Osmohomeostasis and vacuole biogenesis genes in the yeast saccharomyces cerevisiae

Abstract

The putative role of the S. cerevisiae vacuole in osmohomeostasis, as well as its biogenesis was analysed by taking a mutational approach. 97 mutants unable to tolerate high concentrations of salt were isolated and examined for aberrant vacuolar phenotypes. A comprehensive phenotypic analysis was able to demonstrate that apart from osmosensitivity most mutations conferred other properties such as altered vacuolar morphology, the inability to perform gluconeogenesis and/or the mislocalization of vacuolar proteins to the cell surface. The mutants fall into at least 20 complementation groups, termed ssv for salt sensitive vacuolar mutants, of which 3 genetically overlap with complementation groups isolated by others. This analysis provides evidence that in 5. cerevisiae correct vacuolar biogenesis is required for osmotolerance as well as other important cellular processes. To elucidate vacuolar osmohomeostasis at the molecular level, one gene, SSV7, was cloned from a genomic DNA library by complementation of a ssv7-l mutation and its sequence determined. It encodes a novel 927 amino acid protein with limited structural homology to the functional domains of two nucleotide exchange factors from 5. cerevisiae, namely CDC25 and BUD5. A mutation in the SSV7 gene confers a pleiotropic phenotype including fragmented vacuoles and the absence of a nucleus. The mutation apparently uncouples mitosis from cell growth, which results in an increased cell size of a ssv7-l mutant. The role of SSV7 in the general concept of vacuole biogenesis is discussed, as is the observed fact that a number of mutants in intermediate biosynthetic pathways confer a vacuolar protein sorting defect.