Molecular Determinants of the Coupling between STIM1 and Orai Channels DIFFERENTIAL ACTIVATION OF Orai1-3 CHANNELS BY A STIM1 COILED-COIL MUTANT
Molecular Determinants of the Coupling between STIM1 and Orai Channels DIFFERENTIAL ACTIVATION OF Orai1-3 CHANNELS BY A STIM1 COILED-COIL MUTANT
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DOI：
10.1074/jbc.m109.018408
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发表时间：
2009-08-07
影响因子：
4.8
通讯作者：
Romanin, Christoph
中科院分区：
文献类型：
--
作者：
Frischauf, Irene;Muik, Martin;Romanin, Christoph
据报道，STIM1和Orai1在存储器耗尽时相互作用，最终导致钙释放激活的钙电流激活。最近，STIM1C末端的必需区域已被发现，它包括第二个卷曲结构域，其末端延伸了类似50个氨基酸，并与Orai1C末端显示出强烈的结合。基于Orai家族内的同源性，可以假设Orai2和Orai3通道都是以类似的STIM1依赖方式激活的。电生理学和Foerster共振能量转移显微镜相结合的方法揭示了STIM1与Orai蛋白通讯的一般机制，涉及到各自Orai C末端保守的螺旋线圈结构域和STIM1 C末端的第二螺旋线圈基序。Orai1 C端的螺旋线圈单一突变导致与STIM1 C端的通讯中断，而Orai2和Orai3的类似突变仍允许它们适度激活。然而，通过增加Orai1的功能缺失或通过产生包含Orai3 C末端的Orai1-Orai3嵌合体来增加螺旋线圈的概率，恢复刺激的程度与Orai2/3相似。在STIM1水平上，通过STIM1 C末端的单个突变降低第二螺旋线圈结构域的概率取消了Orai1的激活，但仍能部分刺激Orai2/3通道。STIM1 C末端第二个螺旋螺旋基序的双重突变完全中断了与所有三个Orai通道的通讯。总体而言，STIM1和ORAI通道之间的整体通信受到的损害，由于C末端中任何一个假定的螺旋线圈结构域的概率降低，可能与它们功能上的异构体相互作用的概念相一致。
STIM1 and Orai1 have been reported to interact upon store depletion culminating in Ca2+ release-activated Ca2+ current activation. Recently, the essential region has been identified within the STIM1 C terminus that includes the second coiled-coil domain C-terminally extended by similar to 50 amino acids and exhibits a strong binding to the Orai1 C terminus. Based on the homology within the Orai family, an analogous scenario might be assumed for Orai2 as well as Orai3 channels as both are activated in a similar STIM1-dependent manner. A combined approach of electrophysiology and Foerster resonance energy transfer microscopy uncovered a general mechanism in the communication of STIM1 with Orai proteins that involved the conserved putative coiled-coil domains in the respective Orai C terminus and the second coiled-coil motif in the STIM1 C terminus. A coiled-coil single mutation in the Orai1 C terminus abrogated communication with the STIM1 C terminus, whereas an analogous mutation in Orai2 and Orai3 still allowed for their moderate activation. However, increasing coiled-coil probability by a gain of function deletion in Orai1 or by generating an Orai1-Orai3 chimera containing the Orai3 C terminus recovered stimulation to a similar extent as with Orai2/3. At the level of STIM1, decreasing probability of the second coiled-coil domain by a single mutation within the STIM1 C terminus abolished activation of Orai1 but still enabled partial stimulation of Orai2/3 channels. A double mutation within the second coiled-coil motif of the STIM1 C terminus fully disrupted communication with all three Orai channels. In aggregate, the impairment in the overall communication between STIM1 and Orai channels upon decreasing probabilities of either one of the putative coiled-coil domains in the C termini might be compatible with the concept of their functional, heteromeric interaction.