Mitochondrial genome maintenance: roles for nuclear nonhomologous end-joining proteins in Saccharomyces cerevisiae.
Mitochondrial DNA (mtDNA) deletions are associated with sporadic and inherited diseases and age-associated neurodegenerative disorders. Approximately 85% of mtDNA deletions identified in humans are flanked by short directly repeated sequences; however, mechanisms by which these deletions arise are unknown. A limitation in deciphering these mechanisms is the essential nature of the mitochondrial genome in most living cells. One exception is budding yeast, which are facultative anaerobes and one of the few organisms for which directed mtDNA manipulation is possible. Using this model system, we have developed a system to simultaneously monitor spontaneous direct-repeat-mediated deletions (DRMDs) in the nuclear and mitochondrial genomes. In addition, the mitochondrial DRMD reporter contains a unique KpnI restriction endonuclease recognition site that is not present in otherwise wild-type (WT) mtDNA. We have expressed KpnI fused to a mitochondrial localization signal to induce a specific mitochondrial double-strand break (mtDSB). Here we report that loss of the MRX (Mre11p, Rad50p, Xrs2p) and Ku70/80 (Ku70p, Ku80p) complexes significantly impacts the rate of spontaneous deletion events in mtDNA, and these proteins contribute to the repair of induced mtDSBs. Furthermore, our data support homologous recombination (HR) as the predominant pathway by which mtDNA deletions arise in yeast, and suggest that the MRX and Ku70/80 complexes are partially redundant in mitochondria.
Department of Biology, University of Rochester, Rochester, New York 14627, USA.
Lidza
L
Kalifa
Daniel F
DF
Quintana
Laura K
LK
Schiraldi
Naina
N
Phadnis
Garry L
GL
Coles
Rey A
RA
Sia
Elaine A
EA
Sia
D034961:Antigens, Nuclear / Q000235:genetics / Q000378:metabolism; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded; D059766:DNA End-Joining Repair / Q000502:physiology*; D004260:DNA Repair; D004272:DNA, Mitochondrial / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D004706:Endodeoxyribonucleases / Q000235:genetics / Q000378:metabolism; D005090:Exodeoxyribonucleases / Q000235:genetics / Q000378:metabolism; D023061:Gene Order; D054629:Genome, Mitochondrial; D000072200:Ku Autoantigen; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism; D008957:Models, Genetic; D059645:Mutation Rate; D009687:Nuclear Proteins / Q000378:metabolism*; D010641:Phenotype; D011995:Recombination, Genetic; D012091:Repetitive Sequences, Nucleic Acid; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D015398:Signal Transduction
D034961:Antigens, Nuclear; D004272:DNA, Mitochondrial; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; C086930:RAD50 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C086929:XRS2 protein, S cerevisiae; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases; C094191:MRE11 protein, S cerevisiae; D015252:Deoxyribonucleases, Type II Site-Specific; C029312:GGTACC-specific type II deoxyribonucleases; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen