Robitaille Mélanie, McCullough Michael H, So Choon Leng, Roberts-Thomson Sarah J, Monteith Gregory R
School of Pharmacy and Pharmaceutical Science, The University of Queensland, 20 Cornwall Street, Woolloongabba, QLD 4102, Australia.
Eccles Institute of Neuroscience, John Curtin School of Medical Research, ANU College of Science and Medicine, The Australian National University, 131 Garran Road, Acton 2601, ACT, Australia; School of Computing, ANU College of Systems and Society, The Australian National University, 108 North Road, Acton 2601, ACT, Australia.
Biochim Biophys Acta Mol Cell Res. 2025 Oct;1872(7):120010. doi: 10.1016/j.bbamcr.2025.120010. Epub 2025 Jun 21.
Reduction in extracellular pH is common in pathophysiological conditions including neurological disorders, inflammation, and cancer. Like extracellular acidification, dysregulated calcium homeostasis is linked to various pathophysiological states including cancer. Changes in pH and calcium can theoretically be linked since the function of several calcium transporters is affected by pH changes. One such calcium channel is ORAI1, where changes in extracellular pH modulate gating. Upon depletion of the endoplasmic reticulum (ER) calcium store, coupling of stromal interacting molecule 1 (STIM1) with ORAI1 facilitates calcium influx and refills the ER calcium store, a process known as Store-Operated Calcium Entry (SOCE). ORAI1 expression has been reported to be increased in several cancer types including basal breast cancer. The ORAI1 protein is expressed as two variants, ORAI1α (long) and ORAI1β (short). The differential roles of ORAI1α and ORAI1β are still not fully understood. Previous studies have shown that ORAI1 gating properties are affected by both intracellular and extracellular pH changes, but none have investigated whether ORAI1α and ORAI1β variants differ in sensitivity to pH changes. In this study, we used cell-based calcium flux assays to compare ORAI1α- and ORAI1β-mediated calcium influx at different extracellular pHs. We developed a mathematical model of intracellular calcium dynamics incorporating SOCE to quantify variations in calcium influx. Fitting this model to our data revealed that at pH 6.8, ORAI1α is associated with more sustained calcium influx compared to ORAI1β. Point mutations in the N-terminus of ORAI1α identified potential domains responsible for the differential activation of ORAI1 variants at pH 6.8.
细胞外pH值降低在包括神经疾病、炎症和癌症在内的病理生理状况中很常见。与细胞外酸化一样，钙稳态失调与包括癌症在内的各种病理生理状态有关。理论上，pH值和钙的变化可能有关联，因为几种钙转运蛋白的功能会受到pH值变化的影响。一种这样的钙通道是ORAI1，细胞外pH值的变化会调节其门控。在内质网（ER）钙库耗尽时，基质相互作用分子1（STIM1）与ORAI1的偶联促进钙内流并补充ER钙库，这一过程称为储存-操纵性钙内流（SOCE）。据报道，ORAI1在包括基底型乳腺癌在内的几种癌症类型中表达增加。ORAI1蛋白以两种变体形式表达，即ORAI1α（长型）和ORAI1β（短型）。ORAI1α和ORAI1β的不同作用仍未完全了解。先前的研究表明，ORAI1的门控特性受细胞内和细胞外pH值变化的影响，但尚未有人研究ORAI1α和ORAI1β变体对pH值变化的敏感性是否不同。在本研究中，我们使用基于细胞的钙流测定法来比较不同细胞外pH值下ORAI1α和ORAI1β介导的钙内流。我们建立了一个包含SOCE的细胞内钙动力学数学模型，以量化钙内流的变化。将该模型与我们的数据拟合后发现，在pH 6.8时，与ORAI1β相比，ORAI1α与更持续的钙内流相关。ORAI1α N端的点突变确定了在pH 6.8时负责ORAI1变体差异激活的潜在结构域。
