Dataset Information

Hsp104-dependent ability to assimilate mannitol and sorbitol conferred by a truncated Cyc8 with a C-terminal polyglutamine in Saccharomyces cerevisiae.

ABSTRACT: Tup1-Cyc8 (also known as Tup1-Ssn6) is a general transcriptional corepressor. D-Mannitol (mannitol) and D-sorbitol (sorbitol) are the major polyols in nature. Budding yeast Saccharomyces cerevisiae is unable to assimilate mannitol or sorbitol, but acquires the ability to assimilate mannitol due to a spontaneous mutation in TUP1 or CYC8. In this study, we found that spontaneous mutation of TUP1 or CYC8 also permitted assimilation of sorbitol. Some spontaneous nonsense mutations of CYC8 produced a truncated Cyc8 with a C-terminal polyglutamine. The effects were guanidine hydrochloride-sensitive and were dependent on Hsp104, but were complemented by introduction of CYC8, ruling out involvement of a prion. Assimilation of mannitol and sorbitol conferred by other mutations of TUP1 or CYC8 was guanidine hydrochloride-tolerant. It is physiologically reasonable that S. cerevisiae carries this mechanism to acquire the ability to assimilate major polyols in nature.

SUBMITTER: Tanaka H

PROVIDER: S-EPMC7657529 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Hsp104-dependent ability to assimilate mannitol and sorbitol conferred by a truncated Cyc8 with a C-terminal polyglutamine in Saccharomyces cerevisiae.

Tanaka Hideki H Murata Kousaku K Hashimoto Wataru W Kawai Shigeyuki S

PloS one 20201111 11

Tup1-Cyc8 (also known as Tup1-Ssn6) is a general transcriptional corepressor. D-Mannitol (mannitol) and D-sorbitol (sorbitol) are the major polyols in nature. Budding yeast Saccharomyces cerevisiae is unable to assimilate mannitol or sorbitol, but acquires the ability to assimilate mannitol due to a spontaneous mutation in TUP1 or CYC8. In this study, we found that spontaneous mutation of TUP1 or CYC8 also permitted assimilation of sorbitol. Some spontaneous nonsense mutations of CYC8 produced a ...[more]

PMID: 33175887

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Project description:Saccharomyces cerevisiae normally cannot assimilate mannitol, a promising brown macroalgal carbon source for bioethanol production. To date, the molecular mechanisms underlying this inability remain unknown. Here, we found that cells acquiring mannitol-assimilating ability appeared from wild-type S. cerevisiae strain during prolonged culture in mannitol medium. Our microarray analysis revealed that genes for putative mannitol dehydrogenase and hexose transporters were up-regulated in cells acquiring mannitol-assimilating ability. Take account of our other results including complementation analysis and cell growth data, we demonstrated that this acquisition of mannitol-assimilating ability was due to the spontaneous mutation in the gene encoding Tup1 or Cyc8. Tup1-Cyc8 is the general corepressor complex involved in the repression of many kinds of genes. Thus, it is suggested that the inability of wild-type S. cerevisiae to assimilate mannitol can be attributed to the transcriptional repression of a set of genes involved in mannitol utilization by Tup1-Cyc8 corepressor. In other words, Tup1-Cyc8 is a key regulator of mannitol metabolism in S. cerevisiae. We also showed that S. cerevisiae strain which carries mutant allele of TUP1 or CYC8 produced ethanol from mannitol efficiently. Especially, strain carrying mutant allele of CYC8 showed high tolerance to salt, which is superior to other ethanologenic microorganisms. This characteristic is highly beneficial to produce bioethanol from marine biomass. Taken together, Tup1-Cyc8 can be an ideal target to develop a yeast-algal bioethanol production system. To figure out how Mtl+ strains (cells acquiring ability to grow in mannitol medium) had acquired the ability to assimilate mannitol, we performed genome-wide analysis by using Nimblegen microarrays. Yeast Saccharomyces cerevisiae cells (wild-type BY4742 strain and two Mtl+ strains, MK3619 and MK3683) were grown at 30Â°C to the logarithmic phase in SC or SM media. Total RNA was purified and the 4 RNA samples (BY4742 cells in SC as control, MK3619 cells in SM, MK3683 cells in both SC and SM) were analyzed with Nimblegen microarrays.

Project description:Engineering Saccharomyces cerevisiae for the utilization of pentose sugars is an important goal for the production of second-generation bioethanol and biochemicals. However, S. cerevisiae lacks specific pentose transporters, and in the presence of glucose, pentoses enter the cell inefficiently via endogenous hexose transporters (HXTs). By means of in vivo engineering, we have developed a quadruple hexokinase deletion mutant of S. cerevisiae that evolved into a strain that efficiently utilizes d-xylose in the presence of high d-glucose concentrations. A genome sequence analysis revealed a mutation (Y353C) in the general corepressor CYC8, or SSN6, which was found to be responsible for the phenotype when introduced individually in the nonevolved strain. A transcriptome analysis revealed altered expression of 95 genes in total, including genes involved in (i) hexose transport, (ii) maltose metabolism, (iii) cell wall function (mannoprotein family), and (iv) unknown functions (seripauperin multigene family). Of the 18 known HXTs, genes for 9 were upregulated, especially the low or nonexpressed HXT10, HXT13, HXT15, and HXT16 Mutant cells showed increased uptake rates of d-xylose in the presence of d-glucose, as well as elevated maximum rates of metabolism (Vmax) for both d-glucose and d-xylose transport. The data suggest that the increased expression of multiple hexose transporters renders d-xylose metabolism less sensitive to d-glucose inhibition due to an elevated transport rate of d-xylose into the cell.IMPORTANCE The yeast Saccharomyces cerevisiae is used for second-generation bioethanol formation. However, growth on xylose is limited by pentose transport through the endogenous hexose transporters (HXTs), as uptake is outcompeted by the preferred substrate, glucose. Mutant strains were obtained with improved growth characteristics on xylose in the presence of glucose, and the mutations mapped to the regulator Cyc8. The inactivation of Cyc8 caused increased expression of HXTs, thereby providing more capacity for the transport of xylose, presenting a further step toward a more robust process of industrial fermentation of lignocellulosic biomass using yeast.

Dataset Information

Hsp104-dependent ability to assimilate mannitol and sorbitol conferred by a truncated Cyc8 with a C-terminal polyglutamine in Saccharomyces cerevisiae.

Publications

Hsp104-dependent ability to assimilate mannitol and sorbitol conferred by a truncated Cyc8 with a C-terminal polyglutamine in Saccharomyces cerevisiae.

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