Differential suppression of DNA repair deficiencies of Yeast rad50, mre11 and xrs2 mutants by EXO1 and TLC1 (the RNA component of telomerase).
Rad50, Mre11, and Xrs2 form a nuclease complex that functions in both nonhomologous end-joining (NHEJ) and recombinational repair of DNA double-strand breaks (DSBs). A search for highly expressed cDNAs that suppress the DNA repair deficiency of rad50 mutants yielded multiple isolates of two genes: EXO1 and TLC1. Overexpression of EXO1 or TLC1 increased the resistance of rad50, mre11, and xrs2 mutants to ionizing radiation and MMS, but did not increase resistance in strains defective in recombination (rad51, rad52, rad54, rad59) or NHEJ only (yku70, sir4). Increased Exo1 or TLC1 RNA did not alter checkpoint responses or restore NHEJ proficiency, but DNA repair defects of yku70 and rad27 (fen) mutants were differentially suppressed by the two genes. Overexpression of Exo1, but not mutant proteins containing substitutions in the conserved nuclease domain, increased recombination and suppressed HO and EcoRI endonuclease-induced killing of rad50 strains. exo1 rad50 mutants lacking both nuclease activities exhibited a high proportion of enlarged, G2-arrested cells and displayed a synergistic decrease in DSB-induced plasmid:chromosome recombination. These results support a model in which the nuclease activity of the Rad50/Mre11/Xrs2 complex is required for recombinational repair, but not NHEJ. We suggest that the 5'-3' exo activity of Exo1 is able to substitute for Rad50/Mre11/Xrs2 in rescission of specific classes of DSB end structures. Gene-specific suppression by TLC1, which encodes the RNA subunit of the yeast telomerase complex, demonstrates that components of telomerase can also impact on DSB repair pathways.
Lewis
L Kevin
LK
Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA.
Karthikeyan
G
G
Westmoreland
James W
JW
Resnick
Michael A
MA
D034961:Antigens, Nuclear; D004265:DNA Helicases; D004260:DNA Repair / Q000502:physiology*; D004271:DNA, Fungal / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000502:physiology; D004706:Endodeoxyribonucleases / Q000235:genetics / Q000502:physiology; D005090:Exodeoxyribonucleases / Q000235:genetics / Q000502:physiology; D045585:Flap Endonucleases; D005656:Fungal Proteins / Q000235:genetics / Q000502:physiology; D000072200:Ku Autoantigen; D009687:Nuclear Proteins / Q000502:physiology; D012331:RNA, Fungal / Q000378:metabolism; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D019098:Telomerase / Q000235:genetics / Q000502:physiology*; D016615:Telomere
D034961:Antigens, Nuclear; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D009687:Nuclear Proteins; C086930:RAD50 protein, S cerevisiae; D012331:RNA, Fungal; D029701:Saccharomyces cerevisiae Proteins; C086929:XRS2 protein, S cerevisiae; C520769:YKU70 protein, S cerevisiae; C081647:high affinity DNA-binding factor, S cerevisiae; D019098:Telomerase; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases; D045585:Flap Endonucleases; C094191:MRE11 protein, S cerevisiae; C028450:exodeoxyribonuclease I; C468323:RAD27 protein, S cerevisiae; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen