scholarly journals Pho85 Kinase, a Cyclin-Dependent Kinase, Regulates Nuclear Accumulation of the Rim101 Transcription Factor in the Stress Response of Saccharomyces cerevisiae

2010 ◽  
Vol 9 (6) ◽  
pp. 943-951 ◽  
Author(s):  
Masafumi Nishizawa ◽  
Mirai Tanigawa ◽  
Michio Hayashi ◽  
Tatsuya Maeda ◽  
Yoshiaki Yazaki ◽  
...  

ABSTRACT The budding yeast Saccharomyces cerevisiae alters its gene expression profile in response to changing environmental conditions. The Pho85 kinase, one of the yeast cyclin-dependent kinases (CDK), is known to play an important role in the cellular response to alterations in parameters such as nutrient levels and salinity. Several genes whose expression is regulated, either directly or indirectly, by the Rim101 transcription factor become constitutively activated when Pho85 function is absent,. Because Rim101 is responsible for adaptation to alkaline conditions, this observation suggests an interaction between Pho85 and Rim101 in the response to alkaline stress. We have found that Pho85 affects neither RIM101 transcription, the proteolytic processing that is required for Rim101 activation, nor Rim101 stability. Rather, Pho85 regulates the nuclear accumulation of active Rim101, possibly via phosphorylation. Additionally, we report that Pho85 and the transcription factor Pho4 are necessary for adaptation to alkaline conditions and that PTK2 activation by Pho4 is involved in this process. These findings illustrate novel roles for the regulators of the PHO system when yeast cells cope with various environmental stresses potentially threatening their survival.

2015 ◽  
Vol 81 (11) ◽  
pp. 3688-3698 ◽  
Author(s):  
Kazutaka Sawada ◽  
Tomoya Sato ◽  
Hiroshi Hamajima ◽  
Lahiru Niroshan Jayakody ◽  
Miyo Hirata ◽  
...  

ABSTRACTIn nature, different microorganisms create communities through their physiochemical and metabolic interactions. Many fermenting microbes, such as yeasts, lactic acid bacteria, and acetic acid bacteria, secrete acidic substances and grow faster at acidic pH values. However, on the surface of cereals, the pH is neutral to alkaline. Therefore, in order to grow on cereals, microbes must adapt to the alkaline environment at the initial stage of colonization; such adaptations are also crucial for industrial fermentation. Here, we show that the yeastSaccharomyces cerevisiae, which is incapable of synthesizing glucosylceramide (GlcCer), adapted to alkaline conditions after exposure to GlcCer from koji cereal cultured withAspergillus kawachii. We also show that various species of GlcCer derived from different plants and fungi similarly conferred alkali tolerance to yeast. Although exogenous ceramide also enhanced the alkali tolerance of yeast, no discernible degradation of GlcCer to ceramide was observed in the yeast culture, suggesting that exogenous GlcCer itself exerted the activity. Exogenous GlcCer also increased ethanol tolerance and modified the flavor profile of the yeast cells by altering the membrane properties. These results indicate that GlcCer fromA. kawachiimodifies the physiology of the yeastS. cerevisiaeand demonstrate a new mechanism for cooperation between microbes in food fermentation.


mBio ◽  
2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Andy Hesketh ◽  
Marta Vergnano ◽  
Stephen G. Oliver

