Pathway utilization in response to a site-specific DNA double-strand break in fission yeast.
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.
Prudden
John
J
MRC Radiation and Genome Stability Unit, Harwell, Didcot, Oxon OX11 0RD, UK.
Evans
Joanne S
JS
Hussey
Sharon P
SP
Deans
Bryan
B
O'Neill
Peter
P
Thacker
John
J
Humphrey
Tim
T
D015825:Chromosomes, Fungal; D004249:DNA Damage; D004260:DNA Repair / Q000235:genetics*; D004271:DNA, Fungal / Q000235:genetics* / Q000378:metabolism; D015252:Deoxyribonucleases, Type II Site-Specific / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D005720:Gamma Rays; D005785:Gene Conversion; D005800:Genes, Fungal; D008954:Models, Biological; D009154:Mutation; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D012568:Schizosaccharomyces / Q000235:genetics* / Q000528:radiation effects
D004271:DNA, Fungal; D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; D015252:Deoxyribonucleases, Type II Site-Specific