DOMINANT-NEGATIVE ALLELES OF SEC10 REVEAL DISTINCT DOMAINS INVOLVED IN SECRETION AND MORPHOGENESIS IN YEAST

The accurate targeting of secretory vesicles to distinct sites on the plasma membrane is necessary to achieve polarized growth and to establ ish specialized domains at the surface of eukaryotic cells. Members of a protein complex required for exocytosis, the exocyst, have been loc alized to regions of active secretion in the budding yeast Saccharomyc es cerevisiae where they may function to specify sites on the plasma m embrane for vesicle docking and fusion. In this study we have addresse d the function of one member of the exocyst complex, Sec10p. We have i dentified two functional domains of Sec10p that act in a dominant-nega tive manner to inhibit cell growth upon overexpression. Phenotypic and biochemical analysis of the dominant-negative mutants points to a bif unctional role for Sec10p. One domain, consisting of the amino-termina l two-thirds of Sec10p directly interacts with Sec15p, another exocyst component. Overexpression of this domain displaces the full-length Se c10 from the exocyst complex, resulting in a block in exocytosis and a n accumulation of secretory vesicles. The carboxy-terminal domain of S ec10p does not interact with other members of the exocyst complex and expression of this domain does not cause a secretory defect. Rather, t his mutant results in the formation of elongated cells, suggesting tha t the second domain of Sec10p is required for morphogenesis, perhaps r egulating the reorientation of the secretory pathway from the tip of t he emerging daughter cell toward the mother-daughter connection during cell cycle progression.