Xia Fuzhen, Leung Yuk M, Gaisano Gregory, Gao Xiaodong, Chen Yi, Fox Jocelyn E Manning, Bhattacharjee Alpana, Wheeler Michael B, Gaisano Herbert Y, Tsushima Robert G
Department of Medicine, University of Toronto, Ontario, Canada M5S 1A8.
Endocrinology. 2007 May;148(5):2157-67. doi: 10.1210/en.2006-1296. Epub 2007 Feb 15.
Pancreatic alpha-cells secrete glucagon in response to low glucose to counter insulin actions, thereby maintaining glucose homeostasis. The molecular basis of alpha-cell stimulus-secretion coupling has not been fully elucidated. We investigated the expression of voltage-gated K(+) (K(V)) and Ca(2+) (Ca(V)) channels, and soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins in pancreatic alpha-cells and examined their targeting to specialized cholesterol-rich lipid rafts. In alpha-cells, we detected the expression of K(V)4.1/4.3 (A-type current), K(V)3.2/3.3 (delayed rectifier current), Ca(V)1.2 (L-type current), Ca(V)2.2 (N-type current), and the SNARE (synaptosomal-associated protein of 25 kDa, syntaxin 1A, and vesicle-associated membrane protein 2) and SNARE-associated proteins (Munc-13-1 and Munc-18a). We also detected caveolin-2, a structural protein of cholesterol-rich lipid rafts. Of these proteins, caveolin-2, K(V)4.1/4.3, Ca(V)1.2, and SNARE proteins (syntaxin 1A, synaptosomal-associated protein of 25 kDa, and vesicle-associated membrane protein 2) target to lipid raft domains on alpha-cell plasma membranes. Disruption of lipid rafts by depletion of membrane cholesterol with methyl-beta-cyclodextrin decreased the association of K(V)4.1/4.3, Ca(V)1.2, and SNARE proteins with lipid rafts. This resulted in inhibition of A-type K(V) currents and enhancement of glucagon secretion from alpha-cells. Consistently, capacitance measurements of exocytosis of single alpha-cells showed enhanced exocytosis after membrane cholesterol depletion. Taken together, our results demonstrate the association of K(V)4, Ca(V)1.2, and SNARE proteins with lipid rafts in pancreatic alpha-cells. Glucagon secretion from alpha-cells is regulated by lipid rafts, and the dissociation of SNARE proteins from cholesterol-rich lipid raft domains enhances glucagon secretion.
胰腺α细胞在血糖水平低时分泌胰高血糖素，以对抗胰岛素的作用，从而维持血糖稳态。α细胞刺激-分泌偶联的分子基础尚未完全阐明。我们研究了电压门控钾离子（K(V)）通道、钙离子（Ca(V)）通道以及可溶性N-乙基马来酰亚胺敏感因子附着蛋白受体（SNARE）蛋白在胰腺α细胞中的表达，并检测了它们在富含胆固醇的特殊脂筏中的定位。在α细胞中，我们检测到K(V)4.1/4.3（A型电流）、K(V)3.2/3.3（延迟整流电流）、Ca(V)1.2（L型电流）、Ca(V)2.2（N型电流）以及SNARE（25 kDa突触体相关蛋白、 syntaxin 1A和囊泡相关膜蛋白2）和SNARE相关蛋白（Munc-13-1和Munc-18a）的表达。我们还检测到富含胆固醇脂筏的结构蛋白小窝蛋白-2。在这些蛋白中，小窝蛋白-2、K(V)4.1/4.3、Ca(V)1.2和SNARE蛋白（syntaxin A、25 kDa突触体相关蛋白和囊泡相关膜蛋白2）定位于α细胞质膜上的脂筏结构域。用甲基-β-环糊精耗尽膜胆固醇破坏脂筏，会降低K(V)4.1/4.3、Ca(V)1.2和SNARE蛋白与脂筏的结合。这导致A型K(V)电流受到抑制，α细胞分泌的胰高血糖素增加。同样，对单个α细胞胞吐作用的电容测量显示，膜胆固醇耗尽后胞吐作用增强。综上所述，我们的结果表明K(V)4、Ca(V)1.2和SNARE蛋白在胰腺α细胞中与脂筏相关。α细胞分泌胰高血糖素受脂筏调节，SNARE蛋白从富含胆固醇的脂筏结构域解离会增强胰高血糖素的分泌。
