RESUMO
SIN3 gene product operates as a repressor for a huge amount of genes in Saccharomyces cerevisiae. Sin3 protein with a mass of about 175 kDa is a member of the RPD3 protein complex with an assessed mass of greater than 2 million Da. It was previously shownthat RPD3 gene mutations influence recombination and repair processes in S. cerevisiae yeasts. We studied the impacts of the sin3 mutation on UV-light sensitivity and UV-induced mutagenesis in budding yeast cells. The deletion ofthe SIN3 gene causes weak UV-sensitivity of mutant budding cells as compared to the wild-type strain. These results show that the sin3 mutation decreases both spontaneous and UV-induced levels of levels. This fact is hypothetically related to themalfunction of ribonucleotide reductase activity regulation, which leads to a decrease in the dNTP pool and the inaccurate error-prone damage bypass postreplication repair pathway, which in turn provokes a reduction in the incidence of mutations.
Assuntos
Replicação do DNA/efeitos da radiação , Histona Desacetilases/genética , Proteínas Repressoras/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Transcrição Gênica , Reparo do DNA , Replicação do DNA/genética , Regulação Fúngica da Expressão Gênica/efeitos da radiação , Mutagênese/genética , Mutagênese/efeitos da radiação , Mutação , Raios UltravioletaRESUMO
Long-term storage at +4 degrees C and cultivation at +30 degrees C changes the spontaneous mutation rate of the yeast Saccharomyces cerevisiae double mutants rad52hsm3delta and rad52hsm6-1. Combinations of hsm3 and hsm6 mutations with the rad52 mutation lead to a decrease of the spontaneous mutation rate mediated by DNA repair synthesis in multiply replanted strains in comparison with the same strains investigated right after RAD52 gene decay. Combinations of hsm3 and hsm6 mutations with mutations in other genes of the RAD52 epistatic group did not provide a spontaneous mutation rate decrease.
Assuntos
Chaperonas Moleculares/genética , Proteína Rad52 de Recombinação e Reparo de DNA/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Reparo do DNA/genética , Replicação do DNA/genética , Mutação , Taxa de Mutação , TemperaturaRESUMO
Previously, we isolated mutant yeasts Saccharomyces cerevisiae with an increased rate of spontaneous mutagenesis. Here, we studied the properties of HSM6 gene, the hsm6-1 mutation of which increased the frequency of UV-induced mutagenesis and decreased the level of UV-induced mitotic crossover at the centromere gene region, ADE2. HSM6 gene was mapped on the left arm of chromosome 11 in the region where the PSY4 gene is located. The epistatic analysis has shown that the hsm6-1 mutation represents an allele of PSY4 gene. Sequencing of hsm6-1 mutant allele has revealed a frameshift mutation, which caused the substitution of Lys218Glu and the generation of a stop codon in the next position. The interactions of hsm6-1 and rad52 mutations were epistatic. Our data show that the PSY4 gene plays a key role in the regulation of cell withdrawal from checkpoint induced by DNA disturbances.
Assuntos
Centrômero/metabolismo , Mutagênese , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Alelos , Pareamento Incorreto de Bases/genética , Centrômero/genética , Centrômero/efeitos da radiação , Códon de Terminação , Reparo do DNA/genética , Inativação Gênica , Mitose/efeitos da radiação , Mutação , Proteína Rad52 de Recombinação e Reparo de DNA/genética , Raios UltravioletaRESUMO
It was assumed previously that the mutator phenotype of the hms3 mutant was determined by processes taking place in the D-loop. As a next step, genetic analysis was performed to study the interactions between the hsm3 mutation and mutations of the genes that control the initial steps of the D-loop formation. The mutations of the MMS4 and XRS2 genes, which initiate the double-strand break formation and subsequent repair, were shown to completely block HSM3-dependent UV-induced mutagenesis. Mutations of the RAD51, RAD52, and RAD54 genes, which are also involved in the D-loop formation, only slightly decreased the level of UV-induced mutagenesis in the hsm3 mutant. Similar results were observed for the interaction of hsm3 with the mph1 mutation, which stabilizes the D-loop. In contrast, the shu1 mutation, which destabilizes the D-loop structure, led to an extremely high level of UV-induced mutagenesis and displayed epistatic interactions with the hsm3 mutation. The results made it possible to assume that the hsm3 mutation destabilizes the D-loop, which is a key substrate of both Rad5- and Rad52-dependent postreplicative repair pathways.
