Sayaka Nakai

Genetic control of diploid recovery after γ-irradiation in the yeast Saccharomyces cerevisiae

Abstract Genetic mechanism(s) of γ-ray resistance of the diploid and budding haploid cells of S. cerevisiae were investigated, with special reference to mitotic recombination, by examining 11 rad mutant strains. The radiosensitivity of the diploid was markedly enhanced in certain γ-ray-sensitive rad mutants, whereas the sensitivity of the haploid was not so enhanced in these rad mutants. These enhanced sensitivities of diploids were irrespective of their own haploid sensitivities. From these results, the existence of a mechanism of diploid-specific recovery was postulated. The magnitude of diploid radioresistance in rad mutants was positively correlated with the ability for the induction of mitotic recombinational events which were controlled by RAD genes belonging to the RAD-51 genetic pathway. The genetic mechanism(s) of the diploid recovery after γ-irradiation are probably related to recombinational processes between the homologous chromosomes leading to reciprocal recombination or non-reciprocal gene conversion. Furthermore, the higher radioresistance of budding cells in comparison with the non-budding cells was also correlated to the diploid radioresistance with a few exceptions. Consequently, the mechanism(s) of budding radioresistance similar to teh diploid recovery seems to be related to mitotic recombinational processes.

Two types of radiation-sensitive mutant in yeast

Abstract Three radiation-sensitive mutants designated UV 1 S , UV 2 S and X 1 S were isolated from the yeast Saccharomyces cerevisiae . The three genes responsible for radiation sensitivity are located at different loci. In haploid states, the single mutants, X 1 S , UV 1 S and UV 2 S , the double mutants, UV 1 S UV 2 S , X 1 S UV 1 S and X 1 S UV 2 S , and the triple mutant, X 1 S UV 1 S UV 2 S , have, respectively, 1.5, 27, 27, 27, 80, 60 and 80 times higher sensitivities to killing by ultraviolet (2537 A) irradiation (UV) than the wild-type strain. In diploid states, homozygous mutants, X 1 S /X 1 S , UV 1 S /UV 1 S and UV 2 S / UV 2 S , and heterozygous mutants, X 1 S /+, UV 1 S /+, UV 2 S /+, have, respectively, 2, 28, 28, 1.5, 1 and 1.5 times higher sensitivities to UV-killing than the wild-type. With regard to X-ray sensitivity, mutant genes, UV 1 S and UV 2 S , have no effect but the X 1 S gene has an appreciable and interesting effect. The sensitization effect of the X 1 S gene is unique: it eliminates the “tailing” effect typical in wild-type haploid survival curves, and the “shoulder” in the wild-type diploid survival curves. Experimental results indicate that a similarity exists between the UV S gene in yeast and the UV-sensitive genes giving rise to reduced dark repair ability in E. coli and between the X S gene in yeast and Rec− gene in E. coli .

Induction of spontaneous and UV-induced mutations during commitment to meiosis in Saccharomyces cerevisiae.

Inductions of reversions of nonsense, missense and frameshift-type mutations were investigated in a diploid cell population of Saccharomyces cerevisiae during commitment to meiosis, by using the medium-transfer technique from sporulation medium to vegetative medium. The yields of spontaneous reverse mutations obtained from the cells that were committed to different stages during meiosis were rather constant irrespective of the alleles tested, although the yields of both intergenic and intragenic recombinations markedly increased. The susceptibilities to UV-induced reverse mutations examined during commitment to meiosis were not changed appreciably. It is concluded that induction of base-change-type mutations in meiosis is not essentially different from that in mitosis.

Radiation-induced chromosome aberrations in lymphocytes from man and crab-eating monkey: The dose-response relationships at low doses

To obtain information on the relation between yield of chromosome aberrations and dose at low-dose levels, experiments were conducted with 5, 10, 20, 30 and 50 rad of 137Cs gamma-rays, on lymphocytes from man and crab-eating monkey (Macaca fascicularis). The dose-response relationship for dicentrics was obtained from the combined data of these low-dose experiments with those of our previous ones at high doses (100-400 rad). When the difference between observed yields and those expected from the linear-quadratic model were computed, the dose-response curve had a good fit for man, but not for the monkey. The linear regression lines between 0 and 30 rad were calculated, because the expected values of alpha/beta for man and monkey would be about 100 and 60 rad. The human data gave a satisfactory fit to a linear model, i.e., a linear increase in aberration frequency with dose, whereas this was not so for those of the monkey. Furthermore, there was some suggestive evidence for the existence of a plateau in dicentric yields between 10 and 30 rad for the monkey and between 20 and 30 rad for human lymphocytes, but more data would be needed to verify this suggestion, particularly for human lymphocytes.

Evaluation of radiation-induced chromosomal aberrations in human peripheral blood lymphocytes in vitro: result of an IAEA-coordinated programme.

The results of an IAEA coordinated programme on radiation induced chromosomal aberrations in human peripheral blood lymphocytes in vitro are presented. In a master experiment, a whole blood sample from one donor was irradiated with 200 R of X-rays. Different fixation times from 46 to 82 h were used. The progression of cells into mitosis was monitored by BrdUrd incorporation. 14 investigators took part in the scoring of chromosomal aberrations. The main conclusions of this study are: (1) The mean frequencies of aberrations changed with fixation time. (2) The number of cells scored as aberrant by different laboratories was very similar, but there was variability in the number of aberrations scored per aberrant cell. (3) The differences in the frequencies of aberrations between laboratories were minimal when the scoring was restricted to the first major peak of mitotic activity and sufficient cells were scored. It is concluded that using controlled experimentals conditions, human peripheral blood lymphocytes can effectively be used as a reliable biological dosimeter for absorbed radiation dose.

