A novel dual mechanism controlling the localization and function of exocytic v-SNAREs

SNARE proteins are essential for membrane fusion but their regulation is not yet fully understood. We have previously shown that the amino-terminal Longin domain [Filippini, 2001 #8431] of the v-SNARE TI-VAMP/VAMP7 plays an inhibitory role in neurite outgrowth [Martinez-Arca, 2001 #8432; Martinez-Arca, 2000 #828]. The goal of this study was to investigate the regulation of TI-VAMP as a model of v-SNARE regulation. We show here that the Longin domain of TI-VAMP plays a dual role. Firstly, it negatively regulates the ability of TI-VAMP and the Longin/Synaptobrevin2 chimera to participate in SNARE complexes. Secondly, it interacts with the AP-3 adaptor complex and this interaction targets TI-VAMP to late endosomes. Accordingly, in mocha cells lacking AP-3d, TI-VAMP is retained in an early endosomal compartment. Furthermore, TI-VAMPc, a novel isoform of TI-VAMP lacking a one third of the Longin domain, does not interact with AP-3 and therefore it is not targeted to late endosomes, but this shorter Longin domain yet inhibits SNARE complex formation in vivo. These findings support a novel mechanism controlling both localization and function of exocytic v-SNAREs through the Longin domain and clathrin adaptors. They point to the amino-terminal domains of SNAREs as multifunctional modules responsible for the fine-tuning of SNARE function.