ENDOCYTOSIS IS REQUIRED FOR THE GROWTH OF VACUOLAR H+-ATPASE-DEFECTIVE YEAST - IDENTIFICATION OF 6 NEW END GENES

Yeast mutants that are defective in acidification of the lysosome-like vacuole are able to grow at pH 5.5, but not at pH 7. Here, we present evidence that endocytosis is required for this low pH-dependent growth and use this observation to develop a screen for mutants defective in endocytosis. By isolating mutants that cannot grow when they lack the 60-kD vacuolar ATPase subunit (encoded by the VAT2 gene), we isolated a number of vat2-synthetic lethal (Vsl(-)) mutant strains. Seven of the Vsl(-) mutants are defective in endocytosis. Four of these mutant strains(end8-1, end9-1, end10-1, and end11-1) show altered up take of the endocytosed ligand, alpha-factor, and three (end12-1, end12-2, and ead13-1) are probably defective in transfer of internalized material to the vacuole. Most of the mutations also confer a strong Ts(-) growth defect. The mutants defective in uptake of alpha-factor sort newly synthesized vacuolar proteins correctly, while those which may be defective in subsequent transport steps secrete at least a fraction of the newly synthesized soluble vacuolar proteins. The mutations that result in a defect in alpha-factor uptake are not allelic to any of the genes previously shown to encode endocytic functions.