ABSTRACT Correlations between gene transcription and the abundance of high-energy purine nucleotides in Saccharomyces cerevisiae have often been noted. However, there has been no systematic investigation of this phenomenon in the absence of confounding factors such as nutrient status and growth rate, and there is little hard evidence for a causal relationship. Whether transcription is fundamentally responsive to prevailing cellular energetic conditions via sensing of intracellular purine nucleotides, independently of specific nutrition, remains an important question. The controlled nutritional environment of chemostat culture revealed a strong correlation between ATP and GTP abundance and the transcription of genes required for growth. Short pathways for the inducible and futile consumption of ATP or GTP were engineered into S. cerevisiae, permitting analysis of the transcriptional effect of an increased demand for these nucleotides. During steady-state growth using the fermentable carbon source glucose, the futile consumption of ATP led to a decrease in intracellular ATP concentration but an increase in GTP and the guanylate energy charge (GEC). Expression of transcripts encoding proteins involved in ribosome biogenesis, and those controlled by promoters subject to SWI/SNF-dependent chromatin remodelling, was correlated with these nucleotide pool changes. Similar nucleotide abundance changes were observed using a nonfermentable carbon source, but an effect on the growth-associated transcriptional programme was absent. Induction of the GTP-cycling pathway had only marginal effects on nucleotide abundance and gene transcription. The transcriptional response of respiring cells to glucose was dampened in chemostats induced for ATP cycling, but not GTP cycling, and this was primarily associated with altered adenine nucleotide levels. IMPORTANCE This paper investigates whether, independently of the supply of any specific nutrient, gene transcription responds to the energy status of the cell by monitoring ATP and GTP levels. Short pathways for the inducible and futile consumption of ATP or GTP were engineered into the yeast Saccharomyces cerevisiae, and the effect of an increased demand for these purine nucleotides on gene transcription was analyzed. The resulting changes in transcription were most consistently associated with changes in GTP and GEC levels, although the reprogramming in gene expression during glucose repression is sensitive to adenine nucleotide levels. The results show that GTP levels play a central role in determining how genes act to respond to changes in energy supply and that any comprehensive understanding of the control of eukaryotic gene expression requires the elucidation of how changes in guanine nucleotide abundance are sensed and transduced to alter the global pattern of transcription.


2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Qun Ren ◽  
Rebecca Brenner ◽  
Thomas C. Boothby ◽  
Zhaojie Zhang

Abstract Background Anhydrobiotes, such as the yeast Saccharomyces cerevisiae, are capable of surviving almost total loss of water. Desiccation tolerance requires an interplay of multiple events, including preserving the protein function and membrane integrity, preventing and mitigating oxidative stress, maintaining certain level of energy required for cellular activities in the desiccated state. Many of these crucial processes can be controlled and modulated at the level of organelle morphology and dynamics. However, little is understood about what organelle perturbations manifest in desiccation-sensitive cells as a consequence of drying or how this differs from organelle biology in desiccation-tolerant organisms undergoing anhydrobiosis. Results In this study, electron and optical microscopy was used to examine the dynamic changes of yeast cells during the desiccation process. Dramatic structural changes were observed during the desiccation process, including the diminishing of vacuoles, decrease of lipid droplets, decrease in mitochondrial cristae and increase of ER membrane, which is likely caused by ER stress and unfolded protein response. The survival rate was significantly decreased in mutants that are defective in lipid droplet biosynthesis, or cells treated with cerulenin, an inhibitor of fatty acid synthesis. Conclusion Our study suggests that the metabolism of lipid droplets and membrane may play an important role in yeast desiccation tolerance by providing cells with energy and possibly metabolic water. Additionally, the decrease in mitochondrial cristae coupled with a decrease in lipid droplets is indicative of a cellular response to reduce the production of reactive oxygen species.


2020 ◽  
Vol 86 (7) ◽  
Author(s):  
Rui Yao ◽  
Pei Zhou ◽  
Chengjin Wu ◽  
Liming Liu ◽  
Jing Wu

ABSTRACT In Saccharomyces cerevisiae, Y family DNA polymerase Rev1 is involved in the repair of DNA damage by translesion DNA synthesis (TLS). In the current study, to elucidate the role of Rev1 in oxidative stress-induced DNA damage in S. cerevisiae, REV1 was deleted and overexpressed; transcriptome analysis of these mutants along with the wild-type strain was performed to screen potential genes that could be associated with REV1 during response to DNA damage. When the yeast cells were treated with 2 mM H2O2, the deletion of REV1 resulted in a 1.5- and 2.8-fold decrease in the survival rate and mutation frequency, respectively, whereas overexpression of REV1 increased the survival rate and mutation frequency by 1.1- and 2.9-fold, respectively, compared to the survival rate and mutation frequency of the wild-type strain. Transcriptome and phenotypic analyses identified that Sml1 aggravated oxidative stress in the yeast cells by inhibiting the activity of Rev1. This inhibition was due to the physical interaction between the BRCA1 C terminus (BRCT) domain of Rev1 and amino acid residues 36 to 70 of Sml1; the cell survival rate and mutation frequency increased by 1.8- and 3.1-fold, respectively, when this interaction was blocked. We also found that Sml1 inhibited Rev1 phosphorylation under oxidative stress and that deletion of SML1 increased the phosphorylation of Rev1 by 46%, whereas overexpression of SML1 reduced phosphorylation of Rev1. Overall, these findings demonstrate that Sml1 could be a novel regulator that mediates Rev1 dephosphorylation to inhibit its activity during oxidative stress. IMPORTANCE Rev1 was critical for cell growth in S. cerevisiae, and the deletion of REV1 caused a severe growth defect in cells exposed to oxidative stress (2 mM H2O2). Furthermore, we found that Sml1 physically interacted with Rev1 and inhibited Rev1 phosphorylation, thereby inhibiting Rev1 DNA antioxidant activity. These findings indicate that Sml1 could be a novel regulator for Rev1 in response to DNA damage by oxidative stress.


