School of Medical Sciences, University of Adelaide, Adelaide, South Australia, 5005, Australia.
School of Medicine, University of Adelaide, and South Australian Health and Medical Research Institute, Adelaide, South Australia, 5005, Australia.
Sci Rep. 2017 Aug 29;7(1):9829. doi: 10.1038/s41598-017-06371-0.
Ca release activated Ca (CRAC) channels composed of two cellular proteins, Ca-sensing stromal interaction molecule 1 (STIM1) and pore-forming Orai1, are the main mediators of the Ca entry pathway activated in response to depletion of intracellular Ca stores. Previously it has been shown that the amplitude of CRAC current (I) strongly depends on extracellular and intracellular pH. Here we investigate the intracellular pH (pH) dependence of I mediated by Orai1 and STIM1ectopically expressed in HEK293 cells. The results indicate that pH affects not only the amplitude of the current, but also Ca dependent gating of CRAC channels. Intracellular acidification changes the kinetics of I, introducing prominent re-activation component in the currents recorded in response to voltage steps to strongly negative potentials. I with similar kinetics can be observed at normal pH if the expression levels of Orai1 are increased, relative to the expression levels of STIM1. Mutations in the STIM1 inactivation domain significantly diminish the dependence of I kinetics on pH, but have no effect on pH dependence of I amplitude, implying that more than one mechanism is involved in CRAC channel regulation by intracellular pH.
钙释放激活钙 (CRAC) 通道由两种细胞蛋白组成，即钙感应基质相互作用分子 1 (STIM1) 和形成孔的 Orai1，是细胞内钙库耗竭时激活的钙内流途径的主要介导者。先前已经表明，CRAC 电流 (I) 的幅度强烈依赖于细胞外和细胞内 pH 值。在这里，我们研究了在 HEK293 细胞中过表达的 Orai1 和 STIM1 介导的 I 对细胞内 pH 值 (pH) 的依赖性。结果表明，pH 值不仅影响电流的幅度，还影响 CRAC 通道的 Ca 依赖性门控。细胞内酸化改变了 I 的动力学，在响应电压阶跃至强负电位的电流中引入了明显的再激活成分。如果 Orai1 的表达水平相对于 STIM1 的表达水平增加，则可以在正常 pH 值下观察到具有相似动力学的 I。STIM1 失活结构域的突变显著降低了 I 动力学对 pH 值的依赖性，但对 I 幅度的 pH 值依赖性没有影响，这意味着细胞内 pH 值对 CRAC 通道的调节涉及多种机制。
