1. Friedrich-Miescher-Laboratorium der Max-Planck-Gesellschaft, Spemannstrasse 37-39, 72076 Tübingen, Germany
2. Department of Molecular Biology, University of Southern Denmark, Campusvej 55, 5230 Odense M, Denmark
3. These authors contributed equally to this work

Correspondence to: Andreas Mayer¹ Correspondence and requests for materials should be addressed to A.M. (e-mail: Email: andreas.mayer@tuebingen.mpg.de).

Abstract

SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) and Rab-GTPases, together with their cofactors, mediate the attachment step in the membrane fusion of vesicles. But how bilayer mixing—the subsequent core process of fusion—is catalysed remains unclear. Ca²⁺/calmodulin controls this terminal process in many intracellular fusion events. Here we identify V0, the membrane-integral sector of the vacuolar H⁺-ATPase, as a target of calmodulin on yeast vacuoles. Between docking and bilayer fusion, V0 sectors from opposing membranes form complexes. V0 trans-complex formation occurs downstream from trans-SNARE pairing, and depends on both the Rab-GTPase Ypt7 and calmodulin. The maintenance of existing complexes and completion of fusion are independent of trans-SNARE pairs. Reconstituted proteolipids form sealed channels, which can expand to form aqueous pores in a Ca²⁺/calmodulin-dependent fashion. V0 trans-complexes may therefore form a continuous, proteolipid-lined channel at the fusion site. We propose that radial expansion of such a protein pore may be a mechanism for intracellular membrane fusion.