A Coiled-coil Clamp Controls Both Conformation and Clustering of Stromal Interaction Molecule 1 (STIM1)

Fahrner, Marc; Muik, Martin; Schindl, Rainer; Butorac, Carmen; Stathopulos, Peter B.; Zheng, Lirong; Jardín, Isaac; Ikura, Mitsuhiko; Romanin, Christoph

doi:10.1074/jbc.m114.610022

Cited by 109 publications

(236 citation statements)

References 48 publications

(123 reference statements)

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“…Moreover, the proposed Orai1-interacting sequences were exactly those same residues involved in SOAR-SOAR interactions that are buried deeply in the SOAR dimer. Another factor is that the site proposed earlier (23,24) did not include the exposed and highly active SOAR region surrounding the F394 residue that we suggest is a crucial locus for Orai1 interactions. Our surprising results with heteromeric SOAR concatemers suggested that a “unimolecular” interaction of SOAR with Orai1 is sufficient to activate the channel.…”

Section: A Unimolecular Coupling Model For Stim1-orai1 Interactionsmentioning

confidence: 78%

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The STIM-Orai coupling interface and gating of the Orai1 channel

et al. 2017

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In virtually all cells, store-operated Ca2+ entry signals are vital in controlling a spectrum of functions. The signals are mediated by STIM proteins in the ER and Orai channels in the PM which undergo a dynamic coupling process within discrete ER-PM junctional regions. This coupling is initiated by depletion of ER stored Ca2+ triggering STIM proteins to undergo an intricate activation process. Thereafter, STIM proteins become trapped in the ER-PM junctions where they tether and gate PM Orai Ca2+ channels. STIM1 exists as a dimer, with a single STIM-Orai activating region (SOAR) buried in the resting protein that becomes exposed upon activation. An exposed region on SOAR including the Phe-394 residue forms a critical Orai1 interacting site. Using dimeric SOAR concatemers, we reveal only one of the two sites in the SOAR dimer is needed for Orai1 activation. This unimolecular interaction of SOAR with Orai1 suggests STIM1 can cross-link Orai channels with important significance for Ca2+ signaling. A critical “nexus” region in Orai1 close to the STIM1-binding site can be mutated to constitutively activate the channel mimicking the gating action of STIM1. This indicates STIM1 remotely controls Orai1 channel gating through an allosteric switch triggered by STIM1 binding only to the exposed C-terminal tail of the Orai1 channel.

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Section: A Unimolecular Coupling Model For Stim1-orai1 Interactionsmentioning

confidence: 78%

“…2A. Recent NMR studies were considered to support this bimolecular model (23,24). However, those studies were based on short SOAR-derived peptides that had no Orai1-activating function.…”

Section: A Unimolecular Coupling Model For Stim1-orai1 Interactionsmentioning

confidence: 98%

The STIM-Orai coupling interface and gating of the Orai1 channel

et al. 2017

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“…The entire cytosolic C-terminal doman (STIM1ct), or just the STIM1-Orai1 Activating Region (SOAR) or CRAC-Activating Domain (CAD) of STIM1, each exist as cytosolic dimers when expressed (reviewed in (5)). Indeed, the SOAR/CAD region of STIM1 is essential for oligomerization and puncta formation of STIM1 (30, 31). Our results show that 2-APB does not induce any significant changes in FRET signals between STIM1ct (Fig.…”

Section: Resultsmentioning

confidence: 99%

Molecular mechanisms underlying inhibition of STIM1-Orai1-mediated Ca2+ entry induced by 2-aminoethoxydiphenyl borate

Wei

Zhou

Sun

et al. 2016

Pflugers Arch - Eur J Physiol

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Store operated Ca2+ entry (SOCE) mediated by STIM1 and Orai1 is crucial for Ca2+ signaling and homeostasis in most cell types. 2-aminoethoxydiphenyl borate (2-APB) is a well described SOCE inhibitor but its action remains largely elusive. Here we show that 2-APB does not affect the dimeric state of STIM1, but enhances the intramolecular coupling between the coiled-coil 1 (CC1) and STIM-Orai Activating Region (SOAR) of STIM1, with subsequent reduction in the formation of STIM1 puncta in the absence of Orai1overexpression. 2-APB also inhibits Orai1 channels, directly inhibiting Ca2+ entry through the constitutively active, STIM1-independent Orai1 mutants, Orai1-P245T and Orai1-V102A. When unbound with STIM1, the constitutively active Orai1-V102C mutant is not inhibited by 2-APB. Thus we used Orai1-V012C as a tool to examine whether 2-APB can also inhibit the coupling between STIM1 and Orai1. We find that the functional coupling between STIM1 and Orai1-V102C is inhibited by 2-APB. This inhibition on coupling is indirect, arising from 2-APB’s action on STIM1, and it is most likely mediated by gating residues in Orai1 N terminus. Overall, our finding of this two-direct-site inhibition may help better understanding of Orai1-activation by STIM1 and future drug design.

