Epsilon Open Archive

Abstract

Syntaxins are required for fusion of membranes in eukaryotic cells and belong to a group of proteins known as t-SNAREs. This thesis primarily focuses on the role of the plasma membrane syntaxins Sso1p and Sso2p in the yeast Saccharomyces cerevisiae. The plasma membrane syntaxins are required for viability in yeast, but in the vegetatively growing cell, the Sso proteins have seemingly reduntant functions. We generated a mutant allele of SSO2, sso2-1, that has a conditional lethal phenotype in the absence of SSO1. Overexpression of genes coding for other SNARE proteins; Sec9p, Snc1p and Snc2p, suppressed the lethal phenotype. The corresponding mutant allele of SSO1, sso1-1, is also temperature-sensitive and interacts synthetically with a disruption of MSO1, which codes for a Sec1p interacting protein. Most notably, both SSO1 and MSO1, but not SSO2, were shown to be necessary for spore formation during meiosis. Mapping of functions within the Sso1p protein showed that a region in the N-terminus of Sso1p is needed for efficient sporulation. Unexpectedly, the 3’-untranslated region of SSO1 is absolutely required for sporulation and also sufficient to enable some spore formation when fused to the SSO2 open reading frame. Inspection of the sso1/sso1 phenotype during sporulation using transmission electron microscopy showed that prospore membrane assembly at the meiotic plaque of the spindle pole body is completely blocked in the mutant. A second part of this thesis deals with screening for uncharacterized genes involved in intracellular transport by exposing deletion mutants for drugs known to inhibit intracellular transport. The screen identified two new genes whose deletions made the cell sensitive to monensin, and those were given the names MON1 and MON2. Five new genes caused sensitivity to Brefeldin A when deleted, and were named BRE1-BRE5.