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Biomedical subjects

M Brendel

Publications and source records attributed to M Brendel.

At least 109 records · Page 6Linked to original sources
PubMed1982

Nucleic acid metabolism in yeast VI. Utilisation of exogenous dAMP.

It is shown that mutants of Saccharomyces cerevisiae able to efficiently utilise exogenous dTMP can also utilise exogenous dAMP. Under extracellular conditions permissive for dTMP uptake label stemming from offered [8-3H]dAMP is incorporated preferentially into alkali-resistant, high molecular weight material (putative DNA); only about 30% of high molecular weight cell-bound dAMP label was found to be sensitive towards mild alkali hydrolysis. This putative RNA label can be minimised to practically zero when greater than or equal to mM Ade is employed in a dAMP labelling assay. Exogenous dAMP at much greater than 10 microM was found to be cytostatic similarly to much greater than microM dTMP and similarly to inhibit effectively import of exogenous Pi. We conclude from our results that there exists a yeast cytoplasmic membrane permease able to import dAMP. A model of this hypothetical permease system is presented.

Cell Membrane↗
PubMed1982

The cytotoxic action of activated and non-activated cyclophosphamide in yeast: comparison of induced DNA damage.

The cytotoxic and DNA-damaging effects of cyclophosphamide (CP) and its 'activated' derivative 4-OOH-CP were studied using a series of strains of S. cerevisiae which allow a phenotypical classification of genotoxic characteristics as well as direct physicochemical demonstration of key DNA lesions. The concurring results of biological and biochemical experiments indicate that (i) non-activated CP has a weak but detectable monofunctional alkylating potency, leading to DNA strand breaks and (ii) 4-OOH-CP has the ability to induce both DNA strand breaks and interstrand cross-links. The activity of CP is probably due to spontaneous decomposition in aqueous solution.

Alkylation↗
PubMed1981

Nucleic acid metabolism in yeast II. Metabolism of thymidylate during thymidylate excess death.

A discrete class of strains of Saccharomyces cerevisiae, able to utilize, highly efficiently, exogenous deoxythymidine-5'-monophosphate (dTMP),was found to be sensitive to concentrations greater than 10 Micro M dTMP in an otherwise complete growth medium. Excess dTMP is cytostatic and cytotoxic: 90% of exponentially growing cells lose colony forming ability within 1 h of exposure to excess dTMP is a growth medium. Uptake of dTMP, adenine, histidine, and leucine does occur during this thymidylate excess death (TED). dTMP is anabolized to higher phosphorylated synthesis is blocked under TED-conditions but not RNA and protein biosynthesis.

Biological Transport↗
PubMed1981

Isolation of yeast mutants sensitive to the bifunctional alkylating agent nitrogen mustard.

Mutants of Saccharomyces cerevisiae with enhanced sensitivity to the DNA cross-linking agent nitrogen mustard (HN2) have been isolated and partially characterized with respect to their phenotypic and genetic properties. The screening technique, based on HN2-sensitivity as sole criterion, yields approxiamtely 1 sensitive isolate in 200 clones when applied to an intensively mutagenized population of a resistant parent strain. Mutants characterized so far are all due to recessive nuclear genes and represent at least seven complementation groups. They exhibit different degrees as well as different patterns of sensitivity towards monofunctional and bifunctional alkylating agents, and ultraviolet light.

Alkylating Agents↗
PubMed1981

Toxicity, interstrand cross-links and DNA fragmentation induced by 'activated' cyclophosphamide in yeast.

