Activity of plasma membrane H+-ATPase and expression of PMA1 and PMA2 genes in Saccharomyces cerevisiae cells grown at o
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© Springer-Verlag 1996
O R I G I N A L PA P E R
V. Carmelo · P. Bogaerts · I. Sá-Correia
Activity of plasma membrane H+-ATPase and expression of PMA1 and PMA2 genes in Saccharomyces cerevisiae cells grown at optimal and low pH
Received: 22 April 1996 / Accepted: 6 August 1996
Abstract Cells of Saccharomyces cerevisiae grown in media with an initial pH of 2.5–6.0, acidified with a strong acid (HCl), exhibited the highest plasma membrane H+-ATPase-specific activity at an initial pH of 6.0. At a lower pH (above pH 2.5) ATPase activity (62–83% of the maximum level) still allowed optimal growth. At pH 2.5, ATPase activity was about 30% of the maximum value and growth was impaired. Quantitative immunoassays showed that the content of ATPase protein in the plasma membrane was similar across the entire pH range tested, although slightly lower at pH 2.5. The decrease of plasma membrane ATPase activity in cells grown at low pH was partially accounted for by its in vitro stability, which decreased sharply at pH below 5.5, although the reduction of activity was far below the values expected from in vitro measurements. Yeast growth under acid stress changed the pattern of gene expression observed at optimal pH. The level of mRNA from the essential plasma-membrane-ATPase-encoding gene PMA1 was reduced by 50% in cells grown at pH 2.5 as compared with cells grown at the optimal pH 5.0, although the content of ATPase in the plasma membrane was only modestly reduced. As observed in response to other kinds of stress, the PMA2 promoter at the optimal pH was up to eightfold more efficient in cells grown at pH 2.5, although it remained several hundred times less efficient than that of the PMA1 gene.
V. Carmelo · I. Sá-Correia (Y) Laboratório de Engenharia Bioquímica, Centro de Engenharia Biológica e Química, Instituto Superior Técnico, Av. Rovisco Pais, P-1096 Lisboa Codex, Portugal Tel. +351-1-8417233; Fax +351-1-8480072 e-mail: [email protected] P. Bogaerts Unité de Biochimie Physiologique, Université Catholique de Louvain, Place Croix du Sud 2–20, B-1348 Louvain-la-Neuve, Belgium
Key words Plasma membrane H+-ATPase · Saccharomyces cerevisiae · Low pH · PMA1 gene expression · PMA2 gene expression
Introduction Most Saccharomyces strains grow at pH values of 2.5–8.5, and the kinetics of growth and fermentation are not affected between pH 3.5 and 6.0 because of the tight control of intracellular pH (Matthews and Webb 1991). The proton-translocating ATPase in the plasma membrane of yeast regulates intracellular pH by pumping protons out of the cells. The proton electrochemical potential created drives the secondary transport of nutrients (Goffeau and Slayman 1981; Serrano 1988). An appropriate level of activity is essential and rate limiting to cell growth (Serrano et al. 1986; Serrano 1988; Rao et al. 1993). Like glucose (Serrano 1983), several types of environmental stress, such as subcritical inhibitory concentrations of ethanol (Rosa and Sá-Correia 1991; Monteiro et al. 1994) or octanoic acid (Viegas and Sá-Correia 1991; Vieg
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