Genotoxic effects of structurally related beta-carboline alkaloids.

beta-Carboline alkaloids, found in medicinal plants, tobacco smoke and well-cooked foods, have shown a variety of actions in biological systems related to their interaction with DNA. Therefore, these alkaloids can be considered potentially mutagenic. In this work, the genotoxic, mutagenic, and cytotoxic activities of three aromatic beta-carboline alkaloids (harman, harmine, and harmol) and two dihydro-beta-carboline alkaloids (harmaline and harmalol) were evaluated by means of the Salmonella/microsome assay (Salmonella typhimurium TA98, TA97, TA100, and TA102) and SOS chromotest (Escherichia coli PQ37) with and without metabolic activation. Moreover, harman and harmine were analyzed by the micronucleus assay in vivo. It was shown that genotoxicity was inhibited by the addition of S9 mix for aromatic beta-carbolines harman and harmol in TA97. However, harmine showed signs of mutagenicity only in the presence of S9 mix in TA98 and TA97 frameshift strains. In the SOS chromotest, only harman induced SOS functions in the absence of S9 mix. Dihydro-beta-carbolines were not genotoxic in any of the microorganisms used. The negative responses obtained in the micronucleus assay indicated that harman and harmine were not able to induce chromosomal mutations.

The PSO4 gene of S. cerevisiae is important for sporulation and the meiotic DNA repair of photoactivated psoralen lesions.

We have evaluated the effect of the Saccharomyces cerevisiae pso4-1 mutation in sporulation and DNA repair during meiosis. We have found that pso4-1 cells were arrested in an early step of meiosis, before premeiotic DNA synthesis, and hence did not produce spores. These results suggest that the PSO4 gene may act at the start point of the cell cycle, as do some SPO and CDC genes. The pso4-1 mutant cells are specifically sensitive to 8-MOP- and 3-CPs-photoinduced lesions, and are found to be severely affected in meiotic recombination as well as impaired in the mutagenic response, as previously described for mitosis. This means that the PSO4 gene is important for the repair 8-MOP-photoinduced lesions, mainly double-strand breaks, and the processing of these lesions into recombinogenic intermediates.

The SOS function-inducing activity of the new nitrothiophenic derivatives in Escherichia coli.

Genotoxicity and cytotoxic effects of eight new nitrothiophenic compounds with trypanocidal activity were determined by means of the SOS Chromotest assay. Our results indicate that all nitro compounds with one aromatic ring in the R group were genotoxic without and with metabolic activation. The compounds with two aromatic rings in the R group, except for indazol-1-yl, were strongly cytotoxic but they were unable to induce SOS functions without metabolic activation. However, after metabolic activation no cytotoxic effect was observed for these compounds. The role of the nitroreductases to explain the different genotoxic responses of these nitrothiophenic derivatives is discussed.

Genetic effects of photoactivated psoralens during meiosis in DNA repair mutant pso3-1 of Saccharomyces cerevisiae.

The influence of the DNA repair gene PSO3 on photoactivated psoralen-induced meiotic recombination, gene conversion, reverse mutation, and on survival, was assayed in diploid strains of Saccharomyces cerevisiae homozygous for the wild-type or the pso3-1 mutant allele. Sporulation was normal in the pso3-1 diploid. Wild-type and mutant strains had the same sensitivity to photoactivated monofunctional psoralen (3-CPs + UVA) in meiosis-uncommitted and meiosis-committed stages. The mutant showed higher sensitivity to photoactivated bifunctional psoralen (8-MOP + UVA) during all stages of the meiotic cycle. Mutation induction by 3-CPs + UVA or 8-MOP + UVA in meiosis-committed cells revealed no significant differences between wild-type and the pso3-1 mutant. The status of the PSO3 gene has no influence on the kinetics of induction of gene conversion and crossing-over after 3-CPs + UVA treatment in meiosis-committed cells: gene conversion was blocked while recombination was induced. After treatment with 8-MOP + UVA gene conversion was also blocked in both strains while crossing-over could only be observed in meiosis-committed wild-type cells.

Genotoxic, mutagenic and recombinogenic effects of rauwolfia alkaloids.

In the last decade, the possible correlation between the use of reserpine and rauwolfia drugs as antihypertensive agents and breast cancer incidence has been investigated. For the purpose of evaluating the mutagenic and genotoxic effects of these drugs, reserpine and ajmalicine were studied using the SOS Chromotest and the induction of gene conversion, crossing-over and reverse mutation in the yeast diploid strain XS2316. The results indicated a lack of genotoxic, mutagenic and recombinogenic effects.

PSO4: a novel gene involved in error-prone repair in Saccharomyces cerevisiae.

The haploid xs9 mutant, originally selected for on the basis of a slight sensitivity to the lethal effect of X-rays, was found to be extremely sensitive to inactivation by 8-methoxypsoralen (8MOP) photoaddition, especially when cells are treated in the G2 phase of the cell cycle. As the xs9 mutation showed no allelism with any of the 3 known pso mutations, it was now given the name of pso4-1. Regarding inactivation, the pso4-1 mutant is also sensitive to mono- (HN1) or bi-functional (HN2) nitrogen mustards, it is slightly sensitive to 254 nm UV radiation (UV), and shows nearly normal sensitivity to 3-carbethoxypsoralen (3-CPs) photoaddition or methyl methanesulfonate (MMS). Regarding mutagenesis, the pso4-1 mutation completely blocks reverse and forward mutations induced by either 8MOP or 3CPs photoaddition, or by gamma-rays. In the cases of UV, HN1, HN2 or MMS treatments, while reversion induction is still completely abolished, forward mutagenesis is only partially inhibited for UV, HN1, or MMS, and it is unaffected for HN2. Besides severely inhibiting induced mutagenesis, the pso4-1 mutation was found to be semi-dominant, to block sporulation, to abolish the diploid resistance effect, and to block induced mitotic recombination, which indicates that the PSO4 gene is involved in a recombinational pathway of error-prone repair, comparable to the E. coli SOS repair pathway.

Induction of the cytoplasmic 'petite' mutation in pso mutants of Saccharomyces cerevisiae by photoaddition of furocoumarins or by ultraviolet radiation.

The induction of the cytoplasmic 'petite' mutation (or rho-) after photoaddition of either 8-methoxypsoralen (8-MOP) or 3-carbethoxypsoralen (3-CPs), after 254 nm u.v. and after 6-N-hydroxyaminopurine treatment was examined in three pso mutants in comparison to wild-type Saccharomyces cerevisiae. In three pso mutants which are defective in the induction of nuclear reverse and forward mutations, the photoaddition of 8-MOP enhanced the induction of rho-. This was true for cells in both exponential and stationary phases of growth. After photoaddition of 3-CPs in both growth phases the frequency of rho- was enhanced in pso3-1 whereas pso1-1 showed the same response as the wild-type. In pso2-1 the frequency of rho- was reduced. After treatment with 254 nm u.v. in the stationary phase of growth, rho- induction was increased in pso1-1 and pso3-1 cells as compared to wild-type cells. However, when treated in the exponential phase of growth all three pso mutants showed reduced rho- frequency. The data indicate that the defect in the repair of furocoumarins plus light-induced lesions controlled by nuclear genes (pso) interferes to various extents with the fate of mitochondrial lesions. The frequency of rho- mutants induced in the pso mutants by an analogue of adenine, 6-N-hydroxyaminopurine, was similar to that observed in the wild-type strain, suggesting that this drug may also act at the mitochondrial level as a direct mutagen in yeast.