Dynamic interaction of SARAF with STIM1 and Orai1 to modulate store-operated calcium entry.
Dynamic interaction of SARAF with STIM1 and Orai1 to modulate store-operated calcium entry.
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DOI：
10.1038/srep24452
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发表时间：
2016-04-12
影响因子：
4.6
通讯作者：
Rosado JA
中科院分区：
文献类型：
--
作者：
Albarran L;Lopez JJ;Amor NB;Martin-Cano FE;Berna-Erro A;Smani T;Salido GM;Rosado JA
钙离子内流是细胞内钙稳态和细胞功能的主要机制。在这里，我们提出了SOCE相关调节因子(SARAF)、STIM1和Orai1之间动态相互作用的证据。在内源性表达STIM1和Orai1的细胞中，SARAF的过表达减弱了SOCE和STIM1-Orai1的相互作用，而RNAi介导的SARAF沉默则诱导了相反的作用。Saraf抑制了Orai1与共表达的Orai1激活的小片段STIM1在NG115-401L细胞中的表达。用thapsigargin或生理激动剂处理细胞后，Saraf与Orai1直接相关。不依赖STIM1的Saraf与Orai1的相互作用导致该通道的激活。在内源性表达STIM1和Orai1的细胞中，在thapsigargin处理约30s后，钙库枯竭导致Saraf与STIM1解离，这与Saraf-Orai1相互作用的增加平行，随后与STIM1重新相互作用，并与Orai1解离。Saraf与Orai1或各种N端缺失Orai1突变体的共表达并不改变Saraf与Orai1的相互作用；然而，C端缺失Orai1突变体的表达或阻断Orai1的C末端会削弱与Saraf的相互作用。这些观察结果表明，Saraf最初在激活SOCE中发挥积极作用，随后在促进Orai1的SCDI中发挥作用。
Ca2+ influx by store-operated Ca2+ channels is a major mechanism for intracellular Ca2+ homeostasis and cellular function. Here we present evidence for the dynamic interaction between the SOCE-associated regulatory factor (SARAF), STIM1 and Orai1. SARAF overexpression attenuated SOCE and the STIM1-Orai1 interaction in cells endogenously expressing STIM1 and Orai1 while RNAi-mediated SARAF silencing induced opposite effects. SARAF impaired the association between Orai1 and the Orai1-activating small fragment of STIM1 co-expressed in the STIM1-deficient NG115-401L cells. Cell treatment with thapsigargin or physiological agonists results in direct association of SARAF with Orai1. STIM1-independent interaction of SARAF with Orai1 leads to activation of this channel. In cells endogenously expressing STIM1 and Orai1, Ca2+ store depletion leads to dissociation of SARAF with STIM1 approximately 30s after treatment with thapsigargin, which paralleled the increase in SARAF-Orai1 interaction, followed by reinteraction with STIM1 and dissociation from Orai1. Co-expression of SARAF and either Orai1 or various N-terminal deletion Orai1 mutants did not alter SARAF-Orai1 interaction; however, expression of C-terminal deletion Orai1 mutants or blockade of the C-terminus of Orai1 impair the interaction with SARAF. These observations suggest that SARAF exerts an initial positive role in the activation of SOCE followed by the facilitation of SCDI of Orai1.