Modulation of Na-K-2Cl cotransport by intracellular Cl(-) and protein kinase C-delta in Calu-3 cells.
Modulation of Na-K-2Cl cotransport by intracellular Cl(-) and protein kinase C-delta in Calu-3 cells.
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Calu-3 细胞中细胞内 Cl(-) 和蛋白激酶 C-delta 调节 Na-K-2Cl 共转运。
DOI：
10.1152/ajplung.00143.2001
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发表时间：
2002
期刊：
影响因子：
--
通讯作者：
Cole,ThomasS
中科院分区：
文献类型：
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作者：
Liedtke,CaroleM;Papay,Robert;Cole,ThomasS
在这项研究中，我们测试了细胞内Cl−（Cl）调节蛋白激酶C（PKC）-δ活性并因此激活Calu-3细胞系中Na-K-Cl共转运（NKCC 1）的假设。α1-肾上腺素能激动剂甲氧胺（MOX）和高渗蔗糖分别增加Cl和增加或减少细胞内体积，而不改变Cl浓度（[Cl−]i）。在制霉菌素透化的细胞单层中，[Cl−] i从20-140 mM的滴定不影响PKC-δ、PKC-β或罗特勒林敏感性NKCC 1的基线活性。在200 mM Cl−时，rottlerin敏感的NKCC 1被激活，PKC同种型主要定位于颗粒部分。MOX诱导NKCC 1活性和PKC-δ活性的双相增加以及PKC-δ和-κ B的颗粒定位。NKCC 1和PKC-δ的活性随Cl浓度的增加而降低，在140-200 mM Cl浓度时则明显升高，这是高[Cl-]水平的累加效应。Rottlerin抑制MOX的作用，这表明PKC-δ是激活NKCC 1所必需的。结果表明，在气道上皮细胞中，单独的Cl-电化学梯度不足以刺激NKCC 1活性;相反，PKC-δ活性的升高是必要的。此外，高Cl水平诱导PKC-δ的亚细胞再分布，这导致酶活性增加。
In this study, we tested the hypothesis that intracellular Cl−(Cl) regulates the activity of protein kinase C (PKC)-δ and thus the activation of Na-K-Cl cotransport (NKCC1) in a Calu-3 cell line. The α1-adrenergic agonist methoxamine (MOX) and hypertonic sucrose increased Cland increased or decreased intracellular volume, respectively, without changing Clconcentration ([Cl−]i). Titration of [Cl−]ifrom 20–140 mM in nystatin-permeabilized cell monolayers did not affect the baseline activity of PKC-δ, PKC-ζ, or rottlerin-sensitive NKCC1. At 200 mM Cl−, rottlerin-sensitive NKCC1 was activated, and PKC isotypes were localized predominantly to a particulate fraction. MOX induced a biphasic increase in NKCC1 activity and PKC-δ in activity and particulate localization of PKC-δ and -ζ. Activity of NKCC1 and PKC-δ decreased with increasing Clfrom 20 to 80 mM Clthen increased at 140–200 mM Clapparently as an additive effect to high [Cl−]ilevels. Rottlerin inhibited the effects of MOX, which indicates that PKC-δ was required for activation of NKCC1. The results indicate that, in airway epithelial cells, a Clelectrochemical gradient alone is not sufficient to stimulate NKCC1 activity; rather, elevated activity of PKC-δ is necessary. Further, high Cllevels induce a subcellular redistribution of PKC-δ, which results in increased enzyme activity.