MOLECULAR-PROPERTIES OF NEURONAL G-PROTEIN-ACTIVATED INWARDLY RECTIFYING K+ CHANNELS
MOLECULAR-PROPERTIES OF NEURONAL G-PROTEIN-ACTIVATED INWARDLY RECTIFYING K+ CHANNELS
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DOI：
10.1074/jbc.270.48.28660
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发表时间：
1995-12-01
影响因子：
4.8
通讯作者：
LAZDUNSKI, M
中科院分区：
文献类型：
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作者：
LESAGE, F;GUILLEMARE, E;LAZDUNSKI, M
最近已经克隆了四种编码cDNA的G激活的内向整流K+通道（Kubo，Y.，Eugeny，E.，斯莱辛格，宾夕法尼亚州，Jan，Y. N.，和Jan，L. Y.（1993）Nature 364，802-806; Lesage，F.，Duprat，F.，芬克，M.，Guillemare，E.，科波拉，T.，Lazdunski，M.，和Hugnot，J.P.（1994）FEES Lett. 353，37-42; Krapivinsky，G.，戈尔登，E. A.、威克曼，K.，Velimirovic，B.，克拉皮文斯基湖和Clapham，D. E.（1995）Nature 374，135-141）。我们报告了小鼠GIRK 2剪接变异体的克隆，标记为mGIRK 2A。两种通道蛋白在单独或与GIRK 1 cRNA组合注射其cRNA后在爪蟾卵母细胞中功能性表达。三个GIRK通道mGIRK 1 -3显示存在于大脑中。mGIRK 1和mGIRK 2在相同神经元中的共定位支持了天然通道由异聚亚基组装制成的假设。GIRK 3通道尚未成功表达，即使在其他类型的亚基存在下也是如此。然而，具有GIRK 2的氨基和羧基末端的GIRK 3嵌合体在GIRK 1存在下功能性表达。表达的mGIRK 2和mGIRK 1，-2电流被Ba ~（2+）和Cs ~（+）离子阻断。它们不受蛋白激酶A和蛋白激酶C的调节。通道活动在由内而外的切除斑块中下降，需要ATP来防止这种下降。由于不可水解的ATP类似物AMP-PCP也是活性的，并且由于激酶A和C以及碱性磷酸酶的加入不会改变ATP的作用，因此可以得出结论，ATP水解是不需要的。ATP结合过程似乎是维持神经元内向整流钾通道功能状态所必需的。膜的细胞质面上的Na+结合位点与ATP结合位点协同作用以稳定通道活性。
Four cDNA-encoding G-activated inwardly rectifying K+ channels have been cloned recently (Kubo, Y., Reuveny, E., Slesinger, P. A., Jan, Y. N., and Jan, L. Y. (1993) Nature 364, 802-806; Lesage, F., Duprat, F., Fink, M., Guillemare, E., Coppola, T., Lazdunski, M., and Hugnot, J. P. (1994) FEES Lett. 353, 37-42; Krapivinsky, G., Gor don, E. A., Wickman, K., Velimirovic, B., Krapivinsky, L., and Clapham, D. E. (1995) Nature 374, 135-141). We report the cloning of a mouse GIRK2 splice variant, noted mGIRK2A. Both channel proteins are functionally expressed in Xenopus oocytes upon injection of their cRNA, alone or in combination with the GIRK1 cRNA. Three GIRK channels, mGIRK1-3, are shown to be present in the brain. Colocalization in the same neurons of mGIRK1 and mGIRK2 supports the hypothesis that native channels are made by an heteromeric subunit assembly. GIRK3 channels have not been expressed successfully, even in the presence of the other types of subunits. However, GIRK3 chimeras with the amino and carboxyl-terminal of GIRK2 are functionally expressed in the presence of GIRK1. The expressed mGIRK2 and mGIRK1, -2 currents are blocked by Ba2+ and Cs+ ions. They are not regulated by protein kinase A and protein kinase C. Channel activity runs down in inside-out excised patches, and ATP is required to prevent this rundown. Since the nonhydrolyzable ATP analog AMP-PCP is also active and since addition of kinases A and C as well as alkaline phosphatase does not modify the ATP effect, it is concluded that ATP hydrolysis is not required. An ATP binding process appears to be essential for maintaining a functional state of the neuronal inward rectifier K+ channel. A Na+ binding site on the cytoplasmic face of the membrane acts in synergy with the ATP binding site to stabilize channel activity.