Sec18p (NSF)-driven release of Sec17p (alpha-SNAP) can precede docking and fusion of yeast vacuoles.
S. cerevisiae inherits its vacuole by projecting vacuole-derived membrane vesicles and tubules into the bud, where they fuse to establish the daughter vacuole. This homotypic fusion event can be assayed in vitro. It requires Sec17p and Sec18p, the homologs of the mammalian alpha-SNAP and NSF, which cooperate in multiple steps of membrane trafficking. We now report that Sec17p, Sec18p, and ATP are only needed for an early stage of the reaction that results in Sec17p release. Sec17p and Sec18p actions precede, and are needed for, the step employing the Ras-like GTPase Ypt7p. Sec18p-driven release of Sec17p can even precede vacuole docking, as it can occur prior to mixing of vacuoles and is insensitive to vacuole concentration. Sec17p and Sec18p thus may function in a predocking stage of the reaction, rather than in bilayer fusion per se.
Mayer
A
A
Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire, 03755-3844 USA.
Wickner
W
W
Haas
A
A
D000251:Adenosine Triphosphatases; D002352:Carrier Proteins / Q000378:metabolism*; D005656:Fungal Proteins / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism; D007700:Kinetics; D008561:Membrane Fusion / Q000502:physiology; D008565:Membrane Proteins / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D013997:Time Factors; D014617:Vacuoles / Q000737:chemistry / Q000378:metabolism*; D033921:Vesicular Transport Proteins; D020691:rab GTP-Binding Proteins; D018631:ras Proteins / Q000378:metabolism
D002352:Carrier Proteins; D005656:Fungal Proteins; D008565:Membrane Proteins; C075137:SEC17 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D033921:Vesicular Transport Proteins; D000251:Adenosine Triphosphatases; D019204:GTP-Binding Proteins; C064404:SEC18 protein, S cerevisiae; C079005:YPT7 protein, S cerevisiae; D020691:rab GTP-Binding Proteins; D018631:ras Proteins