Responses of radiation-sensitive mutants of Saccharomyces cerevisiae to lethal effects of bleomycin

Moore, Carol W.

doi:10.1016/s0027-5107(78)80016-5

Cited by 50 publications

(28 citation statements)

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“…A preliminary test showed that higher concentrations of the drug resulted in complete growth inhibition of all strains under our experimental conditions. For survival experiments, solid YEDA medium containing BLM was prepared according to a previously described method (1,22,26 concentrations of the drug (3.75 to 30 ,g/ml) were employed. Although plates were used within 24 h after preparation, similar surviving fractions were obtained with plates used 10 days after preparation.…”

Section: Methodsmentioning

confidence: 99%

“…With phages, bacteria, and yeast cells, it was demonstrated that this drug is recombinogenic and mutagenic (11,23,39,42). Analysis of BLM-induced mutagenesis in repackaged lambda phage suggests SOS repair dependence (33).BLM-induced DNA damage in prokaryotic and eukaryotic cells is partially reparable (3,22,27,40). For human fibroblasts, a long-patch mode of excision repair of BLM-induced damage was recently described (4, 5).…”

mentioning

confidence: 99%

“…BLM-induced DNA damage in prokaryotic and eukaryotic cells is partially reparable (3,22,27,40). For human fibroblasts, a long-patch mode of excision repair of BLM-induced damage was recently described (4,5).…”

mentioning

confidence: 99%

“…For human fibroblasts, a long-patch mode of excision repair of BLM-induced damage was recently described (4,5). Using haploid and diploid radiosensitive strains of Saccharomyces cerevisiae, Moore (22) showed that all rad single mutants sensitive to BLM tested, with the exception of rad1S, are sensitive to X rays. Likewise, BLM-sensitive (blm) mutants exhibit crosssensitivity to ionizing radiation and hydrogen peroxide (26).…”

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Effects of bleomycin on growth kinetics and survival of Saccharomyces cerevisiae: a model of repair pathways

1992

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In order to analyze the roles of some repair genes in the processing of bleomycin-induced DNA damage and, especially, the interrelationships among the involved repair pathways, we investigated the potentially lethal effect of bleomycin on radiosensitive mutants of Saccharomyces cerevisiae defective in recombination, excision, and R4D6-dependent DNA repair. Using single, double, and triple rad mutants, we analyzed growth kinetics and survival curves as a function of bleomycin concentration. Our results indicate that genes belonging to the three epistasis groups interact in the repair of bleomycin-induced DNA damage to different degrees depending on the concentration of bleomycin. The most important mechanisms involved are recombination and postreplication repair. The initial action of a potentially inducible excision repair gene could provide intermediate substrates for the R4D6-and RAD52-dependent repair processes. Interaction between R4D6 and RADS2 genes was epistatic for low bleomycin concentrations. RA1D3 and RAD52 genes act independently in processing DNA damage induced by high concentrations of bleomycin. The synergistic interaction observed at high concentrations in the triple mutant rad2-6 rad6-1 rad52-1 indicates partial independence of the involved repair pathways, with possible common substrates. On the basis of the present results, we propose a heuristic model of bleomycin-induced DNA damage repair.Bleomycin (BLM) is a radiomimetic glycopeptide antibiotic used extensively in tumor therapy (32,48). This drug binds to DNA and through a free-radical-based mechanism produces DNA base loss and single-and double-strand breaks (43,44). In different lines of eukaryotic cells, BLM produces inhibition of DNA synthesis (10) as well as cell cycle G2 delay (19,38). With phages, bacteria, and yeast cells, it was demonstrated that this drug is recombinogenic and mutagenic (11,23,39,42). Analysis of BLM-induced mutagenesis in repackaged lambda phage suggests SOS repair dependence (33).BLM-induced DNA damage in prokaryotic and eukaryotic cells is partially reparable (3,22,27,40). For human fibroblasts, a long-patch mode of excision repair of BLM-induced damage was recently described (4, 5). Using haploid and diploid radiosensitive strains of Saccharomyces cerevisiae, Moore (22) showed that all rad single mutants sensitive to BLM tested, with the exception of rad1S, are sensitive to X rays. Likewise, BLM-sensitive (blm) mutants exhibit crosssensitivity to ionizing radiation and hydrogen peroxide (26). These results suggest that some aspects of the repair of BLM and ionizing radiation DNA damage in yeast cells may be similar. Furthermore, DNA ligase (the product of the CDC9 gene) is required in the overall process of restoring full-size DNA molecules from DNA fragments produced by BLM (24, 25).We have recently shown that BLM induces at least one error-prone type of repair in S. cerevisiae and that the PS04 (XS9) gene acts as a mutation-triggering factor (39). Furthermore, the combination of UV light and BLM produces di...