Treatment of yeast cells with 4-hydroperoxy-cyclophosphamide (4-OOH-CP), the chemically activated form of cyclophosphamide, results in cell killing, induction of DNA interstrand cross-links and DNA fragmentation. Toxicity of 4-OOH-CP is greatly influenced by the cell's capacity of DNA dark-repair: genetic blocking of non-epistatic pathways of DNA repair results in an increase of sensitivity of several orders of magnitude. DNa primary lesions have been measured using a haploid, excision deficient, dTMP-uptaking mutant of S. cerevisiae. In this strain, a significant extent of DNA cross-linking can already be observed at a survival of 88%. At a concentration of 100 nmol/ml 4-OOH-CP, renaturability of DNA increases up to 12 h of drug exposure and drops to lower values upon further incubation. In contrast to the time course of renaturability, DNA double-strand breakage is seen at later stages of drug treatment and continuously increases as a function of incubation time. Whereas inactivation of cells and induction of strand breakage continue upon postincubation of cells, comparable effects are much less pronounced for DNA renaturability.

Cyclophosphamide↗
PubMed1981

Comparative study on the effects of cyclophosphamide on yeast in vitro and in the host-mediated assay: DNA damage and biological response.

Cyclophosphamide (CP), whether applied in its chemically activated form as 4-hydroperoxy-cyclophosphamide (4-OOH-CP) in vitro or in the host-mediated assay (HMA) using rats, exhibits toxic and mutagenic effects on excision deficient yeast cells. The expression of these effects is examined during a prolonged postincubation in buffer and compared with the ability of activated CP to induce interstrand cross-links and DNA fragmentation. At comparable doses, we observed a close similarity of biological and biochemical effects in either test system.

Animals↗
PubMed1979

Formation and fate of cross-links induced by polyfunctional anticancer drugs in yeast.

A method to detect low levels of interstrand cross-links in DNA of Saccharomyces cerevisiae is described. Isopycnic ultracentrifugation of alkali-treated, unpurified Eaton press homogenates allows the detection of less than one cross-link per yeast chromosome. Efficient separation of single- and double-stranded DNA requires low cell density and addition of glycerol during homogenization. Using a yeast strain defective in excision repair, a dose dependent formation of interstrand cross-links after treatment of cells with biological doses of nitrogen mustard, Triaziquone and Chloramubil could be demonstrated. The most powerful of these alkylating agents is Triziquone: half of the DNA molecules are shown to be cross-linked after a 12 min exposure to 9 X 10(-9) g/ml of the drug. The cross-linking reaction continues after excessive alkylating agent is removed. After having reached a maximum the fraction of renaturable DNA decreases upon further incubation. The speed of this "after-reaction" depends on temperature: 48 h after the end of treatment renaturability of DNA has almost completely disappeared when cells are kept at 36 degrees C.

Alkylation↗
PubMed1979

Biological and chemical effects of mustard gas in yeast.

Mustard gas induces inactivation and mutation in yeast. Both effects are dose-proportional, indicating single-hit events. Induction of both effects is influenced by the cell's capacity for DNA dark-repair, whereby the probability of reversion is highest in repair-proficient cells. Binding of mustard gas to cells and probably to DNA is independent of DNA-repair systems. The number of inter-strand cross-links, as determined by assaying for renaturability of alkalidenatured DNA, increases in a dose-proportional manner. At 37% survival an excision-deficient strain contains 55 inter-strand cross-links. Chromatographic analysis yields several alkylation products of DNA. Their relative frequencies resemble the values reported for E. coli and bacteriophage T7.

Centrifugation, Density Gradient↗
PubMed1979

Mutagenesis by cytostatic alkylating agents in yeast strains of differing repair capacities.

