Xia Fuzhen, Gao Xiaodong, Kwan Edwin, Lam Patrick P L, Chan Lillian, Sy Keiyan, Sheu Laura, Wheeler Michael B, Gaisano Herbert Y, Tsushima Robert G
Departments of Medicine and Physiology, University of Toronto, Toronto, Ontario M5S 1A8,Canada.
J Biol Chem. 2004 Jun 4;279(23):24685-91. doi: 10.1074/jbc.M314314200. Epub 2004 Apr 8.
In pancreatic beta-cells, the predominant voltage-gated Ca(2+) channel (Ca(V)1.2) and K(+) channel (K(V)2.1) are directly coupled to SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor) proteins. These SNARE proteins modulate channel expression and gating and closely associate these channels with the insulin secretory vesicles. We show that K(V)2.1 and Ca(V)1.2, but not K(V)1.4, SUR1, or Kir6.2, target to specialized cholesterol-rich lipid raft domains on beta-cell plasma membranes. Similarly, the SNARE proteins syntaxin 1A, SNAP-25, and VAMP-2, but not Munc-13-1 or n-Sec1, are associated with lipid rafts. Disruption of the lipid rafts by depleting membrane cholesterol with methyl-beta-cyclodextrin shunts K(V)2.1, Ca(V)1.2, and SNARE proteins out of lipid rafts. Furthermore, methyl-beta-cyclodextrin inhibits K(V)2.1 but not Ca(V)1.2 channel activity and enhances single-cell exocytic events and insulin secretion. Membrane compartmentalization of ion channels and SNARE proteins in lipid rafts may be critical for the temporal and spatial coordination of insulin release, forming what has been described as the excitosome complex.
在胰腺β细胞中，主要的电压门控Ca(2+)通道（Ca(V)1.2）和K(+)通道（K(V)2.1）直接与SNARE（可溶性N - 乙基马来酰亚胺敏感因子附着蛋白（SNAP）受体）蛋白偶联。这些SNARE蛋白调节通道表达和门控，并使这些通道与胰岛素分泌囊泡紧密相连。我们发现K(V)2.1和Ca(V)1.2，而非K(V)1.4、SUR1或Kir6.2，定位于β细胞质膜上富含胆固醇的特殊脂筏结构域。同样，SNARE蛋白 syntaxin 1A、SNAP - 25和VAMP - 2，而非Munc - 13 - 1或n - Sec1，与脂筏相关。用甲基 - β - 环糊精耗尽膜胆固醇来破坏脂筏，会使K(V)2.1、Ca(V)1.2和SNARE蛋白从脂筏中分流出来。此外，甲基 - β - 环糊精抑制K(V)2.1通道活性但不抑制Ca(V)1.2通道活性，并增强单细胞胞吐事件和胰岛素分泌。脂筏中离子通道和SNARE蛋白的膜区室化对于胰岛素释放的时间和空间协调可能至关重要，形成了所谓的兴奋小体复合物。
