Department of Molecular Pharmacology and Biological Chemistry, Northwestern University School of Medicine, 303 E Chicago Avenue, Ward 8-296, Chicago, Illinois 60611, USA.
Nature. 2012 Jan 25;482(7384):241-5. doi: 10.1038/nature10752.
Two defining functional features of ion channels are ion selectivity and channel gating. Ion selectivity is generally considered an immutable property of the open channel structure, whereas gating involves transitions between open and closed channel states, typically without changes in ion selectivity. In store-operated Ca(2+) release-activated Ca(2+) (CRAC) channels, the molecular mechanism of channel gating by the CRAC channel activator, stromal interaction molecule 1 (STIM1), remains unknown. CRAC channels are distinguished by a very high Ca(2+) selectivity and are instrumental in generating sustained intracellular calcium concentration elevations that are necessary for gene expression and effector function in many eukaryotic cells. Here we probe the central features of the STIM1 gating mechanism in the human CRAC channel protein, ORAI1, and identify V102, a residue located in the extracellular region of the pore, as a candidate for the channel gate. Mutations at V102 produce constitutively active CRAC channels that are open even in the absence of STIM1. Unexpectedly, although STIM1-free V102 mutant channels are not Ca(2+)-selective, their Ca(2+) selectivity is dose-dependently boosted by interactions with STIM1. Similar enhancement of Ca(2+) selectivity is also seen in wild-type ORAI1 channels by increasing the number of STIM1 activation domains that are directly tethered to ORAI1 channels, or by increasing the relative expression of full-length STIM1. Thus, exquisite Ca(2+) selectivity is not an intrinsic property of CRAC channels but rather a tuneable feature that is bestowed on otherwise non-selective ORAI1 channels by STIM1. Our results demonstrate that STIM1-mediated gating of CRAC channels occurs through an unusual mechanism in which permeation and gating are closely coupled.
离子通道的两个定义性功能特征是离子选择性和通道门控。离子选择性通常被认为是开放通道结构的不变特性，而门控涉及开放和关闭通道状态之间的转变，通常不会改变离子选择性。在储存操作的钙（Ca2+）释放激活的钙（CRAC）通道中，CRAC 通道激活剂基质相互作用分子 1（STIM1）对通道门控的分子机制仍然未知。CRAC 通道的特点是具有非常高的 Ca2+选择性，对于许多真核细胞中产生持续的细胞内钙浓度升高是必不可少的，这对于基因表达和效应功能至关重要。在这里，我们探讨了人类 CRAC 通道蛋白 ORAI1 中 STIM1 门控机制的核心特征，并确定了位于孔外部区域的 V102 残基是通道门的候选者。V102 处的突变产生组成型激活的 CRAC 通道，即使在没有 STIM1 的情况下也保持开放。出乎意料的是，尽管没有 STIM1 的 V102 突变体通道没有 Ca2+选择性，但它们的 Ca2+选择性可通过与 STIM1 的相互作用而呈剂量依赖性增强。通过增加直接与 ORAI1 通道连接的 STIM1 激活结构域的数量，或通过增加全长 STIM1 的相对表达，也可以在野生型 ORAI1 通道中看到类似的 Ca2+选择性增强。因此，精美的 Ca2+选择性不是 CRAC 通道的固有特性，而是一种可调节的特性，通过 STIM1 赋予否则非选择性的 ORAI1 通道。我们的结果表明，CRAC 通道的 STIM1 介导的门控通过一种不寻常的机制发生，其中渗透和门控紧密耦合。
