Lin Y F, Jan Y N, Jan L Y
Howard Hughes Medical Institute, Departments of Physiology and Biochemistry, University of California, San Francisco, CA 94143-0725, USA.
EMBO J. 2000 Mar 1;19(5):942-55. doi: 10.1093/emboj/19.5.942.
ATP-sensitive potassium (K(ATP)) channels regulate insulin secretion, vascular tone, heart rate and neuronal excitability by responding to transmitters as well as the internal metabolic state. K(ATP) channels are composed of four pore-forming alpha-subunits (Kir6.2) and four regulatory beta-subunits, the sulfonylurea receptor (SUR1, SUR2A or SUR2B). Whereas protein kinase A (PKA) phosphorylation of serine 372 of Kir6.2 has been shown biochemically by others, we found that the phosphorylation of T224 rather than S372 of Kir6.2 underlies the catalytic subunits of PKA (c-PKA)- and the D1 dopamine receptor-mediated stimulation of K(ATP) channels expressed in HEK293 cells. Specific changes in the kinetic properties of channels treated with c-PKA, as revealed by single-channel analysis, were mimicked by aspartate substitution of T224. The T224D mutation also reduced the sensitivity to ATP inhibition. Alteration of channel gating and a decrease in the apparent affinity for ATP inhibition thus underlie the positive regulation of K(ATP) channels by PKA phosphorylation of T224 in Kir6.2, which may represent a general mechanism for K(ATP) channel regulation in different tissues.
ATP敏感性钾（K(ATP)）通道通过对递质以及细胞内代谢状态做出反应来调节胰岛素分泌、血管张力、心率和神经元兴奋性。K(ATP)通道由四个形成孔道的α亚基（Kir6.2）和四个调节性β亚基即磺脲类受体（SUR1、SUR2A或SUR2B）组成。虽然其他人已通过生化方法证明了Kir6.2的丝氨酸372的蛋白激酶A（PKA）磷酸化，但我们发现，在HEK293细胞中表达的K(ATP)通道，其受PKA催化亚基（c-PKA）和D1多巴胺受体介导的刺激作用的基础是Kir6.2的苏氨酸224而非丝氨酸372的磷酸化。单通道分析显示，用c-PKA处理的通道动力学特性的特定变化可被苏氨酸224的天冬氨酸替代所模拟。T224D突变也降低了对ATP抑制的敏感性。因此，通道门控的改变以及对ATP抑制的表观亲和力的降低是Kir6.2中苏氨酸224的PKA磷酸化对K(ATP)通道进行正向调节的基础，这可能代表了不同组织中K(ATP)通道调节的一般机制。
