THE EXOCYST IS A MULTIPROTEIN COMPLEX REQUIRED FOR EXOCYTOSIS IN SACCHAROMYCES-CEREVISIAE

In the yeast Saccharomyces cerevisiae, the products of at least 15 gen es are involved specifically in vesicular transport from the Golgi app aratus to the plasma membrane, Previously, we have shown that three of these genes, SEC6, SEC8 and SEC15, encode components of a multisubuni t complex which localizes to the tip of the bud, the predominant site of exocytosis in S. cerevisiae. Mutations in three more of these genes , SEC3, SEC5 and SEC10, were found to disrupt the subunit integrity of the Sec6-Sec8-Sec15 complex, indicating that these genes may encode s ome of the remaining components of this complex, To examine this possi bility, we cloned and sequenced the SECS and SEC10 genes, disrupted th em, and either epitope tagged them (Sec5p) or prepared polyclonal anti sera (Sec10p) to them for co-immunoprecipitation studies. Concurrently , we biochemically purified the remaining unidentified polypeptides of the Sec6-Sec8-Sec15 complex for peptide microsequencing, The genes en coding these components were identified by comparison of predicted ami no acid sequences with those obtained from peptide microsequencing of the purified complex components, In addition to Sec6p, Sec8p and Sec15 p, the complex contains the proteins encoded by SEC3, SECS, SEC10 and a novel gene, EXO70, Since these seven proteins function together in a complex required for exocytosis, and not other intracellular traffick ing steps, we have named it the Exocyst.