Cloning and functional characterization of inwardly rectifying K^+ channels expressed in pancreatic beta-cells

胰腺β细胞中ATP敏感性钾通道（K_<ATP>channels）是调节葡萄糖诱导的胰岛素分泌的重要通道，也是磺酰脲类降糖药的作用靶点，而磺酰脲类降糖药广泛应用于非胰岛素依赖型糖尿病（NIDDM）的治疗，但K_ channels的分子基础<ATP>尚不清楚。磺酰脲类药物受体（SUR 1）是ATP结合盒（ABC）超家族的一员，近年来被分离出来，但它不能引起钾<ATP>通道电流。我们克隆了内向整流钾离子通道家族的一个成员uK_<ATP>-1（Kir6.1），它在包括胰岛在内的多种组织中表达，但在克隆的β细胞系中不表达。随后，我们还克隆了BIR（Kir6.2），其与Kir6.1属于相同的亚家族，并且主要在胰岛和克隆β细胞系中表达。我们已经证明，<ATP>由SUR 1和Kir6.2重建的K_通道的性质与天然胰腺β细胞的性质相同，表明胰腺β细胞K_<ATP>通道是由Kir6.2和SUR 1组成的复合物。我们还分离了一个新的磺酰脲受体（SUR 2），它与SUR 1有68%的氨基酸同源性。SUR 2 mRNA在心脏和骨骼肌中含量丰富。SUR 2和Kir6.2在COS 1细胞中的共表达重建了<ATP>天然心肌和骨骼肌中的K_通道，表明K_通道的药理学特性<ATP>是由磺酰脲类受体亚基家族决定的。我们还确定了人Kir6.2基因的结构和染色体定位，并表明人SUR 1和Kir6.2基因聚集在染色体11p15.1。我们发现了Kir6.2基因的几种多态性和接近Kir6.2基因的微卫星DNA多态性，并表明Kir6.2基因的遗传变异在日本人和高加索人对NIDDM的易感性中并不起主要作用。

In pancreatic beta-cells, ATP-sensitive potassium channels (K_<ATP> channels) are crucial for the regulation of glucoseinduced insulin secretion and are the target for the sulfonylureas, oral hypoglycemic agents widely used in the treatment of non-insulin-dependent diabetes mellitus (NIDDM), but the molecular basis of the K_<ATP> channels was unknown. Recently, a high affinity receptor for sulfonylureas (SUR1), a member of the ATP-binding cassette (ABC) superfamily, has been isolated, but it did not elicit K_<ATP> channel currents. We have cloned a member of the inward rectifier K^+ channel family, uK_<ATP>-1 (Kir6.1), which is expressed in various tissues including pancreatic islets, but not in clonal beta-cell lines. Subsequently, we also have cloned BIR (Kir6.2), which belongs to the same subfamily as Kir6.1 and is expressed predominantly in pancreatic islets and clonal beta-cell lines. We have shown that the properties of the K_<ATP> channel reconstituted from SUR1 and Kir6.2 are those described for native pancreatic beta-cells, demonstrating that the pancreatic beta-cell K_<ATP> channel is a complex composed of Kir6.2 and SUR1. We also have isolated a novel sulfonylurea receptor (SUR2) which shares 68% amino acisd identity with SUR1. SUR2 mRNA is abundant in heart and skeletal muscle. Coexpression of SUR2 and Kir6.2 in COS1 cells reconstitutes K_<ATP> channels found in native cardiac and skeletal muscle, indicating that the pharmacological properties of K_<ATP> channels are determined by a family of sulfonylurea receptor subunits. We also have determined the structure and chromosomal localization of human Kir6.2 gene, and have shown that human SUR1 and Kir6.2 genes are clustered at chromosome 11p15.1. We have found several polymorphysms in the Kir6.2 gene and microsatellite DNA polymorphysms close to the Kir6.2 gene, and have shown that genetic variation in the Kir6.2 gene does not play a major role in susceptibility to NIDDM in Japanese and Caucasian.