Pierce G N, Panagia V
Division of Cardiovascular Sciences, St. Boniface General Hospital Research Centre, Winnipeg, Canada.
J Biol Chem. 1989 Sep 15;264(26):15344-50.
The purpose of this investigation was to study the effects of a distinct type of phospholipase C on sarcolemmal Na+-Ca2+ exchange. With this phospholipase C (Staphylococcus aureus), treatment of cardiac sarcolemmal vesicles resulted in a specific hydrolysis of membrane phosphatidylinositol. This hydrolysis of phosphatidylinositol also released two proteins (110 and 36 kDa) from the sarcolemmal membrane. Phospholipase C pretreatment of the sarcolemma resulted in an unexpected stimulation of Na+-Ca2+ exchange. The Vmax of Na+-Ca2+ exchange was increased but the Km for Ca2+ was not altered. This stimulation was specific to the Na+-Ca2+ exchange pathway. ATP-dependent Ca2+ uptake was depressed after phospholipase C treatment, but passive membrane permeability to Ca2+ was unaffected. Sarcolemmal Na+,K+-ATPase activity was not altered, whereas passive Ca2+ binding was modestly decreased after phospholipase C pretreatment. The stimulation of Na+-Ca2+ exchange after phosphatidylinositol hydrolysis was greater in inside-out vesicles than in a total population of vesicles of mixed orientation. This finding suggests that the cardiac sarcolemmal Na+-Ca2+ exchanger is functionally asymmetrical. The results also suggest that membrane phosphatidylinositol is inhibitory to the Na+-Ca2+ exchanger or, alternatively, this phospholipid may anchor an endogenous inhibitory protein in the sarcolemmal membrane. The observation that a transsarcolemmal Ca2+ flux pathway may be stimulated solely by phosphatidylinositol hydrolysis independently of phosphoinositide metabolic products like inositol triphosphate is novel.
本研究的目的是研究一种独特类型的磷脂酶C对肌膜钠钙交换的影响。用这种磷脂酶C（金黄色葡萄球菌）处理心脏肌膜囊泡，导致膜磷脂酰肌醇发生特异性水解。这种磷脂酰肌醇的水解还从肌膜释放出两种蛋白质（110和36 kDa）。对肌膜进行磷脂酶C预处理导致钠钙交换出现意外的刺激。钠钙交换的Vmax增加，但对钙离子的Km未改变。这种刺激对钠钙交换途径具有特异性。磷脂酶C处理后，ATP依赖性钙摄取受到抑制，但膜对钙离子的被动通透性未受影响。肌膜钠钾ATP酶活性未改变，而磷脂酶C预处理后，被动钙结合略有减少。磷脂酰肌醇水解后对钠钙交换的刺激在内外翻转囊泡中比在混合取向的总囊泡群体中更大。这一发现表明心脏肌膜钠钙交换器在功能上是不对称的。结果还表明，膜磷脂酰肌醇对钠钙交换器具有抑制作用，或者，这种磷脂可能在肌膜中锚定一种内源性抑制蛋白。仅通过磷脂酰肌醇水解而不依赖于肌醇三磷酸等磷酸肌醇代谢产物来刺激跨肌膜钙通量途径这一观察结果是新颖的。