Assuntos
Chaperonas Moleculares/genética , Reparo de DNA por Recombinação/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Quebras de DNA de Cadeia Dupla/efeitos da radiação , Mutagênese , Mutação , Raios UltravioletaRESUMO
In eukaryotes, damage tolerance of matrix DNA is mainly determined by the repair pathway under the control of the RAD6 epistatic group of genes. T this pathway is also a main source of mutations generated by mutagenic factors. The results of our recent studies show that gene HSM3 participating in the control of adaptive mutagenesis increases the frequency of mutations induced by different mutagens. Mutations rad18, rev3, and mms2 controlling various stages of the RAD6 pathway are epistatic with mutation hsm3 that decreases UV-induced mutagenesis to the level typical for single radiation-sensitive mutants. The level of mutagenesis in the double mutant srs2 hsm3 was lower than in both single mutants. Note that a decrease in the level of mutagenesis relative to the single mutant srs2 depends on the mismatch repair, since this level in the triple mutant srs2 hsm3 pms 1 corresponds to that in the single mutant srs2. These data show that the mutator phenotype hsm3 is probably determined by processes occurring in a D loop. In a number of current works, the protein Hsm3 was shown to participate in the assembly of the proteasome complex S26. The assembly of proteasomes is governed by the N-terminal domain. Our results demonstrated that the Hsm3 protein contains at least two domains; the N-terminal part of the domain is responsible for the proteasome assembly, whereas the C-terminal portion of the protein is responsible for mutagenesis.
Assuntos
Epistasia Genética/efeitos da radiação , Chaperonas Moleculares/metabolismo , Mutagênese/efeitos da radiação , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Enzimas de Conjugação de Ubiquitina/metabolismo , Raios Ultravioleta , Reparo do DNA/efeitos da radiação , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , DNA Polimerase Dirigida por DNA/genética , DNA Polimerase Dirigida por DNA/metabolismo , Epistasia Genética/fisiologia , Chaperonas Moleculares/genética , Mutação , Complexo de Endopeptidases do Proteassoma/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Estrutura Terciária de Proteína , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Enzimas de Conjugação de Ubiquitina/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Gene HSM3 encodes the Hsm3 protein involved in the minor branch in the system responsible for the correction of mismatched bases in DNA structure and controls replicative and reparative spontaneous mutagenesis in yeast Saccharomyces cerevisiae. Spontaneous and UV-induced mutagenesis was studied in three mutant alleles of gene HSM3, and repair effectivity of artificial heteroduplexes was assessed in DNA molecule. The resuts of these studies allowed establishment of the protein domain structure of protein Hsm3 and functions of each domain: the N-terminal domain is responsible for binding to mispaired bases, and the C-terminal domain ensures the interaction with other proteins involved in the system of mismatched base correction.
Assuntos
Reparo do DNA/genética , Chaperonas Moleculares/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Alelos , Reparo do DNA/efeitos da radiação , Replicação do DNA/genética , Replicação do DNA/efeitos da radiação , Chaperonas Moleculares/metabolismo , Mutagênese/genética , Mutagênese/efeitos da radiação , Estrutura Terciária de Proteína , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Raios UltravioletaRESUMO
Sensitivity to the lethal action of the anticancer substance cisplatin was studied in the yeast mutants himl, hsm2, hsm3, and hsm6, deficient for repair of spontaneous and induced mutations. The himl and hsm3 mutants were as resistant to the agent under study as the wild-type strain. The survival of the double mutant rad2 hsm3 was higher than that of the single mutant rad2. The hsm2 and hsm6 mutants were more cisplatin-sensitive than the wild type. Cisplatin was shown to have high mutagenic and recombinogenic effects on yeast cells.
