Lurtz Monica M, Louis Charles F
Department of Cell Biology and Neuroscience, University of California, Riverside, CA 92521, USA.
Am J Physiol Cell Physiol. 2007 Dec;293(6):C1806-13. doi: 10.1152/ajpcell.00630.2006. Epub 2007 Sep 26.
The mechanism by which intracellular Ca(2+) concentration (Ca(2+)) regulates the permeability of gap junctions composed of connexin43 (Cx43) was investigated in HeLa cells stably transfected with this connexin. Extracellular addition of Ca(2+) in the presence of the Ca(2+) ionophore ionomycin produced a sustained elevation in Ca(2+) that resulted in an inhibition of the cell-to-cell transfer of the fluorescent dye Alexa fluor 594 (IC(50) of 360 nM Ca(2+)). The Ca(2+) dependency of this inhibition of Cx43 gap junctional permeability is very similar to that described in sheep lens epithelial cell cultures that express the three sheep lens connexins (Cx43, Cx44, and Cx49). The intracellular Ca(2+)-mediated decrease in cell-to-cell dye transfer was prevented by an inhibitor of calmodulin action but not by inhibitors of Ca(2+)/calmodulin-dependent protein kinase II or protein kinase C. In experiments that used HeLa cells transfected with a Cx43 COOH-terminus truncation mutant (Cx43(Delta257)), cell-to-cell coupling was similarly decreased by an elevation of Ca(2+) (IC(50) of 310 nM Ca(2+)) and similarly prevented by the addition of an inhibitor of calmodulin. These data indicate that physiological concentrations of Ca(2+) regulate the permeability of Cx43 in a calmodulin-dependent manner that does not require the major portion of the COOH terminus of Cx43.
在稳定转染了连接蛋白43（Cx43）的HeLa细胞中，研究了细胞内钙离子浓度（Ca(2+)）调节由Cx43组成的间隙连接通透性的机制。在钙离子载体离子霉素存在的情况下，细胞外添加钙离子会使Ca(2+)持续升高，导致荧光染料Alexa fluor 594的细胞间转移受到抑制（钙离子的半数抑制浓度（IC(50)）为360 nM）。这种对Cx43间隙连接通透性的抑制作用的钙离子依赖性，与在表达三种绵羊晶状体连接蛋白（Cx43、Cx44和Cx49）的绵羊晶状体上皮细胞培养物中所描述的情况非常相似。细胞内钙离子介导的细胞间染料转移减少可被钙调蛋白作用抑制剂阻止，但不能被钙离子/钙调蛋白依赖性蛋白激酶II或蛋白激酶C的抑制剂阻止。在使用转染了Cx43羧基末端截短突变体（Cx43(Delta257)）的HeLa细胞的实验中，Ca(2+)升高同样会使细胞间偶联减少（钙离子的IC(50)为310 nM），添加钙调蛋白抑制剂也同样能阻止这种减少。这些数据表明，生理浓度的Ca(2+)以钙调蛋白依赖性方式调节Cx43的通透性，且不需要Cx43羧基末端的主要部分。
