Identification of an endogenous inhibitor of the cardiac Na+/Ca2+ exchanger, phospholemman
Identification of an endogenous inhibitor of the cardiac Na+/Ca2+ exchanger, phospholemman
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DOI：
10.1074/jbc.m414703200
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发表时间：
2005-05-20
影响因子：
4.8
通讯作者：
Cheung, JY
中科院分区：
文献类型：
--
作者：
Ahlers, BA;Zhang, XQ;Cheung, JY
快速和精确地控制Na+/Ca 2+交换器（NCX 1）活性对于维持心肌细胞搏动间Ca 2+稳态是必不可少的。在这里，我们表明，磷脂（PLM），一个15 kDa的整合肌膜磷蛋白，是一种新的内源性蛋白抑制剂的心脏NCX 1。使用一个异源表达系统，是缺乏内源性PLM和NCX 1，我们首先证明了通过共聚焦免疫荧光研究，外源性PLM和NCX 1共定位于质膜。相互免疫共沉淀研究揭示了PLM和NCX 1之间的特异性蛋白质-蛋白质相互作用。PLM与NCX 1直接结合的功能结果是抑制NCX 1活性，如测量NCX 1电流密度的全细胞膜片钳研究和评估Na+依赖性Ca-45（2+）摄取的放射性示踪剂通量测定所证明的。PLM对NCX 1的抑制是特异性的，因为PLM中丝氨酸68变为丙氨酸的单一突变导致NCX 1电流抑制的完全丧失，尽管PLM突变体与NCX 1的关联没有改变。在天然成人心肌细胞中，PLM与NCX 1共免疫沉淀。我们得出结论，PLM，已知调节Na+-K+-ATP酶的FXYD家族的小离子转运调节剂的成员，也调节心脏中的Na+/Ca 2+交换。
Rapid and precise control of Na+/Ca2+ exchanger (NCX1) activity is essential in the maintenance of beat-to-beat Ca2+ homeostasis in cardiac myocytes. Here, we show that phospholemman (PLM), a 15-kDa integral sarcolemmal phosphoprotein, is a novel endogenous protein inhibitor of cardiac NCX1. Using a heterologous expression system that is devoid of both endogenous PLM and NCX1, we first demonstrated by confocal immunofluorescence studies that both exogenous PLM and NCX1 co-localized at the plasma membrane. Reciprocal co-immunoprecipitation studies revealed specific protein-protein interaction between PLM and NCX1. The functional consequences of direct association of PLM with NCX1 was the inhibition of NCX1 activity, as demonstrated by whole-cell patch clamp studies to measure NCX1 current density and radiotracer flux assays to assess Na+-dependent Ca-45(2+) uptake. Inhibition of NCX1 by PLM was specific, because a single mutation of serine 68 to alanine in PLM resulted in a complete loss of inhibition of NCX1 current, although association of the PLM mutant with NCX1 was unaltered. In native adult cardiac myocytes, PLM co-immunoprecipitated with NCX1. We conclude that PLM, a member of the FXYD family of small ion transport regulators known to modulate Na+-K+-ATPase, also regulates Na+/Ca2+ exchange in the heart.