Wild-type NM23-H1, but not its S120 mutants, suppresses desensitization of muscarinic potassium current.
Wild-type NM23-H1, but not its S120 mutants, suppresses desensitization of muscarinic potassium current.
复制标题
野生型 NM23-H1（而非其 S120 突变体）抑制毒蕈碱钾电流的脱敏。
DOI：
10.1016/s0167-4889(99)00009-9
复制
发表时间：
1999
期刊：
影响因子：
--
通讯作者：
Steeg,PS
中科院分区：
文献类型：
--
作者：
Otero,AS;Doyle,MB;Hartsough,MT;Steeg,PS
NM23(NDP激酶)通过一种不同于GTP再生的机制来调节激动剂对M胆碱K+通道的门控。为了更好地确定NM23在这一途径中的功能，并确定NM23中对其在M胆碱型K+通道功能中的作用至关重要的位置，我们利用了低水平表达NM23-H1的人乳腺癌细胞MDA-MB-435。M2受体和GIRK1/GIRK4通道亚基在稳定表达载体(对照)、野生型NM23-H1或几个NM23-H1突变体的细胞中共表达。所有被测细胞株的裂解物都显示出类似的二磷酸核苷(NDP)激酶活性。全细胞膜片钳记录显示，在过量表达NM23-H1的细胞中，M胆碱钾电流的急性脱敏显著减少。突变体NM23-H1P96S和NM23-H1S44A在降低脱敏能力方面与野生型NM23-H1相似。而突变体NM23-H1S120G和NM23-H1S120A则完全阻断NM23-H1对M胆碱钾电流的脱敏作用。此外，NM23-H1S120G加强了急性脱敏，表明该突变体保留了与毒碱途径相互作用的能力，但具有与野生型蛋白相反的特性。我们的结论是，NM23对导致M+电流脱敏的过程起着抑制作用，Ser-120对其作用是必不可少的。
NM23 (NDP kinase) modulates the gating of muscarinic K+channels by agonists through a mechanism distinct from GTP regeneration. To better define the function of NM23 in this pathway and to identify sites in NM23 that are important for its role in muscarinic K+channel function, we utilized MDA-MB-435 human breast carcinoma cells that express low levels of NM23-H1. M2 muscarinic receptors and GIRK1/GIRK4 channel subunits were co-expressed in cells stably transfected with vector only (control), wild-type NM23-H1, or several NM23-H1 mutants. Lysates from all cell lines tested exhibit comparable nucleoside diphosphate (NDP) kinase activity. Whole cell patch clamp recordings revealed a substantial reduction of the acute desensitization of muscarinic K+currents in cells overexpressing NM23-H1. The mutants NM23-H1P96Sand NM23-H1S44Aresembled wild-type NM23-H1 in their ability to reduce desensitization. In contrast, mutants NM23-H1S120Gand NM23-H1S120Acompletely abolished the effect of NM23-H1 on desensitization of muscarinic K+currents. Furthermore, NM23-H1S120Gpotentiated acute desensitization, indicating that this mutant retains the ability to interact with the muscarinic pathway, but has properties antithetical to those of the wild-type protein. We conclude that NM23 acts as a suppressor of the processes leading to the desensitization of muscarinic K+currents, and that Ser-120 is essential for its actions.