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Section: Methodsmentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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Effects of bleomycin on growth kinetics and survival of Saccharomyces cerevisiae: a model of repair pathways

1992

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“…Washed cells, 107 per ml of deionized water (pH 4.5 to 5.0), were incubated with bleomycin at 23°C with aeration, after which EDTA was added to 0.1 M. Cells were subsequently sedimented at 4°C in an RC-SB Sorvall SS34 rotor at 3,000 x g, washed VOL. 33,1989 Chromatin cleavage and degradation are higher during stationary phase. A representative electrophoretic analysis of dose-dependent internucleosomal cleavage and degradation of chromatin from cells incubated with low concentrations of bleomycin during the stationary phase of growth is shown in Fig.…”

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Growth phase dependency of chromatin cleavage and degradation by bleomycin

Moore

Jones

Wall

1989

Antimicrob Agents Chemother

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Preferential cleavage of Saccharomyces cerevisiae chromosomes in internucleosomal (linker) regions and nonspecific degradation of chromatin by an anticancer antibiotic which degrades DNA were investigated and found to increase in consecutive stages of growth. Cleavage of DNA in internucleosomal regions and intensities and multiplicities of nucleosomal bands were dependent on drug concentration, growth phase of the cells, and length of incubation. Cellular DNA was least degraded during logarithmic phase. After cells progressed only one generation in logarithmic phase, low concentrations (6.7 x 10-7 to 3.4 x 10-6 M) of bleomycin produced approximately three to seven times more DNA breaks. Internucleosomal cleavage was highest, and the most extended oligonucleosomal series and extensive chromatin degradation were observed during stationary phase. It is concluded that the growth phase of cells is critical in determining amounts of the highly preferential cleavage in internucleosomal regions and overall breakage and degradation of DNA. Mononucleosomal bands were most intense, indicating the greatest accumulation of DNA of this size. Mean mononucleosomal lengths were 165.9 + 3.9 base pairs, in agreement with yeast mononucleosomal lengths. As high-molecular-weight chromatin was digested by bleomycin, oligonucleosomes and, eventually, mononucleosomes became digested. Therefore, it is also concluded that bleomycin degradation of oligonucleosomes and trimming of DNA linker regions proceed to degradation of the monosomes (core plus linker DNA).We consider in the current investigation that the growth phase of cells could be a crucial factor in determining lesions produced in cellular chromatin. The chromatin conformation characteristic for chromosomal organization in eucaryotic cells certainly can be very important in determining the nature and extent of chemical reactivity with genomic structures. For example, DNA is preferentially cleaved in internucleosomal (linker) regions of chromatin in isolated mammalian nuclei (18,22,23), in intact human K562 cells (46), and in Saccharomyces cerevisiae (40) by the unique chemical action of the low-molecular-weight glycopeptides (Mr, approximately 1,500 to 1,600 [ Fig. 1]) in the bleomycin family of anticancer antibiotics (4,12,14,20,21,30,31,39,47,59,60,62). Bleomycins form single-and double-strand breaks in DNA (e.g., see references 2, 7, 15, 16, 29, 39, 41, 44, and 60 sensitive to killing by bleomycin in plateau phase than when growing exponentially, but the reverse was true for mouse tumor cells in an earlier study (71) and for Chinese hamster V79 (28), Burkitt's lymphoma (32,66), and human colon carcinoma (8) (10,36,39,44,51) from starting inocula of 8 x 106 cells per ml. Cells were harvested by centrifugation at 4°C in a Dupont Sorvall RC-SB SS34 rotor at 2,500 x g and washed twice with deionized water.

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Bleomycin

Moore¹

2002

Encyclopedia of Molecular Biology

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Responses of radiation-sensitive mutants of Saccharomyces cerevisiae to lethal effects of bleomycin

Cited by 50 publications

References 30 publications

Effects of bleomycin on growth kinetics and survival of Saccharomyces cerevisiae: a model of repair pathways

Effects of bleomycin on growth kinetics and survival of Saccharomyces cerevisiae: a model of repair pathways

Growth phase dependency of chromatin cleavage and degradation by bleomycin

Bleomycin

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