Differential roles of developmentally distinct SNAP-25 isoforms in the neurotransmitter release process

The role of SNAP-25 (synaptosomal associated protein of 25 kDa) isotypes in the neurotransmitter release process was examined by varying their relativ e abundance during PC12 cell differentiation induced by nerve growth factor (NGF). Norepinephrine release by NGF-differentiated PC12 cells is more sen sitive to type A botulinum toxin (BoNT/A) than by nondifferentiated cells, while both differentiated and nondifferentiated PC12 cells are equally sens itive to type E botulinum toxin (BoNT/E). The differential sensitivity to B oNT/A corresponds to an altered susceptibility of SNAP-25 isotypes to BoNT/ A cleavage in vitro, whereas both isotypes are equally vulnerable to cleava ge by BoNT/E. Using recombinant SNAP-25 preparations, we show that BoNT/A c leaves SNAP-25b (present in differentiated cells) 2-fold more readily than SNAP-25a (present in both differentiated and nondifferentiated cells). Stru ctural studies using far-ultraviolet circular dichroism (UV-CD) and thermal denaturation suggest a difference in the polypeptide folding as the underl ying molecular basis for the differential sensitivity of SNAP-25b and SNAP- 25a to BoNT/A cleavage. We propose differential roles for SNAP-25b and SNAP 25a in the neurotransmitter release process since our results suggest that BoNT/A inhibits neurotransmitter release by primarily cleaving SNAP-25b.