Structure and function of mitochondrial ATP-sensitive K^+ channel.

ATP敏感的K~+(K_&lt；ATP&gt；)通道的开启和关闭取决于细胞质中的ATP浓度，并控制细胞的兴奋性。我们先前已经证明，胰腺β细胞的K_1通道由ABC蛋白的成员SUR1和内向整流性K通道的成员Kir6.2组成。在心肌、骨骼肌和血管中，具有相似特性的K&lt；ATP&gt；通道包括SUR2A/B和Kir6.x，它们调节肌肉在缺氧或缺血时的收缩。尽管线粒体表达类似的K&lt；ATP&gt；通道，并调节能量代谢，但其组成分子尚不清楚。我们利用膜片钳技术，记录了Jurkat细胞线粒体内膜的离子通道活动。观察到电导为55ps的K~(++)离子选择性通道，其活性可被2 mM的ATP抑制，这些特性和猝发动力学与含有Kir6.1分子的通道相似。在另一项研究中，是否在低…期间诱导了Kir6.1分子更多的血管来源的XIA、A7r5细胞在含有Co~(2+)和Gt~(2+)的培养液中培养，模拟低氧条件。用抗Kir6.1抗体进行免疫印迹显示，在加入这些离子后3-4天，Kir6.1蛋白开始表达。但在血管肌质膜上未观察到具有Kir6.1活性的通道活动。在这些结果的基础上，目前正在研究缺血诱导的Kir6.1分子组成线粒体K_lt；ATP&gt；通道的可能性。在其他一系列实验中，我们研究了蛋白激酶A介导的由SUR1和Kir6.2组成的K_&lt；ATP&gt；通道的磷酸化作用。最后，研究了抗糖尿病药物曲格列酮、吡格列酮和KAD-1229对重组K_&lt；ATP&gt；通道的影响。这些研究有助于了解质膜和线粒体中K&lt；ATP&Gt；通道的结构与功能关系。较少

ATP-sensitive K^+ (K_<ATP>) channels open and close depending on ATP concentration in the cytoplasm, and control excitability of the cells. We have previously shown that K_<ATP> channel of pancreatic β-cell comprises SUR1, a member of ABC protein, and Kir6.2, a member of inward rectifier K channels. In heart and skeletal muscles, and blood vessels, K_<ATP> channels of similar properties comprise SUR2A/B and Kir6.x, regulating muscle contraction during hypoxia or ischemia.Although similar K_<ATP> channel is expressed in mitochondria, and regulating energy metabolism, its constituting molecules are unknown.Using the patch-clamp technique, we recorded ion channel activity from the mitochondrial inner membrane of Jurkat cells. K^+ ion selective channels with conductance of 55 pS were observed, of which activity being suppressed by 2 mM ATP.These properties and burst kinetics are similar to the channel comprising Kir6.1 molecule. In other to study if Kir6.1 molecules are induced during hypo … More xia, A7r5 cells, derived from blood vessel were cultured in medium containing Co^<2+> ions, mimicking hypoxic conditions. Western blotting using anti Kir6.1 antibody revealed that Kir6.1 protein became expressed in 3-4 days after an addition of the ions. However, no activity of channels comprising activity Kir6.1 was observed in the plasma membrane of heart of blood vessel muscles. Following these results, a possibility that Kir6.1 molecule induced in ischemia composes mitochondrial K_<ATP> channel is currently investigated.In other series of experiments effects of protein kinase A mediated phosphorylation of K_<ATP> channels comprising SUR1 and Kir6.2 were studied. We found both subunits are phosphorylated and channel properties are modulated in a different manner.Finally, effects of anti-diabetic reagents, troglitazone, pioglitazone, and KAD-1229, were studied on reconstituted K_<ATP> channels. The studies are helpful in understanding structure-function relationship of K_<ATP> channels in the plasma membrane as well as mitochondria. Less