Structural and Functional Interactions between Transient Receptor Potential Vanilloid Subfamily 1 and Botulinum Neurotoxin Serotype A
Structural and Functional Interactions between Transient Receptor Potential Vanilloid Subfamily 1 and Botulinum Neurotoxin Serotype A
复制标题
瞬时受体电位香草酸亚家族 1 与肉毒杆菌神经毒素血清型 A 之间的结构和功能相互作用
DOI：
10.1371/journal.pone.0143024
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发表时间：
2016-01-08
期刊：
影响因子：
3.7
通讯作者：
Coffield, Julie A.
中科院分区：
文献类型：
--
作者：
Li, Xiaqing;Coffield, Julie A.
肉毒神经毒素是由肉毒梭菌产生的一种神经毒素。有八种血清学上不同的肉毒杆菌神经毒素亚型(血清型A-H型)。目前，A型肉毒神经毒素(BoNT/A)通常用于治疗许多疾病，如多动肌肉骨骼疾病、肌张力障碍和疼痛。然而，BONT/A的止痛效果以及介导BONT/A镇痛作用的机制尚不清楚。采用免疫荧光、免疫共沉淀法和免疫印迹法研究了原代培养的小鼠胚胎背根节神经元中BONT/A与瞬时受体电位香草样亚家族1(TRPV1)的相互作用。结果1)培养3周的小鼠背根神经节神经元高表达瞬时TRPV1、突触小泡2A(SV2A)和突触小体相关蛋白25(SNAP-25)。SV2A和SNAP-25分别是BONT/A的结合受体和靶蛋白。2)在背根神经节细胞培养中加入BONT/A后，TRPV1与BONT/A和断裂的SNAP-25共定位。3)免疫沉淀小球经BONT/A处理(1nmol/L)24小时后，免疫印迹检测到TRPV1和BONT/A阳性条带。4)用特异性抗体阻断TRPV1可减少BONT/A对SNAP-25的切割作用。结论BoNT/A在体外培养的小鼠胚胎背根神经节神经元中与TRPV1在结构和功能上存在相互作用。这些结果表明，BONT/A使用了另一种机制来调节疼痛缓解。
BackgroundBotulinum neurotoxins are produced by Clostridium botulinum bacteria. There are eight serologically distinct botulinum neurotoxin isoforms (serotypes A-H). Currently, botulinum neurotoxin serotype A (BoNT/A) is commonly used for the treatment of many disorders, such as hyperactive musculoskeletal disorders, dystonia, and pain. However, the effectiveness of BoNT/A for pain alleviation and the mechanisms that mediate the analgesic effects of BoNT/A remain unclear. To define the antinociceptive mechanisms by which BoNT/A functions, the interactions between BoNT/A and the transient receptor potential vanilloid subfamily 1 (TRPV1) were investigated using immunofluorescence, co-immunoprecipitation, and western blot analysis in primary mouse embryonic dorsal root ganglion neuronal cultures.Results1) Three-week-old cultured dorsal root ganglion neurons highly expressed transient TRPV1, synaptic vesicle 2A (SV2A) and synaptosomal-associated protein 25 (SNAP-25). SV2A and SNAP-25 are the binding receptor and target protein, respectively, of BoNT/A. 2) TRPV1 colocalized with both BoNT/A and cleaved SNAP-25 when BoNT/A was added to dorsal root ganglia neuronal cultures. 3) After 24 hours of BoNT/A treatment (1 nmol/l), both TRPV1 and BoNT/A positive bands were detected in western blots of immunoprecipitated pellets. 4) Blocking TRPV1 with a specific antibody decreased the cleavage of SNAP-25 by BoNT/A.ConclusionBoNT/A interacts with TRPV1 both structurally and functionally in cultured mouse embryonic dorsal root ganglion neurons. These results suggest that an alternative mechanism is used by BoNT/A to mediate pain relief.