A Loss-of-Function Mutation in the PAS Kinase Rim15p Is Related to Defective Quiescence Entry and High Fermentation Rates of Saccharomyces cerevisiae Sake Yeast Strains

Abstract

b Sake 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 in Saccharomyces 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-length RIM15 gene in a laboratory strain led to a defective stress response, decreased synthesis of the storage carbohydrates trehalose and glycogen, and impaired G 1 arrest, which together closely resemble the characteristic phenotypes of sake yeast. Notably, expression of a functional RIM15 gene 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 the RIM15 gene may be the key genetic determinant of the increased ethanol production rates in modern sake yeast strains.

Assessing the Mechanisms Responsible for Differences between Nitrogen Requirements of Saccharomyces cerevisiae Wine Yeasts in Alcoholic Fermentation

“…The differences in gene expression and the physiological responses are reminiscent of the observations of Watanabe et al (41), who analyzed the effects of a mutated form of the PAS kinase RIM15 in sake yeast. They showed that the altered form of RIM15 reduced the yeast stress response and the amount of storage carbohydrate and enhanced the fermentation rate.…”

Accelerated Alcoholic Fermentation Caused by Defective Gene Expression Related to Glucose Derepression inSaccharomyces cerevisiae

“…In sake yeast strain K701, unknown defects in the latter mechanism might have caused the observed decrease in the activities of Adr1p and Cat8p. We postulate that the Rim15p-Msn2/4p pathway, which is specifically inactivated in modern sake strains, 3,4) is indirectly involved in the maintenance of Adr1p and Cat8p activities. As shown in Table 2, however, deletion of the RIM15 gene did not decrease the ADH2 mRNA level in X2180-1A.…”

“…We have found that defects in the stress responses mediated by Rim15p, 3) Msn2/ 4p, 4) and Hsf1p 52) contribute to enhancement of the initial fermentation rate in modern sake yeast strains. These findings, together with our present results, indicate that sake yeast has multiple cellular functions that have been impaired during years of selection and domestication, and that are unnecessary for or negatively affect efficient alcoholic fermentation.…”

“…5,6) Due to mutations in the RIM15 and MSN4 genes, modern sake yeast cells in the stationary phase share common phenotypes, including low thermotolerance, ethanol susceptibility, low buoyant density, decreased production of reserve carbohydrates trehalose and glycogen, and impaired cell-cycle G1 arrest. 3,4,7,8) Furthermore, deletion of these genes significantly enhances the fermentation rates of non-sake yeast strains, 3,4,9) suggesting that the rim15 5055insA and msn4…”

“…In previous studies, we identified modern sake yeastspecific loss-of-function mutations in the RIM15 (rim15 5055insA ) 3) and MSN4 (msn4 C1540T ) 4) genes, both of which are involved in entry into quiescence during the stationary phase of cell growth. Under stress conditions, yeast Per-Arnt-Sim kinase (PASK) Rim15p activates a quiescent gene expression program mediated by redundant stress-responsive transcription factors Msn2p and Msn4p (Msn2/4p).…”