A CONDITIONAL STEROL ESTERIFICATION DEFECT IN YEAST HAVING EITHER A SEC1 OR SEC5 MUTATION IN THE SECRETORY PATHWAY

Two temperature-conditional secretory mutations, sec1 and sec5 cause t he accumulation of post-Golgi vesicles when strains containing these m utations are grown at 37 degrees C. In addition to accumulating vesicl es, the mutants do not esterify free sterol on rich media at the restr ictive temperature. It is the high level of inositol in the media that causes this condition in the yeast Saccharomyces cerevisiae, not a de fective steryl ester synthase or lack of substrates. When strains cont aining the SEC1 or SEC5 mutation were transformed separately with a pl asmid carrying SEC1 and SECS, the esterification and secretory defects were alleviated. Double mutants containing sec6, sec14 or sec18 with either a sec1 or sec5 mutation have normal esterification levels. Stra ins with suppressor mutations were isolated that grew at 37 degrees C, esterified sterols and had diminished accumulation of vesicles, when grown at the restrictive temperature on defined media with additional inositol. Electron microscopy was used to examine vesicle accumulation . the number of lipid droplets, and to further characterize the esteri fication defect. When grown at 37 degrees C on defined medium, the str ains with sec5 or sec1 accumulated the usual secretory vesicles, but w hen grown under similar conditions with elevated levels of inositol, a ccumulated an additional vesicular-like body. (C) 1997 by John Wiley & Sons, Ltd.