A newly identified DNA ligase of Saccharomyces cerevisiae involved in RAD52-independent repair of DNA double-strand breaks.
Eukaryotic DNA ligases are ATP-dependent DNA strand-joining enzymes that participate in DNA replication, repair, and recombination. Whereas mammalian cells contain several different DNA ligases, encoded by at least three distinct genes, only one DNA ligase has been detected previously in either budding yeast or fission yeast. Here, we describe a newly identified nonessential Saccharomyces cerevisiae gene that encodes a DNA ligase distinct from the CDC9 gene product. This DNA ligase shares significant amino acid sequence homology with human DNA ligase IV; accordingly, we designate the yeast gene LIG4. Recombinant LIG4 protein forms a covalent enzyme-AMP complex and can join a DNA single-strand break in a DNA/RNA hybrid duplex, the preferred substrate in vitro. Disruption of the LIG4 gene causes only marginally increased cellular sensitivity to several DNA damaging agents, and does not further sensitize cdc9 or rad52 mutant cells. In contrast, lig4 mutant cells have a 1000-fold reduced capacity for correct recircularization of linearized plasmids by illegitimate end-joining after transformation. Moreover, homozygous lig4 mutant diploids sporulate less efficiently than isogenic wild-type cells, and show retarded progression through meiotic prophase I. Spore viability is normal, but lig4 mutants appear to produce a higher proportion of tetrads with only three viable spores. The mutant phenotypes are consistent with functions of LIG4 in an illegitimate DNA end-joining pathway and ensuring efficient meiosis.
Schär
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Imperial Cancer Research Fund, Clare Hall Laboratories, South Mimms, UK.
Herrmann
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Daly
G
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Lindahl
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T
D000595:Amino Acid Sequence; D004249:DNA Damage; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000235:genetics; D004260:DNA Repair / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000502:physiology; D005656:Fungal Proteins / Q000502:physiology; D005800:Genes, Fungal / Q000235:genetics; D008540:Meiosis / Q000235:genetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames / Q000235:genetics; D010641:Phenotype; D010957:Plasmids / Q000235:genetics; D051721:Rad52 DNA Repair and Recombination Protein; D011993:Recombinant Fusion Proteins; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D013172:Spores, Fungal
C000607890:CDC9 protein, S cerevisiae; D004268:DNA-Binding Proteins; C000607891:DNL4 protein, S cerevisiae; D005656:Fungal Proteins; C000607893:LIG4 protein, human; C497197:RAD52 protein, S cerevisiae; D051721:Rad52 DNA Repair and Recombination Protein; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D011088:DNA Ligases; D000072481:DNA Ligase ATP