The C terminus of SNAP25 is essential for Ca2+-dependent binding of synaptotagmin to SNARE complexes.

The plasma membrane soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins syntaxin and synaptosome-associated protein of 25 kDa (SNAP25) and the vesicle SNARE protein vesicleassociated membrane protein (VAMP) are essential for a late Ca21-dependent step in regulated exocytosis, but their precise roles and regulation by Ca21 are poorly understood. Botulinum neurotoxin (BoNT) E, a protease that cleaves SNAP25 at Arg180-Ile181, completely inhibits this late step in PC12 cell membranes, whereas BoNT A, which cleaves SNAP25 at Gln197-Arg198, is only partially inhibitory. The difference in toxin effectiveness was found to result from a reversal of BoNT A but not BoNT E inhibition by elevated Ca21 concentrations. BoNT A treatment essentially increased the Ca21 concentration required to activate exocytosis, which suggested a role for the C terminus of SNAP25 in the Ca21 regulation of exocytosis. Synaptotagmin, a proposed Ca21 sensor for exocytosis, was found to bind SNAP25 in a Ca21-stimulated manner. Ca21-dependent binding was abolished by BoNT E treatment, whereas BoNT A treatment increased the Ca21 concentration required for binding. The C terminus of SNAP25 was also essential for Ca21- dependent synaptotagmin binding to SNAP25zsyntaxin and SNAP25zsyntaxinzVAMP SNARE complexes. These results clarify classical observations on the Ca21 reversal of BoNT A inhibition of neurosecretion, and they suggest that an essential role for the C terminus of SNAP25 in regulated exocytosis is to mediate Ca21-dependent interactions between synaptotagmin and SNARE protein complexes.

Main Author: Gerona, Roy.
Other Authors: Larsen, Eric., Kowalchyk, Judith., Martin, Thomas.
Language: English
Published: 1999
Online Access: http://ezproxy.villanova.edu/login?url=https://digital.library.villanova.edu/Item/vudl:178027