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 relative abundance during PC12 cell differentiation induced by nerve growth factor (NGF). Norepinephrine release by NGF-differentiated PC12 cells is more sensitive to type A botulinum toxin (BoNT/A) than by nondifferentiated cells, while both differentiated and nondifferentiated PC12 cells are equally sensitive to type E botulinum toxin (BoNT/E). The differential sensitivity to BoNT/A corresponds to an altered susceptibility of SNAP-25 isotypes to BoNT/A cleavage in vitro, whereas both isotypes are equally vulnerable to cleavage by BoNT/E. Using recombinant SNAP-25 preparations, we show that BoNT/A cleaves SNAP-25b (present in differentiated cells) 2-fold more readily than SNAP-25a (present in both differentiated and nondifferentiated cells). Structural studies using far-ultraviolet circular dichroism (UV--CD) and thermal denaturation suggest a difference in the polypeptide folding as the underlying 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.
Puffer
E B
EB
Department of Chemistry and Biochemistry University of Massachusetts at Dartmouth, Dartmouth, Massachusetts 02747, USA.
Lomneth
R B
RB
Sarkar
H K
HK
Singh
B R
BR
D000818:Animals; D001905:Botulinum Toxins / Q000494:pharmacology; D019274:Botulinum Toxins, Type A / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects; D002942:Circular Dichroism; D008565:Membrane Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000502:physiology*; D020932:Nerve Growth Factor / Q000494:pharmacology; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000502:physiology*; D009638:Norepinephrine / Q000037:antagonists & inhibitors / Q000378:metabolism*; D016716:PC12 Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D020033:Protein Isoforms / Q000096:biosynthesis / Q000737:chemistry / Q000502:physiology; D051381:Rats; D050825:Synaptosomal-Associated Protein 25; D013816:Thermodynamics
D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D020033:Protein Isoforms; C494210:Snap25 protein, rat; D050825:Synaptosomal-Associated Protein 25; D020932:Nerve Growth Factor; D001905:Botulinum Toxins; D019274:Botulinum Toxins, Type A; C096326:botulinum toxin type E; D009638:Norepinephrine