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EMBO J. 2008 Aug 6;27(15):2031-42. doi: 10.1038/emboj.2008.139. Epub 2008 Jul 24.

Reconstituted membrane fusion requires regulatory lipids, SNAREs and synergistic SNARE chaperones.

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1
Department of Biochemistry, Dartmouth Medical School, Hanover, NH 03755-3844, USA.

Abstract

The homotypic fusion of yeast vacuoles, each with 3Q- and 1R-SNARE, requires SNARE chaperones (Sec17p/Sec18p and HOPS) and regulatory lipids (sterol, diacylglycerol and phosphoinositides). Pairs of liposomes of phosphatidylcholine/phosphatidylserine, bearing three vacuolar Q-SNAREs on one and the R-SNARE on the other, undergo slow lipid mixing, but this is unaffected by HOPS and inhibited by Sec17p/Sec18p. To study these essential fusion components, we reconstituted proteoliposomes of a more physiological composition, bearing vacuolar lipids and all four vacuolar SNAREs. Their fusion requires Sec17p/Sec18p and HOPS, and each regulatory lipid is important for rapid fusion. Although SNAREs can cause both fusion and lysis, fusion of these proteoliposomes with Sec17p/Sec18p and HOPS is not accompanied by lysis. Sec17p/Sec18p, which disassemble SNARE complexes, and HOPS, which promotes and proofreads SNARE assembly, act synergistically to form fusion-competent SNARE complexes, and this synergy requires phosphoinositides. This is the first chemically defined model of the physiological interactions of these conserved fusion catalysts.

PMID:
18650938
PMCID:
PMC2516887
DOI:
10.1038/emboj.2008.139
[Indexed for MEDLINE]
Free PMC Article

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