2 NEW YPT GTPASES ARE REQUIRED FOR EXIT FROM THE YEAST TRANS-GOLGI COMPARTMENT

Small GTPases of the Ypt/rab family are involved in the regulation of vesicular transport. These GTPases apparently function during the targ eting of vesicles to the acceptor compartment. Two members of the Ypt/ rab family, Ypt1p and Sec4p, have been shown to regulate early and lat e steps of the yeast exocytic pathway, respectively. Here we tested th e role of two newly identified GTPases, Ypt31p and Ypt32p. These two p roteins share 81% identity and 90% similarity, and belong to the same protein subfamily as Ypt1p and Sec4p. Yeast cells can tolerate deletio n of either the YPT31 or the YPT32 gene, but not both. These observati ons suggest that Ypt31p and Ypt32p perform identical or overlapping fu nctions. Cells deleted for the YPT31 gene and carrying a conditional y pt32 mutation exhibit protein transport defects in the late exocytic p athway, but not in vacuolar protein sorting. The ypt31/32 mutant secre tory defect is clearly downstream from that displayed by a ypt1 mutant and is similar to that of sec4 mutant cells. However, electron micros copy revealed that while sec4 mutant cells accumulate secretory vesicl es, ypt31/32 mutant cells accumulate aberrant Golgi structures. The yp t31/32 phenotype is epistatic to that of a sec1 mutant, which accumula tes secretory vesicles. Together, these results indicate that the Ypt3 1/32p GTPases are required for a step that occurs in the trans-Golgi c ompartment, between the reactions regulated by Ypt1p and Sec4p. This s tep might involve budding of vesicles from the trans-Golgi. Alternativ ely, Ypt31/32p might promote secretion indirectly, by allowing fusion of recycling vesicles with the trans-Golgi compartment.