Suppressors Reveal Two Classes of Glucose Repression Genes in the Yeast Saccharomyces Cerevisiae

We selected and analyzed extragenic suppressors of mutations in four genes-GRR1, REG1, GAL82 and GAL83-required for glucose repression of the GAL genes in the yeast Saccharomyces cerevisiae. The suppressors restore normal or nearly normal glucose repression of GAL1 expression in these glucose repression mutants. Tests of the ability of each suppressor to cross-suppress mutations in the other glucose repression genes revealed two groups of mutually cross-suppressed genes: (1) REG1, GAL82 and GAL83 and (2) GRR1. Mutations of a single gene, SRG1, were found as suppressors of reg1, GAL83-2000 and GAL82-1, suggesting that these three gene products act at a similar point in the glucose repression pathway. Mutations in SRG1 do not cross-suppress grr1 or hxk2 mutations. Conversely, suppressors of grr1 (rgt1) do not cross-suppress any other glucose repression mutation tested. These results, together with what was previously known about these genes, lead us to propose a model for glucose repression in which Grr1p acts early in the glucose repression pathway, perhaps affecting the generation of the signal for glucose repression. We suggest that Reg1p, Gal82p and Gal83p act after the step(s) executed by Grr1p, possibly transmitting the signal for repression to the Snf1p protein kinase.     

端粒植入胃癌7901细胞对细胞生长、端粒和端粒酶的影响

研究外源端粒片段植入胃癌7901细胞后对细胞生长、端粒长度和端粒酶活性的影响.采用lipofectTM2000介导的转染方式,将含有端粒片段质粒pSXneo-1.6-T2AG3转染胃癌细胞SGC7901,PCR在基因水平上鉴定外源性端粒片段的植入后,采用TRAP法检测转染细胞端粒酶活性变化,TRF法检测转染细胞端粒长度变化,MTT法检测细胞生长曲线,RT-PCR测定转染细胞hTERT表达变化.染色体核型分析细胞染色体变化.结果显示端粒片段成功导入SGC7901细胞后获得稳定的细胞株,端粒片段植入后细胞生长变慢,端粒长度延长不明显,端粒酶活性明显降低,hTERT mRNA表达水平下降,核型分析显示转染前后细胞染色体数目无明显变化.实验成功将携带了1600 bp端粒TTAGGG重复序列的真核表达载体pSX-T2AG3-neo稳定转染至人胃癌7901细胞中,端粒植入降低细胞端粒酶的活性和下调端粒酶活性亚单位hTERT的表达,但对端粒长度无明显影响.     

Genetic Interactions at the Fla10 Locus: Suppressors and Synthetic Phenotypes That Affect the Cell Cycle and Flagellar Function in Chlamydomonas Reinhardtii

Through the isolation of suppressors of temperature-sensitive flagellar assembly mutations at the FLA10 locus of Chlamydomonas reinhardtii, we have identified six other genes involved in flagellar assembly. Mutations at these suppressor loci, termed SUF1-SUF6, display allele specificity with respect to which fla10- mutant alleles they suppress. An additional mutation, apm1-122, which confers resistance to the plant herbicides amiprophos-methyl and oryzalin, was also found to interact with mutations at the FLA10 locus. The apm1-122 mutation in combination with three fla10- mutant alleles results in synthetic cold-sensitive cell division defects, and in combination with an additional pseudo-wild-type fla10- allele yields a synthetic temperature-sensitive flagellar motility phenotype. Based upon the genetic interactions of these loci, we propose that the FLA10 gene product interacts with multiple components of the flagellar apparatus and plays a role both in flagellar assembly and in the cell cycle.     

Isolation and Characterization of Omnipotent Suppressors in the Yeast Saccharomyces Cerevisiae

Approximately 290 omnipotent suppressors, which enhance translational misreading, were isolated in strains of the yeast Saccharomyces cerevisiae containing the psi+ extrachromosomal determinant. The suppressors could be assigned to 8 classes by their pattern of suppression of five nutritional markers. The suppressors were further distinguished by differences in growth on paromomycin medium, hypertonic medium, low temperatures (10 degrees), nonfermentable carbon sources, alpha-aminoadipic acid medium, and by their dominance and recessiveness. Genetic analysis of 12 representative suppressors resulted in the assignment of these suppressors to 6 different loci, including the three previously described loci SUP35 (chromosome IV), SUP45 (chromosome II) and SUP46 (chromosome II), as well as three new loci SUP42 (chromosome IV), SUP43 (chromosome XV) and SUP44 (chromosome VII). Suppressors belonging to the same locus had a wide range of different phenotypes. Differences between alleles of the same locus and similarities between alleles of different loci suggest that the omnipotent suppressors encode proteins that effect different functions and that altered forms of each of the proteins can effect the same function.     