γ-Ray-induced reciprocal translocations in spermatogonia of the crab-eating monkey (Macaca fascicularis)

Abstract The yield of translocations induced by γ-rays in the crab-eating monkey (Macaca fascicularis) spermatogonia were studied by cytological analysis in spermatocytes derived from them. The frequencies of translocations were 0.09% at 0 Gy, 1.9% at 1 Gy, 2.5% at 2 Gy and 1.3% at 3 Gy, showing a humped dose-response curve with a peak yield around 2 Gy. No remarkable inter-seasonal or inter-animal variations in the induction of translocation were observed. The frequencies in the crab-eating monkey were significantly higher than those in the same Macaca genus, the rhesus monkey (Macaca mulatta) (van Buul, 1976, 1980). This inter-species difference in radiosensitivity might be affected by the condition of spermatogonial stem cells at the time of exposure to radiation, depending on the seasonal change in spermatogenetic activity.

Dose-response relations for dicentric yields in G0 lymphocytes on man and crab-eating monkey following acute and chronic γ-irradiations

A comparison has been made of dicentric yields in GO lymphocytes between man and crab-eating monkey, Macaca fascicularis, after acute and chronic gamma-irradiations. With acute irradiation (49.6 rad/min) there was no significant difference between them, but for the chronic irradiation (17.1 rad/h) a significant difference was observed between the species. When the dose-response relations were fitted to the linear-quadratic model (Y = alpha D + beta D2), the species-difference observed for chronic irradiation was almost entirely due to change in the value of beta. In addition, after chronic irradiation the beta-value for monkey was almost negligible, but that for man was significant. Post-irradiation incubation experiment showed that cells with dicentrics were partly eliminated during the course of chronic irradiation, because there were appreciable reductions of dicentric yields (ca. 25% for both man and monkey at 400 rad) together with mitotic indices (ca. 30% and 60% for man and monkey, respectively, at 400 rad). Accordingly, it would be reasonable to postulate that GO repair for dicentrics other than selection mechanism must play a major role in the effects of low dose rate. It can be further suggested that GO-repair capacity for chromosal damages leading to dicentrics may be different among different primate species.

Dose-response relationship of γ-ray-induced reciprocal translocations at low doses in spermatogonia of the crab-eating monkey (Macaca fascicularis)

Abstract The yield of translocations induced by acute γ-irradiation at low doses (0.25 and 0.50 Gy) in the crab-eating monkeys (Macaca fascicularis) spermatogonia was examined. The frequencies of translocations per cell were 0.53% at 0.25 Gy and 1.07% at 0.50 Gy. Over the low dose range from 0 to 1 Gy, the dose-response relationship for translocation yield was a linear one with a regression coefficient of 1.79 · 10−2. To estimate the sensitivity to the induction of translocations in the crab-eating monkeys spermatogonia, the slope of the regression line was compared with those in other mammalian species. Consequently, over the low dose range below 1 Gy, the sensitivity of the crab-eating monkeys spermatogonia to translocation induction was similar to several mammalian species, the mouse. Chinese hamster, and the rabbit, but significantly higher than that of the rhesus monkey and lower than that of the marmoset.

Unusual dose-response of chromosome aberrations induced in human lymphocytes by very low dose exposures to tritium

Leukocyte cultures of human peripheral blood were chronically exposed for 48 h to tritiated water and [3H]thymidine using a wide range of tritium doses, and aberrations in lymphocyte chromosomes at the first metaphases were examined. In the experimental conditions, the types of aberrations induced by radiation emitted from both tritiated water and [3H]thymidine were mostly chromatid types, such as chromatid gaps and deletions. The dose--response relations for chromatid breaks per cell exhibited unusual dose-dependency in both cases. It was demonstrated that at higher dose range the yields of chromatid breaks increased linearly with dose, while those at lower dose range were significantly higher than would be expected by a downward extrapolation from the linear relation. Partial-hit or partial-target kinetic events appears at very low dose exposure.

Differential effect of UV irradiation on induction of intragenic and intergenic recombination during commitment to meiosis in Saccharomyces cerevisiae

A comparison was made between the induction of intragenic and intergenic recombinations during meiosis in a wild-type diploid of Saccharomyces cerevisiae. Under non-irradiated normal conditions, production of both intragenic and intergenic recombinants greatly increased in the cells with commitment to meiosis. The susceptibility of cells to the induction of both the spontaneous intra- and intergenic recombinations in meiotic cells was similar. However, under condition of UV irradiation, there were striking differences between intra- and intergenic recombinations. Susceptibility to induction of intragenic recombination by UV irradiation was not enhanced at meiosis compared with mitosis, and was not altered through commitment to meiotic processes. In contrast, however, susceptibility to the induction of intergenic recombination by UV irradiation was enhanced markedly during commitment to meiosis compared with mitosis. Genetic analysis suggested that the enhanced susceptibility to recombination during meiosis is specifically concerned with reciprocal-type recombination (crossing-over) but not non-reciprocal-type recombination (gene conversion). Hence it is concluded that the meiotic process appears to be intimately concerned with the mechanism(s) of induction of recombination, especially reciprocal-type recombination.