The ATPase inhibitor Dio-9 effectively suppressed a number of physiological processes in a wild-type strain of Saccharomyces cerevisiae, X2180-1A. Low levels of the antibiotic inhibited cell growth, amino acid transport, hydrogen ion efflux, and ATPase activity. In addition, Dio-9 acted as a permeabilizing agent for the yeast plasma membrane. A mutant yeast strain, XC24, was selected on the basis of its ability to grow on minimal medium containing 200 μg/ml of Dio-9. Strain XC24 had acquired a pH-conditional ability to resist the permeabilizing effects of Dio-9. In addition, amino acid transport and hydrogen ion pumping exhibited a reduced senstivity to Dio-9 at low pH in the mutant strain. Strain XC24 was also resistant to the permeabilizing effects of the basic polymers protamine and deacylated chitin.