ABSTRACT
Checkpoints, which are integral to the cellular response to DNA damage, coordinate transient cell cycle arrest and the induced expression of DNA repair genes after genotoxic stress. DNA repair ensures cellular survival and genomic stability, utilizing a multipathway network. Here we report evidence that the two systems, DNA damage checkpoint control and DNA repair, are directly connected by demonstrating that the Rad55 double-strand break repair protein of the recombinational repair pathway is a terminal substrate of DNA damage and replication block checkpoints. Rad55p was specifically phosphorylated in response to DNA damage induced by the alkylating agent methyl methanesulfonate, dependent on an active DNA damage checkpoint. Rad55p modification was also observed after gamma ray and UV radiation. The rapid time course of phosphorylation and the recombination defects identified in checkpoint-deficient cells are consistent with a role of the DNA damage checkpoint in activating recombinational repair. Rad55p phosphorylation possibly affects the balance between different competing DNA repair pathways.
Subject(s)
DNA Damage , DNA Repair , DNA, Fungal , DNA-Binding Proteins , Fungal Proteins/genetics , Saccharomyces cerevisiae Proteins , DNA Damage/radiation effects , Fungal Proteins/metabolism , Gene Expression Regulation, Fungal , Phosphorylation , Recombination, Genetic , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Ultraviolet RaysABSTRACT
A new DNA repair gene from Schizosaccharomyces pombe with homology to RecA was identified and characterized. Comparative analysis showed highest similarity to Saccharomyces cerevisiae Rad55p. rhp55(+) (rad homologue pombe 55) encodes a predicted 350-amino-acid protein with an M(r) of 38,000. The rhp55Delta mutant was highly sensitive to methyl methanesulfonate (MMS), ionizing radiation (IR), and, to a lesser degree, UV. These phenotypes were enhanced at low temperatures, similar to deletions in the S. cerevisiae RAD55 and RAD57 genes. Many rhp55Delta cells were elongated with aberrant nuclei and an increased DNA content. The rhp55 mutant showed minor deficiencies in meiotic intra- and intergenic recombination. Sporulation efficiency and spore viability were significantly reduced. Double-mutant analysis showed that rhp55(+) acts in one DNA repair pathway with rhp51(+) and rhp54(+), homologs of the budding yeast RAD51 and RAD54 genes, respectively. However, rhp55(+) is in a different epistasis group for repair of UV-, MMS-, or gamma-ray-induced DNA damage than is rad22(+), a putative RAD52 homolog of fission yeast. The structural and functional similarity suggests that rhp55(+) is a homolog of the S. cerevisiae RAD55 gene and we propose that the functional diversification of RecA-like genes in budding yeast is evolutionarily conserved.
Subject(s)
DNA Repair/genetics , DNA, Fungal/genetics , DNA-Binding Proteins , Escherichia coli/genetics , Fungal Proteins/genetics , Genes, Fungal , Rec A Recombinases/genetics , Recombination, Genetic/genetics , Saccharomyces cerevisiae Proteins , Schizosaccharomyces pombe Proteins , Schizosaccharomyces/genetics , Amino Acid Sequence , DNA Damage , DNA Helicases/genetics , DNA, Fungal/metabolism , Epistasis, Genetic , Fungal Proteins/physiology , Gene Library , Genes, Bacterial , Genetic Complementation Test , Molecular Sequence Data , Rad51 Recombinase , Sequence Alignment , Sequence Deletion , Sequence Homology, Amino Acid , Sequence Homology, Nucleic AcidABSTRACT
We present a refined genetic map of the obligate methylotroph Methylobacillus flagellatum. New, Hfr (high-frequency-of-transfer) donors, and pulsed-field gel electrophoresis, were used to determine that M. flagellatum contains one approximately 3.1-Mb circular chromosome, and no plasmids. A correlation between time-of-entry units and DNA length was established. Using in vivo and in vitro cloning, and sequencing, a number of new genetic markers were identified and mapped; in addition, the nature of some of the previously mapped markers was elucidated.
Subject(s)
Gram-Negative Anaerobic Bacteria/genetics , Chromosome Mapping , Chromosomes, Bacterial , Cloning, Molecular , Genes, Bacterial , Genetic Markers , Transaminases/geneticsABSTRACT
In offsprings of N.C.Y.C. 74, maltose regulatory constitutive MAL6C2 mutation controls alpha-methylglucoside (alpha-mgl) fermentation in the presence of MALx. MAL6C2 MALx system described by ten Berge et al, is analogous in function (polymeric interaction) to, at least, one MGLa gene from the system of complementary alpha-mgl genes MGLa MGLb MGLc identified by ten Berge. Suppressor malx mutation inhibits both the maltose and alpha-mgl activity of MAL6C2 allele. A brief review on participation of maltose genes in alpha-mgl fermentation is presented.