2017 ◽  
Vol 37 (11) ◽  
Author(s):  
Jayasankar Mohanakrishnan Kaimal ◽  
Ganapathi Kandasamy ◽  
Fabian Gasser ◽  
Claes Andréasson

ABSTRACT Protein aggregation is intimately associated with cellular stress and is accelerated during aging, disease, and cellular dysfunction. Yeast cells rely on the ATP-consuming chaperone Hsp104 to disaggregate proteins together with Hsp70. Hsp110s are ancient and abundant chaperones that form complexes with Hsp70. Here we provide in vivo data showing that the Saccharomyces cerevisiae Hsp110s Sse1 and Sse2 are essential for Hsp104-dependent protein disaggregation. Following heat shock, complexes of Hsp110 and Hsp70 are recruited to protein aggregates and function together with Hsp104 in the disaggregation process. In the absence of Hsp110, targeting of Hsp70 and Hsp104 to the aggregates is impaired, and the residual Hsp104 that still reaches the aggregates fails to disaggregate. Thus, coordinated activities of both Hsp104 and Hsp110 are required to reactivate aggregated proteins. These findings have important implications for the understanding of how eukaryotic cells manage misfolded and amyloid proteins.


mSphere ◽  
2017 ◽  
Vol 2 (6) ◽  
Author(s):  
Walters Aji Tebung ◽  
Raha Parvizi Omran ◽  
Debra L. Fulton ◽  
Joachim Morschhäuser ◽  
Malcolm Whiteway

ABSTRACT Candida albicans poses a significant threat to the lives of immunocompromised people. Historically, knowledge has been drawn from studies on Saccharomyces cerevisiae to understand the pathogen, and many Candida albicans genes are named after their S. cerevisiae orthologs. Direct studies on the pathogen have, however, revealed differences in the roles of some orthologous proteins in the two yeasts. We show that the Put3 transcription factor allows the pathogen to completely degrade proline to usable nitrogen and carbon by evading regulatory restrictions imposed on its S. cerevisiae ortholog, which mandates conditional use of proline only as a nitrogen source in the baker’s yeast. The ability of Candida albicans to freely obtain nutrients from multiple sources may help it thrive as a commensal and opportunistic pathogen. The zinc cluster transcription factor Put3 was initially characterized in Saccharomyces cerevisiae as the transcriptional activator of PUT1 and PUT2, two genes acting early in the proline assimilation pathway. We have used phenotypic studies, transcription profiling, and chromatin immunoprecipitation with microarray technology (ChIP-chip) to establish that unlike S. cerevisiae, which only uses proline as a nitrogen source, Candida albicans can use proline as a nitrogen source, a carbon source, or a source of both nitrogen and carbon. However, a C. albicans put3 null mutant cannot grow on proline, suggesting that as in S. cerevisiae, C. albicans Put3 (CaPut3) is required for proline catabolism, and because the C. albicans put3 null mutant grew efficiently on glutamate as the sole carbon or nitrogen source, it appears that CaPut3 also regulates the early genes of the pathway. CaPut3 showed direct binding to the CaPUT1 promoter, and both PUT1 and PUT2 were upregulated in response to proline addition in a Put3-dependent manner, as well as in a C. albicans strain expressing a hyperactive Put3. CaPut3 directs proline degradation even in the presence of a good nitrogen source such as ammonia, which contrasts with S. cerevisiae Put3 (ScPut3)-regulated proline catabolism, which only occurs in the absence of a rich nitrogen source. Thus, while overall proline regulatory circuitry differs between S. cerevisiae and C. albicans, the specific role of Put3 appears fundamentally conserved. IMPORTANCE Candida albicans poses a significant threat to the lives of immunocompromised people. Historically, knowledge has been drawn from studies on Saccharomyces cerevisiae to understand the pathogen, and many Candida albicans genes are named after their S. cerevisiae orthologs. Direct studies on the pathogen have, however, revealed differences in the roles of some orthologous proteins in the two yeasts. We show that the Put3 transcription factor allows the pathogen to completely degrade proline to usable nitrogen and carbon by evading regulatory restrictions imposed on its S. cerevisiae ortholog, which mandates conditional use of proline only as a nitrogen source in the baker’s yeast. The ability of Candida albicans to freely obtain nutrients from multiple sources may help it thrive as a commensal and opportunistic pathogen.


