Abstract
Vam7p, the vacuolar soluble Qc-SNARE, is essential for yeast vacuole fusion. The large tethering complex, homotypic fusion and vacuole protein sorting complex (HOPS), and phosphoinositides, which interact with the Vam7p PX domain, have each been proposed to serve as its membrane receptors. Studies with the isolated organelle cannot determine whether these receptor elements suffice and whether ligands or mutations act directly or indirectly on Vam7p binding to the membrane. Using pure components that are active in reconstituted vacuolar fusion, we now find that Vam7p binds to membranes through its combined affinities for several vacuolar membrane constituents: HOPS, phosphatidylinositol 3-phosphate, SNAREs, and acidic phospholipids. Acidic lipids allow low concentrations of Vam7p to suffice for fusion; without acidic lipids, the block to fusion is partially bypassed by high concentrations of Vam7p.
Highlights
The soluble SNARE Vam7p is recruited to membranes to form trans-SNARE complexes for fusion
The large tethering complex, homotypic fusion and vacuole protein sorting complex (HOPS), and phosphoinositides, which interact with the Vam7p PX domain, have each been proposed to serve as its membrane receptors
We show that Vam7p can be brought to the membrane through the combination of its moderate affinities for phosphoinositides, SNAREs, acidic lipids, and HOPS
Summary
The soluble SNARE Vam7p is recruited to membranes to form trans-SNARE complexes for fusion. Vacuoles undergo an initial reversible tethering that helps bring membranes into apposition, followed by the enrichment of fusion-specific factors around the edge of the apposed membrane domain [16] The small vacuolar Rab GTPase Ypt7p is required for fusion [1] It binds its effector complex, HOPS2 [3], a large multisubunit tethering factor. Certain lipids are important for fusion, including phosphoinositides (19 –21), ergosterol [22], and diacylglycerol [23, 24] We have reconstituted this complex process of yeast vacuole fusion in vitro [12, 13] using purified vacuolar components, the SNAREs, SNARE chaperones, HOPS, Ypt7p, and lipids of vacuolar compositions in reconstituted proteoliposomes. The “true” receptor is a cooperating set of binding interactions with other fusion factors, both peripheral membrane proteins (HOPS) and integral membrane proteins (other SNAREs) as well as specific lipids (PI[3]P and acidic lipids)
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
