Sec18p (NSF)-Driven Release of Sec17p (α-SNAP) Can Precede Docking and Fusion of Yeast Vacuoles

and the plasma membrane in epithelial cells requires S.cerevisiae inherits its vacuole by projecting vacuoleNSF for the basolateral route, but probably not for the derived membrane vesicles and tubules into the bud, apical route (Ikonen et al., 1995). Homotypic ER to ER where they fuse to establish the daughter vacuole. fusion and nuclear fusion during yeast mating depend on This homotypic fusion event can be assayed in vitro. Cdc48p, an NSF-related protein (Latterich et al., 1995). It requires Sec17p and Sec18p, the homologs of the Together with NSF, the mammalian Cdc48 homolog p97 mammalian a-SNAP and NSF, which cooperate in mulis also necessary for rebuilding the Golgi from vesicular tiple steps of membrane trafficking. We now report Golgi fragments (Rabouille et al., 1995; Acharya et al., that Sec17p, Sec18p, and ATP are only needed for 1995). an early stage of the reaction that results in Sec17p Homotypic vacuole fusion, which occurs during vacurelease. Sec17p and Sec18p actions precede, and are ole inheritance in yeast, depends on the a-SNAP homoneeded for, the step employing the Ras-like GTPase log Sec17p and the NSF homolog Sec18p (Haas and Ypt7p. Sec18p-driven release of Sec17p can even preWickner, submitted). Both proteins are present on the cede vacuole docking, as it can occur prior to mixing of vacuoles. Vacuole fusion is sensitive to antibodies or vacuoles and is insensitive to vacuole concentration. Fab fragments against Sec18p or Sec17p. Purified Sec17p and Sec18p thus may function in a predocking Sec18p stimulates the reaction and reverses a block by stage of the reaction, rather than in bilayer fusion per Sec18p antibodies. There is no indication of Cdc48p se. involvement, since Cdc48p antibodies do not inhibit the