biological
  • Mutation
  • Recombination, Genetic
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism
  • Schizosaccharomyces / genetics*
  • Schizosaccharomyces / radiation effects

Substances

  • DNA, Fungal
  • Fungal Proteins
  • Saccharomyces cerevisiae Proteins
  • Deoxyribonucleases, Type II Site-Specific