2016 ◽  
Vol 82 (10) ◽  
pp. 3052-3060 ◽  
Author(s):  
Rosa de Llanos ◽  
Carlos Andrés Martínez-Garay ◽  
Josep Fita-Torró ◽  
Antonia María Romero ◽  
María Teresa Martínez-Pastor ◽  
...  

ABSTRACTFungi, including the yeastSaccharomyces cerevisiae, lack ferritin and use vacuoles as iron storage organelles. This work explored how plant ferritin expression influenced baker's yeast iron metabolism. Soybean seed ferritin H1 (SFerH1) and SFerH2 genes were cloned and expressed in yeast cells. Both soybean ferritins assembled as multimeric complexes, which bound yeast intracellular ironin vivoand, consequently, induced the activation of the genes expressed during iron scarcity. Soybean ferritin protected yeast cells that lacked the Ccc1 vacuolar iron detoxification transporter from toxic iron levels by reducing cellular oxidation, thus allowing growth at high iron concentrations. Interestingly, when simultaneously expressed inccc1Δ cells, SFerH1 and SFerH2 assembled as heteropolymers, which further increased iron resistance and reduced the oxidative stress produced by excess iron compared to ferritin homopolymer complexes. Finally, soybean ferritin expression led to increased iron accumulation in both wild-type andccc1Δ yeast cells at certain environmental iron concentrations.IMPORTANCEIron deficiency is a worldwide nutritional disorder to which women and children are especially vulnerable. A common strategy to combat iron deficiency consists of dietary supplementation with inorganic iron salts, whose bioavailability is very low. Iron-enriched yeasts and cereals are alternative strategies to diminish iron deficiency. Animals and plants possess large ferritin complexes that accumulate, detoxify, or buffer excess cellular iron. However, the yeastSaccharomyces cerevisiaelacks ferritin and uses vacuoles as iron storage organelles. Here, we explored how soybean ferritin expression influenced yeast iron metabolism, confirming that yeasts that express soybean seed ferritin could be explored as a novel strategy to increase dietary iron absorption.


2012 ◽  
Vol 78 (11) ◽  
pp. 4008-4016 ◽  
Author(s):  
Daisuke Watanabe ◽  
Yuya Araki ◽  
Yan Zhou ◽  
Naoki Maeya ◽  
Takeshi Akao ◽  
...  

ABSTRACTSake yeast cells have defective entry into the quiescent state, allowing them to sustain high fermentation rates. To reveal the underlying mechanism, we investigated the PAS kinase Rim15p, which orchestrates initiation of the quiescence program inSaccharomyces cerevisiae. We found that Rim15p is truncated at the carboxyl terminus in modern sake yeast strains as a result of a frameshift mutation. Introduction of this mutation or deletion of the full-lengthRIM15gene in a laboratory strain led to a defective stress response, decreased synthesis of the storage carbohydrates trehalose and glycogen, and impaired G1arrest, which together closely resemble the characteristic phenotypes of sake yeast. Notably, expression of a functionalRIM15gene in a modern sake strain suppressed all of these phenotypes, demonstrating that dysfunction of Rim15p prevents sake yeast cells from entering quiescence. Moreover, loss of Rim15p or its downstream targets Igo1p and Igo2p remarkably improved the fermentation rate in a laboratory strain. This finding verified that Rim15p-mediated entry into quiescence plays pivotal roles in the inhibition of ethanol fermentation. Taken together, our results suggest that the loss-of-function mutation in theRIM15gene may be the key genetic determinant of the increased ethanol production rates in modern sake yeast strains.


1988 ◽  
Vol 8 (9) ◽  
pp. 3717-3725
Author(s):  
M Kornuc ◽  
R Altman ◽  
D Harrich ◽  
J Garcia ◽  
J Chao ◽  
...  

