Mre11 and Ku regulation of double-strand break repair by gene conversion and break-induced replication.
The yeast Mre11-Rad50-Xrs2 (MRX) and Ku complexes regulate single-strand resection at DNA double-strand breaks (DSB), a key early step in homologous recombination (HR). A prior plasmid gap repair study showed that mre11 mutations, which slow single-strand resection, reduce gene conversion tract lengths and the frequency of associated crossovers. Here we tested whether mre11Delta or nuclease-defective mre11 mutations reduced gene conversion tract lengths during HR between homologous chromosomes in diploid yeast. We found that mre11 mutations reduced the efficiency of HR but did not reduce tract lengths or crossovers, despite substantially reduced end-resection at the test (ura3) locus. End-resection is increased in yku70Delta, but this change also had no effect on tract lengths. Thus, heteroduplex formation and tract lengths are not regulated by the extent of end-resection during DSB repair in a chromosomal context. In a plasmid-chromosome DSB repair assay, tract lengths were again similar in wild-type and mre11Delta, but they were reduced in mre11Delta in a gap repair assay. These results indicate that tract lengths are not affected by the extent of end processing when broken ends can invade nearby sites, perhaps because MRX coordination of the two broken ends is dispensable when ends invade nearby sites. Although HR outcome was largely unaffected in mre11 mutants, break-induced replication (BIR) and chromosome loss increased, suggesting that Mre11 function in mitotic HR is limited to early HR stages. Interestingly, yku70Delta suppressed BIR in mre11 mutants. BIR is also elevated in rad51 mutants, but yku70Delta did not suppress BIR in a rad51 background. These results indicate that Mre11 functions in Rad51-independent BIR, and that Ku functions in Rad51-dependent BIR.
Krishna
Sanchita
S
Department of Molecular Genetics and Microbiology and Cancer Research and Treatment Center, University of New Mexico School of Medicine, Albuquerque, NM 87131, United States.
Wagener
Brant M
BM
Liu
Hui Ping
HP
Lo
Yi-Chen
YC
Sterk
Rosa
R
Petrini
John H J
JH
Nickoloff
Jac A
JA
D000483:Alleles; D034961:Antigens, Nuclear / Q000502:physiology*; D002874:Chromosome Mapping; D015825:Chromosomes, Fungal; D053903:DNA Breaks, Double-Stranded; D004249:DNA Damage; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000502:physiology*; D004706:Endodeoxyribonucleases / Q000502:physiology*; D005090:Exodeoxyribonucleases / Q000502:physiology*; D005785:Gene Conversion; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D005838:Genotype; D000072200:Ku Autoantigen; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000502:physiology*
D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases; C094191:MRE11 protein, S cerevisiae; D000072200:Ku Autoantigen