Sequential SNARE disassembly and GATE-16-GOS-28 complex assembly mediated by distinct NSF activities drives Golgi membrane fusion

doi:10.1083/jcb.200202082

Published online 17 June 2002. doi:10.1083/jcb.200202082

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© The Rockefeller University Press, 0021-9525/2002/6/1161 $5.00
The Journal of Cell Biology, Volume 157, Number 7, June 24, 2002 1161-1173

Article

Sequential SNARE disassembly and GATE-16–GOS-28 complex assembly mediated by distinct NSF activities drives Golgi membrane fusion

Joyce M.M. Müller¹, James Shorter², Richard Newman³, Katrin Deinhardt¹, Yuval Sagiv⁴, Zvulun Elazar⁴, Graham Warren² and David T. Shima¹

¹ Endothelial Cell Biology, Cancer Research UK, London WC2A 3PX, United Kingdom
² Department of Cell Biology, Ludwig Institute for Cancer Research, Yale University School of Medicine, New Haven, CT 06520
³ Macromolecular Structure Database Group, European Bioinformatics Institute, Cambridge CB10 1SD, United Kingdom
⁴ Department of Biological Chemistry, The Weizmann Institute of Science, Rehovot, 76100 Israel

Address correspondence to David T. Shima, Endothelial Cell Biology, Cancer Research UK, 44 Lincoln's Inn Field, London WC2A 3PX, UK. Tel.: 44-203-785-4301. Fax: 44-207-269-3417. E-mail: shima{at}cancer.org.uk

Characterization of mammalian NSF (G274E) and Drosophila NSF(comatose) mutants revealed an evolutionarily conserved NSFactivity distinct from ATPase-dependent SNARE disassembly thatwas essential for Golgi membrane fusion. Analysis of mammalianNSF function during cell-free assembly of Golgi cisternae frommitotic Golgi fragments revealed that NSF disassembles GolgiSNAREs during mitotic Golgi fragmentation. A subsequent ATPase-independentNSF activity restricted to the reassembly phase is essentialfor membrane fusion. NSF/ ${alpha}$ -SNAP catalyze the binding of GATE-16to GOS-28, a Golgi v-SNARE, in a manner that requires ATP butnot ATP hydrolysis. GATE-16 is essential for NSF-driven Golgireassembly and precludes GOS-28 from binding to its cognatet-SNARE, syntaxin-5. We suggest that this occurs at the inceptionof Golgi reassembly to protect the v-SNARE and regulate SNAREfunction.

Key Words: NSF; ${alpha}$ -SNAP; GATE-16; GOS-28; Golgi

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