Effect of extracellular acidification on the activity of plasma membrane ATPase and on the cytosolic and vacuolar pH of Saccharomyces cerevisiae

Abstract

The rapid in vivo activation of Saccharomyces cerevisiae plasma membrane H+-ATPase that has been attributed to medium acidification from pH 6.5 to pH 3.5 is not caused by the low pH itself but is induced by the weak organic acid (succinic) used as the acidulant. The activation induced by 50 mM succinic acid at pH 3.5 occurred in both the presence or absence of glucose. Activation at pH 3.5 was also induced by acetic acid and it was maximal at 50 mM concentration. To investigate the role of plasma membrane ATPase activation in pH homeostasis, the internal pH (cytosolic and vacuolar) of yeast cells incubated in media at pH 6.5 or at pH 3.5, acidified either with HCl or acetic acid, were compared by using in vivo (31)P-NMR. Despite plasma membrane ATPase activation by acetic acid, the decrease in cytosolic pH caused by external acidification was much more important when the permeant acetic acid was used instead of HCl as the acidulant. The supplementation of the incubation medium at pH 3.5 with glucose led to higher cytosolic pH values, consistent with the observed in vivo activation of plasma membrane ATPase by glucose. At the external pH value of 6.5 the vacuole was maintained at a mildly acidic pH (around 6) while the cytosol was at about neutral pH; however, when cytoplasmic pH decreased due to external acidification, vacuolar pH accompanied that decrease. Vacuolar pH reached 5.4-5.5 during incubation with HCI and dropped sharply to values below 4.4 in cells incubated with acetic acid. These results indicate that the vacuole also plays a role in homeostasis of the intracellular pH.