title: Promoter engineering of the Saccharomyces cerevisiae RIM15 gene for improvement of alcoholic fermentation rates under stress conditions.
abstract: A loss-of-function mutation in the RIM15 gene, which encodes a Greatwall-like protein kinase, is one of the major causes of the high alcoholic fermentation rates in Saccharomyces cerevisiae sake strains closely related to Kyokai no. 7 (K7). However, impairment of Rim15p may not be beneficial under more severe fermentation conditions, such as in the late fermentation stage, as it negatively affects stress responses. To balance stress tolerance and fermentation performance, we inserted the promoter of a gluconeogenic gene, PCK1, into the 5'-untranslated region (5'-UTR) of the RIM15 gene in a laboratory strain to achieve repression of RIM15 gene expression in the glucose-rich early stage with its induction in the stressful late stage of alcoholic fermentation. The promoter-engineered strain exhibited a fermentation rate comparable to that of the RIM15-deleted strain with no decrease in cell viability. The engineered strain achieved better alcoholic fermentation performance than the RIM15-deleted strain under repetitive and high-glucose fermentation conditions. These data demonstrated the validity of promoter engineering of the RIM15 gene that governs inhibitory control of alcoholic fermentation.
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, 8916-5 Takayama, Ikoma, Nara 630-0192, Japan.
authors.forename: Daisuke
authors.initials: D
authors.lastname: Watanabe
authors.affiliation: Department of Integrated Biosciences, Graduate School of Frontier Sciences, University of Tokyo, 5-1-5 Kashiwanoha, Kashiwa, Chiba 277-8562, Japan.
authors.forename: Akie
authors.initials: A
authors.lastname: Kaneko
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, 8916-5 Takayama, Ikoma, Nara 630-0192, Japan.
authors.forename: Yukiko
authors.initials: Y
authors.lastname: Sugimoto
authors.affiliation: Department of Integrated Biosciences, Graduate School of Frontier Sciences, University of Tokyo, 5-1-5 Kashiwanoha, Kashiwa, Chiba 277-8562, Japan.
authors.forename: Shinsuke
authors.initials: S
authors.lastname: Ohnuki
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, 8916-5 Takayama, Ikoma, Nara 630-0192, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Takagi
authors.affiliation: Department of Integrated Biosciences, Graduate School of Frontier Sciences, University of Tokyo, 5-1-5 Kashiwanoha, Kashiwa, Chiba 277-8562, Japan. Electronic address: ohya@k.u-tokyo.ac.jp.
authors.forename: Yoshikazu
authors.initials: Y
authors.lastname: Ohya
mesh_terms: D000220:Adaptation, Biological / Q000235:genetics; D000434:Alcoholic Beverages; D003001:Cloning, Molecular; D000431:Ethanol / Q000378:metabolism*; D005285:Fermentation / Q000235:genetics*; D005947:Glucose / Q000378:metabolism; D060847:Metabolic Engineering / Q000379:methods*; D030781:Organisms, Genetically Modified; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011494:Protein Kinases / Q000235:genetics* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism; D013312:Stress, Physiological / Q000235:genetics
chemical_list: D029701:Saccharomyces cerevisiae Proteins; D000431:Ethanol; D011494:Protein Kinases; C410736:Rim15 protein, S cerevisiae; D005947:Glucose
keywords: Alcoholic fermentation; Gluconeogenesis; Glucose derepression; Glucose repression; Greatwall protein kinase; PCK1; Promoter engineering; RIM15; Saccharomyces cerevisiae; Stress response