Reversion of two nulcear ochre nonsense alleles and cell inactivation induced by mono-, bi-, and tri-functional alkylating agents and by UV has been investigated in stationary-phase haploid cells of yeast strains with differing capacities for DNA repair. The ability to survive alkylation damage is correlated with UV repair capacity, a UV-resistant and UV-mutable strain (RAD REV) being least and a UV-sensitive and UV-nonmutable strain (radi rev3) most sensitive. Mutagenicity of alkylating agents is highest in the former and is abolished in the latter strain. Deficiency in excision repair (rad1 rad2) or in the RAD18 function does not lead to enhanced mutability. Mutagenesis by the various agents is characterized by a common pattern of induction of locus-specific revertants and suppressor mutants. Induction kinetics are mostly linear, but UV-induced reversion in the RAD REV strain follows higher-than-linear (probably "quadratic") kinetics. The alkylating agent cyclophosphamide, usually considered inactive without metabolic conversion, reduces colony-forming ability and induces revertants in a manner similar but not identical to the other chemicals tested. These findings are taken to support the concept of mutagenesis by misrepair after alkylation, which albeit sharing common features with the mechanism of UV-induced reversion, can be distinguished therefrom.

Alkylating Agents↗
PubMed1978

Genetic activity of chemicals in yeast: DNA alterations and mutations induced by alkylating anti-cancer agents.

The simple eukaryotic organism baker's yeast allows demonstration of primary DNA lesions in parallel with measurement of mutagenicity and lethality after treatment with alkylating chemicals. Several anti-cancer drugs formed cross-linked DNA molecules and were genetically active. The mutagenicity and lethality of these drugs varied substantially and were dependent on the function of some processes of DNA dark-repair.

Alkylating Agents↗
PubMed1976

A simple method for the isolation and characterization of thymidylate uptaking mutants in Saccharomyces cerevisiae.

The mutant tmpl--10ts which confers thermosensitive auxotrophy for thymidylate is employed for the selection of 5'-dTMP uptaking mutants. At the nonpermissive temperature yeast cells phenotypically wild type for thymidylate uptake can grow for only 3 to 4 generations in the presence of 10(-2) M 5'-dTMP. Thymidylate utilizing mutants (tum mutants) were isolated which can grow in the presence of 12 to 24 mug 5'-dTMP/ml. Genetical analysis revealed one of these mutant strains to be a double mutant, tuml tum2. For normal growth haploid thymidylate auxotrophic strains require approximately 360 mug 5'-dTMP/ml when tuml and 24 mug 5'-dTMP when tum2 is present, respectively. Cells prototrophic for thymidylate (TMP) harbouring tuml tum2 will also take up 5'-dTMP and incorporate it specifically into their DNA. Thymidylate utilization in such strains is independent of functional mitochondria, as similar incorporation of labelled 5'-dTMP is found in isogenic strains with rho+, rho- and rho0 status. Optimal stimulation of the 5'-dTMP uptaking principle in haploid TMP strains is found at 4 mug5'-dTMP/ml when tuml and tum2 are present.

Haploidy↗
PubMed1975

Nucleic acid metabolism in yeast. I. Inhibition of RNA and DNA synthesis by high concentrations of exogenous deoxythymidine 5'-monophosphate in 5'-dTMP low requiring strains.

The three haploid yeast strains T2tmpl1-3, T2tmp1-1, and T6tmp1-51 auxotrophic for 5'-dTMP differ in their requirement for thymidylate: 72, 16, and 3 mug 5'-dTMP/ml will restore optimal growth, respectively. Thymidylate low requirement in strain T2tmp1-1 and T6tmp1-51 is termed tlrA and tlrC, respectively. When the growth medium is made 5 x 10(-4) M for 5'-dTMP only strain T6tmp1-51 is severely inhibited in RNA and DNA synthesis. This inhibition is reversible after removal of excessive 5'-dTMP. The inhibitory characteristic is in marked contrast to "thymineless death" due to the lack of 5'-dTMP in strain T6tmp1-51 where only DNA synthesis stops while RNA synthesis continues. The inhibitory effect of 5 x 10(-4) M 5'-dTMP is not due to the 5'-dTMP auxotrophy but to the thymidylate low requiring character (tlrC) in strain T6tmp1-51. The arrest of RNA and DNA synthesis by high concentrations of exogenous 5'-dTMP suggests a regulatory role of either the mono- or triphosphate on nucleoside or nucleotide biosynthesis in yeast.

DNA↗