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“…This was supported by NMR studies that used shorter fragments of the STIM-Orai1-binding domain and a portion of the C-terminus STIM1 binding site of Orai1 (Stathopulos et al 2013; Fahrner et al 2014). These studies using short SOAR-derived peptides, however, selected regions of SOAR that were unable to activate Orai1 when co-expressed and did not include the high-affinity region surrounding the Phe-394 residue that was shown to be critical for STIM1-Orai1 coupling in our experiments (Stathopulos et al 2013; Fahrner et al 2014). Results from our lab that utilized SOAR concatemeric dimers containing F394H mutations to one or both subunits suggest that a “unimolecular” interaction between SOAR/STIM and Orai1 is necessary and sufficient for channel coupling and gating (Fig.…”

Section: 5 Soar-derived Probes To Understand Stim-orai Couplingmentioning

confidence: 90%

“…Interestingly, these experiments have led to the conclusion that the stoichiometry of interaction can vary, with maximum activation occurring at a STIM1:Orai1 ratio of 2:1 (Scrimgeour et al 2009; Hoover and Lewis 2011; Li et al 2011). However, recent structural studies utilizing partial SOAR and C-terminal STIM1-binding Orai1 TM4 fragments suggested that binding occurs in a bimolecular fashion, whereby a STIM1 dimer will bind to two adjacent Orai1 subunits within the same Orai1 hexamer (Stathopulos et al 2013; Fahrner et al 2014; Maus et al 2015). This model would indicate a stoichiometry of 1:1, yet it cannot explain the variable stoichiometry seen in previous experiments.…”

Section: 4 Coupling Models For the Stim-orai Interactionmentioning

confidence: 99%

The STIM-Orai Pathway: Conformational Coupling Between STIM and Orai in the Activation of Store-Operated Ca2+ Entry

Nwokonko

Cai

Loktionova

et al. 2017

Store-Operated Ca²⁺ Entry (SOCE) Pathways

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Store-operated Ca2+ entry fulfills a crucial role in controlling Ca2+ signals in almost all cells. The Ca2+-sensing stromal interaction molecule (STIM) proteins in the endoplasmic reticulum (ER) undergo complex conformational changes in response to depleted ER luminal Ca2+, allowing them to unfold and become trapped in ER-plasma membrane (PM) junctions. Dimers of STIM proteins trap and gate the plasma membrane Orai Ca2+ channels within these junctions to generate discrete zones of high Ca2+ and regulate sensitive Ca2+-dependent intracellular signaling pathways. The STIM-Orai activating region (SOAR) of STIM1 becomes exposed upon store depletion and promotes trapping of Orai1 at the PM. Residue Phe-394 within SOAR forms an integral part of the high-affinity Orai1-interacting site. Our results demonstrate that only a single active site within the dimeric SOAR domain of STIM1 is required for the activation of Orai1 channel activity. This unimolecular model is strongly supported by evidence of variable STIM1:Orai1 stoichiometry reported in many studies. We hypothesize that unimolecular coupling promotes cross-linking of channels, localizing Ca2+ signals, and regulating channel activity. We have also identified a key “nexus” region in Orai1 near the C-terminal STIM1-binding site that can be mutated to constitutively activate Ca2+ entry, mimicking STIM1 activated channels. This suggests that STIM1 mediates gating of Orai1 in an allosteric manner via interaction with the Orai1 C-terminus alone. This model suggests the dual role of STIM1 in regulating both localization and gating of Orai1 channels and has important implications for the regulation of SOCE-mediated downstream signaling and the kinetics of channel activation.

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A Coiled-coil Clamp Controls Both Conformation and Clustering of Stromal Interaction Molecule 1 (STIM1)

Cited by 109 publications

References 48 publications

The STIM-Orai coupling interface and gating of the Orai1 channel

The STIM-Orai coupling interface and gating of the Orai1 channel

Molecular mechanisms underlying inhibition of STIM1-Orai1-mediated Ca2+ entry induced by 2-aminoethoxydiphenyl borate

The STIM-Orai Pathway: Conformational Coupling Between STIM and Orai in the Activation of Store-Operated Ca2+ Entry

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