Abstract
End joining of double-strand breaks (DSBs) requires Ku proteins and frequently involves base pairing between complementary terminal sequences. To define the role of terminal base pairing in end joining, two oppositely oriented HO endonuclease cleavage sites separated by 2.0 kb were integrated into yeast chromosome III, where constitutive expression of HO endonuclease creates two simultaneous DSBs with no complementary end sequence. Lack of complementary sequence in their 3' single-strand overhangs facilitates efficient repair events distinctly different from when the 3' ends have a 4-bp sequence base paired in various ways to create 2- to 3-bp insertions. Repair of noncomplementary ends results in a set of nonrandom deletions of up to 302 bp, annealed by imperfect microhomology of about 8 to 10 bp at the junctions. This microhomology-mediated end joining (MMEJ) is Ku independent, but strongly dependent on Mre11, Rad50, and Rad1 proteins and partially dependent on Dnl4 protein. The MMEJ also occurs when Rad52 is absent, but the extent of deletions becomes more limited. The increased gamma ray sensitivity of rad1Delta rad52Delta yku70Delta strains compared to rad52Delta yku70Delta strains suggests that MMEJ also contributes to the repair of DSBs induced by ionizing radiation.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Antigens, Nuclear / genetics
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Antigens, Nuclear / metabolism
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Base Pairing
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Base Sequence
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Chromosome Breakage
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Chromosomes, Fungal / genetics
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Chromosomes, Fungal / metabolism
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DNA Helicases*
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DNA Ligase ATP
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DNA Ligases / genetics
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DNA Ligases / metabolism
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DNA Repair / genetics
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DNA Repair / physiology*
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DNA Repair Enzymes
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DNA, Fungal / genetics
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DNA, Fungal / metabolism
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism
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Endodeoxyribonucleases / genetics
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Endodeoxyribonucleases / metabolism*
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Endonucleases / genetics
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Endonucleases / metabolism*
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Exodeoxyribonucleases / genetics
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Exodeoxyribonucleases / metabolism*
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Genes, Fungal
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Ku Autoantigen
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Models, Biological
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Molecular Sequence Data
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Mutation
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Rad52 DNA Repair and Recombination Protein
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Saccharomyces cerevisiae / genetics
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Saccharomyces cerevisiae / metabolism
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Saccharomyces cerevisiae Proteins / genetics
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Saccharomyces cerevisiae Proteins / metabolism*
Substances
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Antigens, Nuclear
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DNA, Fungal
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DNA-Binding Proteins
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RAD52 protein, S cerevisiae
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Rad52 DNA Repair and Recombination Protein
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Saccharomyces cerevisiae Proteins
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Endodeoxyribonucleases
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Endonucleases
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Exodeoxyribonucleases
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MRE11 protein, S cerevisiae
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RAD1 protein, S cerevisiae
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DNA Helicases
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XRCC5 protein, human
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Xrcc6 protein, human
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Ku Autoantigen
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DNA Ligases
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DNA Repair Enzymes
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DNA Ligase ATP