Effect of Yeast Cell Morphology,Cell Wall Physical Structure and Chemical Composition on Patulin Adsorption

The capability of yeast to adsorb patulin in fruit juice can aid in substantially reducing the patulin toxic effect on human health. This study aimed to investigate the capability of yeast cell morphology and cell wall internal structure and composition to adsorb patulin. To compare different yeast cell morphologies, cell wall internal structure and composition, scanning electron microscope, transmission electron microscope and ion chromatography were used. The results indicated that patulin adsorption capability of yeast was influenced by cell surface areas, volume, and cell wall thickness, as well as 1,3-β-glucan content. Among these factors, cell wall thickness and 1,3-β-glucan content serve significant functions. The investigation revealed that patulin adsorption capability was mainly affected by the three-dimensional network structure of the cell wall composed of 1,3-β-glucan. Finally, patulin adsorption in commercial kiwi fruit juice was investigated, and the results indicated that yeast cells could adsorb patulin from commercial kiwi fruit juice efficiently. This study can potentially simulate in vitro cell walls to enhance patulin adsorption capability and successfully apply to fruit juice industry.     

Effect of Ethanol on Cell Growth of Budding Yeast: Genes That Are Important for Cell Growth in the Presence of Ethanol

The budding yeast Saccharomyces cerevisiae has been used in the fermentation of various kinds of alcoholic beverages. But the effect of ethanol on the cell growth of this yeast is poorly understood. This study shows that the addition of ethanol causes a cell-cycle delay associated with a transient dispersion of F-actin cytoskeleton, resulting in an increase in cell size. We found that the tyrosine kinase Swe1, the negative regulator of Cdc28-Clb kinase, is related to the regulation of cell growth in the presence of ethanol. Indeed, the increase in cell size due to ethanol was partially abolished in the SWE1-deleted cells, and the amount of Swe1 protein increased transiently in the presence of ethanol. These results indicated that Swe1 is involved in cell size control in the presence of ethanol, and that a signal produced by ethanol causes a transient up-regulation of Swe1. Further we investigated comprehensively the ethanol-sensitive strains in the complete set of 4847 non-essential gene deletions and identified at least 256 genes that are important for cell growth in the presence of ethanol.     

Effects of Glucose Repression on the Transmission and Recombination of Mitochondrial Genes in Yeast (SACCHAROMYCES CEREVISIAE)

Matings of a number of Saccharomyces cerevisiae stocks give different output ratios of mitochondrial genotypes depending on whether the cells are glucose-repressed or derepressed. The effects of glucose repression are independent of cellular mating type and mitochondrial genotype, and take place at least in part after zygotes are formed. An explanation is proposed in terms of changes in the relative numbers of mitochondrial DNA molecules contributed by the a and α parents, modified by selective replication or destruction of molecules inside the zygote.     

Survival and Growth of Yeast without Telomere Capping by Cdc13 in the Absence of Sgs1, Exo1, and Rad9

Maintenance of telomere capping is absolutely essential to the survival of eukaryotic cells. Telomere capping proteins, such as Cdc13 and POT1, are essential for the viability of budding yeast and mammalian cells, respectively. Here we identify, for the first time, three genetic modifications that allow budding yeast cells to survive without telomere capping by Cdc13. We found that simultaneous inactivation of Sgs1, Exo1, and Rad9, three DNA damage response (DDR) proteins, is sufficient to allow cell division in the absence of Cdc13. Quantitative amplification of ssDNA (QAOS) was used to show that the RecQ helicase Sgs1 plays an important role in the resection of uncapped telomeres, especially in the absence of checkpoint protein Rad9. Strikingly, simultaneous deletion of SGS1 and the nuclease EXO1, further reduces resection at uncapped telomeres and together with deletion of RAD9 permits cell survival without CDC13. Pulsed-field gel electrophoresis studies show that cdc13-1 rad9Δ sgs1Δ exo1Δ strains can maintain linear chromosomes despite the absence of telomere capping by Cdc13. However, with continued passage, the telomeres of such strains eventually become short and are maintained by recombination-based mechanisms. Remarkably, cdc13Δ rad9Δ sgs1Δ exo1Δ strains, lacking any Cdc13 gene product, are viable and can grow indefinitely. Our work has uncovered a critical role for RecQ helicases in limiting the division of cells with uncapped telomeres, and this may provide one explanation for increased tumorigenesis in human diseases associated with mutations of RecQ helicases. Our results reveal the plasticity of the telomere cap and indicate that the essential role of telomere capping is to counteract specific aspects of the DDR.     

磁化溶液对酵母细胞生长代谢的影响

将供酵母生长代谢的溶液在恒磁场中静置处理,实验结果表明,磁化后的溶液使酵母繁殖加快、数量提高,且发酵初期的产酒率有一定增加。实验中采用了酵母固定化技术,保证了实验数据的连续性、可比性和重现性