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

在胰腺β细胞中，ATP敏感钾通道（K_<ATP>通道）对葡萄糖诱导的胰岛素分泌的调节至关重要，并且是磺脲类药物的靶点，口服降糖药广泛用于治疗非胰岛素依赖型糖尿病（NIDDM），但K_<ATP>通道的分子基础尚不清楚。最近，一个高亲和受体sulfonylureas (SUR1)， ATP结合盒（ABC）超家族的成员，已被分离出来，但它不引起K_<ATP>通道电流。我们克隆了一个内向整流K^+通道家族的成员，uK_<ATP>-1 (Kir6.1)，它在包括胰岛在内的各种组织中表达，但在克隆β细胞系中不表达。随后，我们还克隆了BIR (Kir6.2)，它与Kir6.1属于同一亚家族，主要在胰岛和克隆β细胞系中表达。我们已经证明，由SUR1和Kir6.2重建的K_<ATP>通道的性质与天然胰腺β细胞的性质相同，这表明胰腺β细胞的K_<ATP>通道是由Kir6.2和SUR1组成的复合物。我们还分离出一种新的磺酰脲受体（SUR2），它与SUR1具有68%的氨基酸同源性。sur2mrna在心脏和骨骼肌中含量丰富。在COS1细胞中，SUR2和Kir6.2的共表达重建了天然心脏和骨骼肌中发现的K_<ATP>通道，表明K_<ATP>通道的药理特性是由磺酰脲受体亚基家族决定的。我们还确定了人类Kir6.2基因的结构和染色体定位，并表明人类SUR1和Kir6.2基因聚集在染色体11p15.1。我们发现了Kir6.2基因的几个多态性和Kir6.2基因附近的微卫星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.