Sadoul K, Lang J, Montecucco C, Weller U, Regazzi R, Catsicas S, Wollheim C B, Halban P A
Laboratoires de Recherche Louis Jeantet, Centre Médical Universitaire, Geneva, Switzerland.
J Cell Biol. 1995 Mar;128(6):1019-28. doi: 10.1083/jcb.128.6.1019.
SNAP-25 is known as a neuron specific molecule involved in the fusion of small synaptic vesicles with the presynaptic plasma membrane. By immunolocalization and Western blot analysis, it is now shown that SNAP-25 is also expressed in pancreatic endocrine cells. Botulinum neurotoxins (BoNT) A and E were used to study the role of SNAP-25 in insulin secretion. These neurotoxins inhibit transmitter release by cleaving SNAP-25 in neurons. Cells from a pancreatic B cell line (HIT) and primary rat islet cells were permeabilized with streptolysin-O to allow toxin entry. SNAP-25 was cleaved by BoNT/A and BoNT/E, resulting in a molecular mass shift of approximately 1 and 3 kD, respectively. Cleavage was accompanied by an inhibition of Ca(++)-stimulated insulin release in both cell types. In HIT cells, a concentration of 30-40 nM BoNT/E gave maximal inhibition of stimulated insulin secretion of approximately 60%, coinciding with essentially complete cleavage of SNAP-25. Half maximal effects in terms of cleavage and inhibition of insulin release were obtained at a concentration of 5-10 nM. The A type toxin showed maximal and half-maximal effects at concentrations of 4 and 2 nM, respectively. In conclusion, the results suggest a role for SNAP-25 in fusion of dense core secretory granules with the plasma membrane in an endocrine cell type- the pancreatic B cell.
SNAP-25是一种神经元特异性分子，参与小突触囊泡与突触前质膜的融合。通过免疫定位和蛋白质印迹分析，现已表明SNAP-25也在胰腺内分泌细胞中表达。肉毒杆菌神经毒素（BoNT）A和E被用于研究SNAP-25在胰岛素分泌中的作用。这些神经毒素通过切割神经元中的SNAP-25来抑制神经递质释放。用链球菌溶血素-O使胰腺β细胞系（HIT）的细胞和原代大鼠胰岛细胞通透，以允许毒素进入。SNAP-25被BoNT/A和BoNT/E切割，分别导致分子量大约偏移1 kD和3 kD。切割伴随着两种细胞类型中钙（++）刺激的胰岛素释放受到抑制。在HIT细胞中，30 - 40 nM的BoNT/E浓度对刺激的胰岛素分泌产生约60%的最大抑制，这与SNAP-25基本上完全被切割相吻合。在5 - 10 nM的浓度下获得了切割和胰岛素释放抑制方面的半数最大效应。A型毒素在4 nM和2 nM的浓度下分别显示出最大效应和半数最大效应。总之，结果表明SNAP-25在内分泌细胞类型——胰腺β细胞中致密核心分泌颗粒与质膜的融合中发挥作用。
