HOPS recognizes each SNARE, assembling ternary trans-complexes for sudden fusion upon engagement with the 4th SNARE

Abstract

ABSTRACTVacuole fusion requires SNAREs, Sec17/18, a Rab, and HOPS. We find that co-incubation of HOPS, proteoliposomes bearing the Rab and R-SNARE, and proteoliposomes with the Rab and any two Q-SNAREs yields a trans complex which includes these 3 SNAREs. The missing Q-SNARE then triggers a burst of fusion, indicating that each HOPS, R-, and QxQy-SNARE trans-complex is an activated intermediate for functional Qz-SNARE incorporation. HOPS can assemble activated fusion intermediates because it recognizes each of the four SNAREs, binding them independently. HOPS-dependent fusion is saturable for each Q-SNARE, indicating saturable functional sites on HOPS. Though a nonspecific tether allows fusion with pre-assembled Q-SNAREs, only HOPS catalyzes fusion when the Q-SNAREs are not pre-assembled by ushering each Q-SNARE into a functional complex. In contrast, there is little spontaneous functional assembly of the 3 Q-SNAREs. HOPS thus recognizes each of the 4 SNAREs to assemble a versatile set of activated fusion intermediates.