TRPM7 activation potentiates SOCE in enamel cells but requires ORAI
TRPM7 activation potentiates SOCE in enamel cells but requires ORAI
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DOI：
10.1016/j.ceca.2020.102187
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发表时间：
2020-05-01
期刊：
影响因子：
4
通讯作者：
Lacruz, Rodrigo S.
中科院分区：
文献类型：
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作者：
Bomfim, Guilherme H. Souza;Costiniti, Veronica;Lacruz, Rodrigo S.
由 STIM1/2 和 ORAI (ORAI1-3) 蛋白介导的钙 (Ca2+) 释放激活 Ca2+ (CRAC) 通道在多种细胞中形成主要的钙池操纵 Ca2+ 进入 (SOCE) 途径。其中，称为成釉细胞的牙釉质形成细胞依靠 CRAC 通道功能来实现 Ca2+ 流入，这对于牙釉质矿化非常重要。 CRAC 通道的这一关键作用得到了缺乏 STIM1 和 ORAI1 的人类突变和动物模型的支持，这些突变和动物模型会导致牙釉质缺陷和矿化不足。最近的许多报告强调了 chanzyme TRPM7（瞬时受体电位黑色素 7）作为 SOCE 调节剂的作用，TRPM7 是一种跨膜蛋白，含有可渗透二价阳离子（Mg2+、Ca2+）的离子通道。这就提出了这样的问题：TRPM7 是否应被视为 Ca2+ 流入的替代途径，或者 TRPM7 是否会改变牙釉质细胞中的 CRAC 通道活性。为了解决这些问题，我们在大鼠原代牙釉质细胞和用毒胡萝卜素刺激的鼠成釉细胞系LS8细胞中，使用药理学TRPM7激活剂naltriben和抑制剂NS8593监测了SOCE介导的Ca2+流入。我们还测量了 ORAI1/2 缺陷 (shOrai1/2) LS8 细胞和 siRNA 敲低 Trpm7 的细胞中的 Ca2+ 动态。我们发现，与对照细胞相比，用 TRPM7 激活剂刺激的原代牙釉质细胞通过 SOCE 增强了 Ca2+ 流入。然而，用 NS8593 阻断 TRPM7 并没有降低 SOCE 峰值。此外，在缺乏 SOCE 的 shOrai1/2 LS8 细胞中激活 TRPM7 无法引起 Ca2+ 内流，并且 Trpm7 敲低对 SOCE 没有影响。综上所述，我们的数据表明 TRPM7 是 SOCE 的正调节剂，可增强牙釉质细胞中 Ca2+ 的流入，但其功能完全依赖于 ORAI 通道的先前激活。
Calcium (Ca2+) release-activated Ca2+ (CRAC) channels mediated by STIM1/2 and ORAI (ORAI1-3) proteins form the dominant store-operated Ca2+ entry (SOCE) pathway in a wide variety of cells. Among these, the enamel-forming cells known as ameloblasts rely on CRAC channel function to enable Ca2+ influx, which is important for enamel mineralization. This key role of the CRAC channel is supported by human mutations and animal models lacking STIM1 and ORAI1, which results in enamel defects and hypomineralization. A number of recent reports have highlighted the role of the chanzyme TRPM7 (transient receptor potential melastanin 7), a transmembrane protein containing an ion channel permeable to divalent cations (Mg2+, Ca2+), as a modulator of SOCE. This raises the question as to whether TRPM7 should be considered an alternative route for Ca2+ influx, or if TRPM7 modifies CRAC channel activity in enamel cells. To address these questions, we monitored Ca2+ influx mediated by SOCE using the pharmacological TRPM7 activator naltriben and the inhibitor NS8593 in rat primary enamel cells and in the murine ameloblast cell line LS8 cells stimulated with thapsigargin. We also measured Ca2+ dynamics in ORAI1/2-deficient (shOrai1/2) LS8 cells and in cells with siRNA knock-down of Trpm7. We found that primary enamel cells stimulated with the TRPM7 activator potentiated Ca2+ influx via SOCE compared to control cells. However, blockade of TRPM7 with NS8593 did not decrease the SOCE peak. Furthermore, activation of TRPM7 in shOrai1/2 LS8 cells lacking SOCE failed to elicit Ca2+ influx, and Trpm7 knock-down had no effect on SOCE. Taken together, our data suggest that TRPM7 is a positive modulator of SOCE potentiating Ca2+ influx in enamel cells, but its function is fully dependent on the prior activation of the ORAI channels.