Different internal metabolites trigger the induction of glycolytic gene expression in Saccharomyces cerevisiae.

In the yeast Saccharomyces cerevisiae, the sugar-induced expression of various genes coding for glycolytic enzymes is triggered by increases in the concentrations of different internal metabolites. Here, we show that the induction of the glycolytic isoenzyme enolase 2 is strictly dependent on the abilities of different mutant strains to increase the level of glucose-6-phosphate after the addition of sugars. In contrast, the induction of alcohol dehydrogenase I is dependent on increasing concentrations of metabolites in the late stages of glycolysis.     

Differential regulation of glutaredoxin gene expression in response to stress conditions in the yeast Saccharomyces cerevisiae

Glutaredoxins are small heat-stable proteins that are active as glutathione-dependent oxidoreductases and are encoded by two genes, designated GRX1 and GRX2, in the yeast Saccharomyces cerevisiae. We report here that the expression of both genes is induced in response to various stress conditions including oxidative, osmotic, and heat stress and in response to stationary phase growth and growth on non-fermentable carbon sources. Furthermore, both genes are activated by the high-osmolarity glycerol pathway and negatively regulated by the Ras-protein kinase A pathway via stress-responsive STRE elements. GRX1 contains a single STRE element and is induced to significantly higher levels compared to GRX2 following heat and osmotic shock. GRX2 contains two STRE elements, and is rapidly induced in response to reactive oxygen species and upon entry into stationary phase growth. Thus, these data support the idea that the two glutaredoxin isoforms in yeast play distinct roles during normal cellular growth and in response to stress conditions.     

PGA基因在酿酒酵母中的表达研究

本文根据GenBank 中巨大芽孢杆菌（Bacillus megaterium）的PGA基因序列设计了上下游引物,通过PCR扩增出巨大芽孢杆菌1.1741中的PGA基因。将该基因连接到T7lac启动子控制下的表达载体pYES2(amp+,ura+)上,构建了重组质粒pYES2-PGA。用LiAc/SSDNA/PEG方法将其转化进酿酒酵母(Saccharomyces cerevisiae)H158中表达,在不需要苯乙酸诱导的重组菌株发酵液中检测到了青霉素酰化酶活性,最高酶活达到0.75 U/ml。将该PGA基因测序结果与GenBank中巨大芽孢杆菌L04471.1、U07682.1和Z37542三株的PGA基因序列比对,表现出很高的同源性,分别达到97.1%、99.8% 和99.8%。     

Low affinity orthophosphate carriers regulate PHO gene expression independently of internal orthophosphate concentration in Saccharomyces cerevisiae

Phosphate is an essential nutrient that must be taken up from the growth medium through specific transporters. In Saccharomyces cerevisiae, both high and low affinity orthophosphate carriers allow this micro-organism to cope with environmental variations. Intriguingly, in this study we found a tight correlation between selenite resistance and expression of the high affinity orthophosphate carrier Pho84p. Our work further revealed that mutations in the low affinity orthophosphate carrier genes (PHO87, PHO90, and PHO91) cause deregulation of phosphate-repressed genes. Strikingly, the deregulation due to pho87Delta, pho90Delta, or pho91Delta mutations was neither correlated to impaired orthophosphate uptake capacity nor to a decrease of the intracellular orthophosphate or polyphosphate pools, as shown by (31)P NMR spectroscopy. Thus, our data clearly establish that the low affinity orthophosphate carriers affect phosphate regulation independently of intracellular orthophosphate concentration through a new signaling pathway that was found to strictly require the cyclin-dependent kinase inhibitor Pho81p. We propose that phosphate-regulated gene expression is under the control of two different regulatory signals as follows: the sensing of internal orthophosphate by a yet unidentified protein and the sensing of external orthophosphate by low affinity orthophosphate transporters; the former would be required to maintain phosphate homeostasis, and the latter would keep the cell informed on the medium phosphate richness.     

Analysis of mutations affecting Ty-mediated gene expression in Saccharomyces cerevisiae

Summary Yeast translocatable, Ty, elements can cause constitutive synthesis of the glucose-repressible alcohol dehydrogenase (ADHII) when inserted upstream from the 5 end of the structural gene, ADR2. These insertion mutations, ADR3 ^c, are unstable and give rise to secondary ADHII^– mutations. The majority of such mutants, adr3, can be attributed to excision of the insertion sequence, leaving behind a single copy of the -sequence which occurs as a direct repeat at the ends of the Ty elements. A few adr3 mutants appear to be generated by DNA-rearrangements in the vicinity of the Ty insertion. The occurrence of recessive mutants, tye, which are unlinked to ADR2 indicates that the constitutive expression of ADR2 caused by the Ty insertions requires the function of trans-acting genes. These results support the idea that regulation of Ty-linked ADR2 is actively mediated by the insertion sequence and is probably not due to a mere disruption of the wild-type controlling site.     

Comparative analysis of fermentation and enzyme expression profiles among industrial Saccharomyces cerevisiae strains

Applied Microbiology and Biotechnology - Industrial diploid strains of Saccharomyces cerevisiae are selected from natural populations and then domesticated by optimizing the preferred properties...     

Cloning of Candida pelliculosa beta-glucosidase gene and its expression in Saccharomyces cerevisiae

Summary Candida pelliculosa var. acetaetherius is a strain of yeast which can utilize cellobiose as the carbon source. From a gene library prepared from this yeast, the -glucosidase gene has been cloned in a S. cerevisiae host using a chromogenic substrate, 5-bromo-4-chloro-3-indolyl--glucoside as an indicator. It was proved by Southern analysis that the DNA fragment carrying the -glucosidase gene originated from C. pelliculosa. -Glucosidase produced by S. cerevisiae transformants was secreted into the periplasmic space. In Candida, -glucosidase was not induced by cellobiose but was derepressed by lowering the concentration of glucose. The regulation of -glucosidase synthesis in S. cerevisiae carrying the cloned -glucosidase was not clear compared with that in Candida, however, the enzyme activity in low glucose medium (0.05%) was reproducibly higher than in high glucose medium (2%). We have found the sequence that controls the expression of the -glucosidase gene negatively in S. cerevisiae.     

Differential function and expression of Saccharomyces cerevisiae B-type cyclins in mitosis and meiosis.

We have studied the patterns of expression of four B-type cyclins (Clbs), Clb1, Clb2, Clb3, and Clb4, and their ability to activate p34cdc28 during the mitotic and meiotic cell cycles of Saccharomyces cerevisiae. During the mitotic cell cycle, Clb3 and Clb4 were expressed and induced a kinase activity in association with p34cdc28 from early S phase up to mitosis. On the other hand, Clb1 and Clb2 were expressed and activated p34cdc28 later in the mitotic cell cycle, starting in late S phase and continuing up to mitosis. The pattern of expression of Clb3 and Clb4 suggests a possible role in the regulation of DNA replication as well as mitosis. Clb1 and Clb2, whose pattern of expression is similar to that of other known Clbs, are likely to have a role predominantly in the regulation of M phase. During the meiotic cell cycle, Clb1, Clb3, and Clb4 were expressed and induced a p34cdc28-associated kinase activity just before the first meiotic division. The fact that Clb3 and Clb4 were not synthesized earlier, in S phase, suggests that these cyclins, which probably have a role in S phase during the mitotic cell cycle, are not implicated in premeiotic S phase. Clb2, the primary mitotic cyclin in S. cerevisiae, was not detectable during meiosis. Sporulation experiments on strains deleted for one, two, or three Clbs indicate, in agreement with the biochemical data, that Clb1 is the primary cyclin for the regulation of meiosis, while Clb2 is not involved at all.