Meng Jianghui, Dolly J Oliver, Wang Jiafu
International Centre for Neurotherapeutics, Dublin City University, Glasnevin, Dublin 9, Ireland.
Mol Neurobiol. 2014 Oct;50(2):574-88. doi: 10.1007/s12035-014-8665-1. Epub 2014 Mar 8.
Oligomerisation of soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complexes is required for synaptic vesicle fusion and neurotransmitter release. How these regulate the release of pain peptides elicited by different stimuli from sensory neurons has not been established. Herein, K(+) depolarization was found to induce multiple sodium dodecyl sulfate (SDS)-resistant SNARE complexes in sensory neurons exposed to botulinum neurotoxins (BoNTs), with molecular weights ranging from 104-288 k (large) to 38-104 k (small). Isoform 1 of vesicle-associated membrane protein 1 (VAMP 1) assembled into stable complexes upon depolarisation and was required for the participation of intact synaptosome-associated protein of relative molecular mass 25 k (SNAP-25) or BoNT/A-truncated form (SNAP-25A) in the large functional and small inactive SDS-resistant SNARE complexes. Cleaving VAMP 1 decreased SNAP-25A in the functional complexes to a much greater extent than the remaining intact SNAP-25. Syntaxin 1 proved essential for the incorporation of intact and SNAP-25A into the large complexes. Truncation of syntaxin 1 by BoNT/C1 caused /A- and/or /C1-truncated SNAP-25 to appear in non-functional complexes and blocked the release of calcitonin gene-related peptide (CGRP) elicited by capsaicin, ionomycin, thapsigargin or K(+) depolarization. Only the latter two were susceptible to /A. Inhibition of CGRP release by BoNT/A was reversed by capsaicin and/or ionomycin, an effect overcome by BoNT/C1. Unlike BoNT/B, BoNT/D cleaved VAMP 1 in addition to 2 and 3 in rat sensory neurons and blocked both CGRP and substance P release. Thus, unlike SNAP-25, syntaxin 1 and VAMP 1 are more suitable targets to abolish functional SNARE complexes and pain peptide release evoked by any stimuli.
可溶性N - 乙基马来酰亚胺敏感因子附着蛋白受体（SNARE）复合体的寡聚化是突触小泡融合和神经递质释放所必需的。这些过程如何调节感觉神经元对不同刺激产生的痛觉肽释放尚未明确。在此，发现钾离子去极化可在暴露于肉毒杆菌神经毒素（BoNTs）的感觉神经元中诱导多种耐十二烷基硫酸钠（SDS）的SNARE复合体，其分子量范围从104 - 288k（大）到38 - 104k（小）。囊泡相关膜蛋白1（VAMP 1）的同工型1在去极化时组装成稳定复合体，并且是完整的相对分子质量25k的突触体相关蛋白（SNAP - 25）或BoNT/A截短形式（SNAP - 25A）参与大的功能性和小的无活性耐SDS的SNARE复合体所必需的。切割VAMP 1使功能性复合体中的SNAP - 25A减少的程度比剩余完整的SNAP - 25大得多。Syntaxin 1被证明是完整的和SNAP - 25A纳入大复合体所必需的。BoNT/C1对Syntaxin 1的截短导致/A - 和/或/C1 - 截短的SNAP - 25出现在无功能复合体中，并阻断了辣椒素、离子霉素、毒胡萝卜素或钾离子去极化所引发的降钙素基因相关肽（CGRP）的释放。只有后两者对/A敏感。BoNT/A对CGRP释放的抑制作用可被辣椒素和/或离子霉素逆转，而BoNT/C1可克服这种作用。与BoNT/B不同，BoNT/D除了切割大鼠感觉神经元中的VAMP 2和VAMP 3外，还切割VAMP 1，并阻断CGRP和P物质的释放。因此，与SNAP - 25不同，Syntaxin 1和VAMP 1是消除功能性SNARE复合体以及任何刺激引发的痛觉肽释放的更合适靶点。
