Protein kinase G regulates potassium chloride cotransporter-3 expression in primary cultures of rat vascular smooth muscle cells
Protein kinase G regulates potassium chloride cotransporter-3 expression in primary cultures of rat vascular smooth muscle cells
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DOI：
10.1074/jbc.m100901200
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发表时间：
2001-06-15
影响因子：
4.8
通讯作者：
Adragna, NC
中科院分区：
文献类型：
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作者：
Di Fulvio, M;Lincoln, TM;Adragna, NC
K-C1共转运（KCC）由一氧化氮供体激活，并似乎受到cGMP信号通路的调节。KCC mRNA（KCC 1-KCC 4）在大鼠血管平滑肌细胞（VSMC）中的表达是未知的。我们已经报道了KCC 1和KCC 3 mRNA的存在下，在原代培养的血管平滑肌细胞特异性逆转录聚合酶链反应。KCC 2 mRNA水平极低。KCC 4 mRNA未检测到。半定量逆转录-聚合酶链反应显示VSMCs中KCC 1/KCC 3 mRNA的比例为2：1。蛋白激酶G（PKG）-1从VSMCs的消耗没有改变KCC 3 mRNA的表达。用PKG-1-催化结构域-和仅载体-转染的缺乏内源性PEG的VSMCs获得类似的结果，表明PKG-1不参与维持基础KCC 3 mRNA表达。然而，8-溴-cGMP，PKG刺激剂，急性增加KCC 3 mRNA的表达，在浓度和时间依赖性的方式，这种效果被阻断的PKG抑制剂KT 5823，但不被放线菌素D。这些研究结果表明，VSMCs主要表达两种mRNA亚型，KCC 1和KCC 3，并建议PEG参与转录后在急性KCC 3 mRNA的调节。KCC 3对细胞体积和电解质稳态响应PKG调节剂的作用仍有待确定。
K-C1 cotransport (KCC) is activated by nitric oxide donors and appears to be regulated by the cGMP signaling pathway. Expression of KCC mRNAs (KCC1-KCC4) in rat vascular smooth muscle cells (VSMCs) is unknown. We have reported the presence of KCC1 and KCC3 mRNAs in primary cultures of VSMCs by specific reverse transcription-polymerase chain reaction. KCC2 mRNA appeared at extremely low levels. KCC4 mRNA was undetectable. Semiquantitative reverse transcription-polymerase chain reaction revealed a 2:1 KCC1/ KCC3 mRNA ratio in VSMCs. Depletion of protein kinase G (PKG)-1 from VSMCs did not change KCC3 mRNA expression. Analogous results were obtained with PKG-1-catalytic domain- and vector only-transfected VSMCs lacking endogenous PEG, suggesting no involvement of PKG-1 in the maintenance of basal KCC3 mRNA expression. However, 8-bromo-cGMP, a PKG stimulator, acutely increased KCC3 mRNA expression in a concentration- and time-dependent fashion; this effect was blocked by the PKG inhibitor KT5823 but not by actinomycin D, These findings show that VSMCs express mainly two mRNA isoforms, KCC1 and KCC3, and suggest that PEG participates post-transcriptionally in the acute KCC3 mRNA regulation. The role of KCC3 on cell volume and electrolyte homeostasis in response to PKG modulators remains to be determined.