Distinct protein domains are responsible for the interaction of Hrs-2 withSNAP-25 - The role of Hrs-2 in 7 S complex formation

Regulated secretion of neurotransmitter at the synapse is likely to be medi ated by dynamic protein interactions involving components of the vesicle (v esicle-associated membrane protein; VAMP) and plasma membrane (syntaxin and synaptosomal associated protein of 25 kDa (SNAP-25)) along with additional molecules that allow for the regulation of this process. Recombinant Hrs-2 interacts with SNAP-25 in a calcium-dependent manner (they dissociate at e levated calcium levels) and inhibits neurotransmitter release. Thus, Hrs-a has been hypothesized to serve a negative regulatory role in secretion thro ugh its interaction with SNAP-25. In this report, we show that Hrs-2 and SN AP-25 interact directly through specific coiled-coil domains in each protei n. The presence of syntaxin enhances the binding of Hrs-2 to SNAP-25. Moreo ver, while both Hrs-2 and VAMP can separately bind to SNAP-25, they cannot bind simultaneously. Additionally, the presence of Hrs-2 reduces the incorp oration of VAMP into the syntaxin SNAP-25.VAMP (7 S) complex. These finding s suggest that Hrs-2 may modulate exocytosis by regulating the assembly of a protein complex implicated in membrane fusion.