The adenovirus early region 3 (E3) promoter is an early viral promoter which is strongly induced by the adenovirus transactivator protein E1A. DNase I footprinting with HeLa cell extracts has identified four factor-binding domains which appear to be involved in basal and E1A-induced transcriptional regulation. These binding domains may bind TATA region-binding factors (site I), the CREB/ATF protein (site II), the AP-1 protein (site III), and nuclear factor I/CTF (site IV). Recently, it has been shown that the DNA-binding domain of transcription factor AP-1 has homology with the yeast transcription factor GCN4 and that the yeast transactivator protein GAL4 is able to stimulate transcription in HeLa cells from promoters containing GAL4-binding sites. These results suggest an evolutionary conservation of both transcription factors and the mechanisms responsible for transcriptional activation in Saccharomyces cerevisiae and higher eucaryotic organisms. To determine whether similar patterns of transcriptional regulation were seen with the E3 promoter in HeLa and yeast cells, the E3 promoter fused to the chloramphenicol acetyltransferase (cat) gene was cloned into a high-copy-number plasmid and stably introduced into yeast cells. S1 analysis revealed that similar E3 promoter mRNA start sites were found in yeast and HeLa cells. DNase I footprinting with partially purified yeast extracts revealed that four regions of the E3 promoter were protected. Several of these regions were similar to binding sites determined by using HeLa cell extracts. Oligonucleotide mutagenesis of these binding domains indicated their importance in the transcriptional regulation of the E3 promoter in yeast cells. These results suggest that similar cellular transcription factor-binding sites may be involved in the regulation of promoters in both yeast and mammalian cells.


2018 ◽  
Vol 85 (1) ◽  
Author(s):  
Daisuke Watanabe ◽  
Takuma Kajihara ◽  
Yukiko Sugimoto ◽  
Kenichi Takagi ◽  
Megumi Mizuno ◽  
...  

ABSTRACT Saccharomyces cerevisiae sake yeast strain Kyokai no. 7 (K7) and its relatives carry a homozygous loss-of-function mutation in the RIM15 gene, which encodes a Greatwall family protein kinase. Disruption of RIM15 in nonsake yeast strains leads to improved alcoholic fermentation, indicating that the defect in Rim15p is associated with the enhanced fermentation performance of sake yeast cells. In order to understand how Rim15p mediates fermentation control, we here focused on target-of-rapamycin protein kinase complex 1 (TORC1) and protein phosphatase 2A with the B55δ regulatory subunit (PP2AB55δ), complexes that are known to act upstream and downstream of Rim15p, respectively. Several lines of evidence, including our previous transcriptomic analysis data, suggested enhanced TORC1 signaling in sake yeast cells during sake fermentation. Fermentation tests of the TORC1-related mutants using a laboratory strain revealed that TORC1 signaling positively regulates the initial fermentation rate in a Rim15p-dependent manner. Deletion of the CDC55 gene, encoding B55δ, abolished the high fermentation performance of Rim15p-deficient laboratory yeast and sake yeast cells, indicating that PP2AB55δ mediates the fermentation control by TORC1 and Rim15p. The TORC1-Greatwall-PP2AB55δ pathway similarly affected the fermentation rate in the fission yeast Schizosaccharomyces pombe, strongly suggesting that the evolutionarily conserved pathway governs alcoholic fermentation in yeasts. It is likely that elevated PP2AB55δ activity accounts for the high fermentation performance of sake yeast cells. Heterozygous loss-of-function mutations in CDC55 found in K7-related sake strains may indicate that the Rim15p-deficient phenotypes are disadvantageous to cell survival. IMPORTANCE The biochemical processes and enzymes responsible for glycolysis and alcoholic fermentation by the yeast S. cerevisiae have long been the subject of scientific research. Nevertheless, the factors determining fermentation performance in vivo are not fully understood. As a result, the industrial breeding of yeast strains has required empirical characterization of fermentation by screening numerous mutants through laborious fermentation tests. To establish a rational and efficient breeding strategy, key regulators of alcoholic fermentation need to be identified. In the present study, we focused on how sake yeast strains of S. cerevisiae have acquired high alcoholic fermentation performance. Our findings provide a rational molecular basis to design yeast strains with optimal fermentation performance for production of alcoholic beverages and bioethanol. In addition, as the evolutionarily conserved TORC1-Greatwall-PP2AB55δ pathway plays a major role in the glycolytic control, our work may contribute to research on carbohydrate metabolism in higher eukaryotes.


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