Measurement of the CRAC Channel Fast Ca2+-Dependent Inactivation (FCDI)

2018 ◽  
pp. 167-173 ◽  
Author(s):  
Grigori Y. Rychkov
Keyword(s):  
Dependent Inactivation ◽  
Crac Channel

scholarly journals Regulation of Store-Operated Ca2+ Entry by SARAF

Cells ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1887
Author(s):  
Inbal Dagan ◽  
Raz Palty
Keyword(s):  
Molecular Mechanisms ◽  
Feedback Mechanism ◽  
Cellular Biology ◽  
Excitable Cells ◽  
Major Pathway ◽  
Negative Feedback Mechanism ◽  
Crac Channels ◽  
Dependent Inactivation ◽  
Crac Channel ◽  
The One

Calcium (Ca2+) signaling plays a dichotomous role in cellular biology, controlling cell survival and proliferation on the one hand and cellular toxicity and cell death on the other. Store-operated Ca2+ entry (SOCE) by CRAC channels represents a major pathway for Ca2+ entry in non-excitable cells. The CRAC channel has two key components, the endoplasmic reticulum Ca2+ sensor stromal interaction molecule (STIM) and the plasma-membrane Ca2+ channel Orai. Physical coupling between STIM and Orai opens the CRAC channel and the resulting Ca2+ flux is regulated by a negative feedback mechanism of slow Ca2+ dependent inactivation (SCDI). The identification of the SOCE-associated regulatory factor (SARAF) and investigations of its role in SCDI have led to new functional and molecular insights into how SOCE is controlled. In this review, we provide an overview of the functional and molecular mechanisms underlying SCDI and discuss how the interaction between SARAF, STIM1, and Orai1 shapes Ca2+ signaling in cells.

Bidirectional regulation of calcium release–activated calcium (CRAC) channel by SARAF

2021 ◽  
Vol 220 (12) ◽  
Author(s):  
Elia Zomot ◽  
Hadas Achildiev Cohen ◽  
Inbal Dagan ◽  
Ruslana Militsin ◽  
Raz Palty
Keyword(s):  
Gene Expression ◽  
Calcium Release ◽  
Cell Receptor ◽  
Crac Channels ◽  
Bidirectional Regulation ◽  
Dependent Inactivation ◽  
Store Operated Calcium Entry ◽  
Crac Channel ◽  
Initial Activation ◽  
Downstream Gene Expression

Store-operated calcium entry (SOCE) through the Ca2+ release–activated Ca2+ (CRAC) channel is a central mechanism by which cells generate Ca2+ signals and mediate Ca2+-dependent gene expression. The molecular basis for CRAC channel regulation by the SOCE-associated regulatory factor (SARAF) remained insufficiently understood. Here we found that following ER Ca2+ depletion, SARAF facilitates a conformational change in the ER Ca2+ sensor STIM1 that relieves an activation constraint enforced by the STIM1 inactivation domain (ID; aa 475–483) and promotes initial activation of STIM1, its translocation to ER–plasma membrane junctions, and coupling to Orai1 channels. Following intracellular Ca2+ rise, cooperation between SARAF and the STIM1 ID controls CRAC channel slow Ca2+-dependent inactivation. We further show that in T lymphocytes, SARAF is required for proper T cell receptor evoked transcription. Taking all these data together, we uncover a dual regulatory role for SARAF during both activation and inactivation of CRAC channels and show that SARAF fine-tunes intracellular Ca2+ responses and downstream gene expression in cells.

scholarly journals Structural and stoichiometric determinants of Ca2+ release-activated Ca2+ (CRAC) channel Ca2+-dependent inactivation

2014 ◽  
Vol 1838 (5) ◽  
pp. 1281-1287 ◽  
Author(s):  
Nathan R. Scrimgeour ◽  
David P. Wilson ◽  
Greg J. Barritt ◽  
Grigori Y. Rychkov
Keyword(s):  
Dependent Inactivation ◽  
Crac Channel

scholarly journals Regulation of Interorganellar Ca2+ Transfer and NFAT Activation by the Mitochondrial Ca2+ Uniporter

2021 ◽  
Author(s):  
Ryan E. Yoast ◽  
Scott M. Emrich ◽  
Xuexin Zhang ◽  
Ping Xin ◽  
Vikas Arige ◽  
...  
Keyword(s):  
Nuclear Translocation ◽  
Channel Activity ◽  
Crac Channels ◽  
Store Depletion ◽  
Agonist Stimulation ◽  
Dependent Inactivation ◽  
Nfat Activation ◽  
Crac Channel ◽  
Cellular Bioenergetics ◽  
Trisphosphate Receptor

Mitochondrial Ca2+ uptake is crucial for coupling receptor stimulation to cellular bioenergetics. Further, Ca2+ uptake by respiring mitochondria prevents Ca2+-dependent inactivation (CDI) of store-operated Ca2+ release-activated Ca2+ (CRAC) channels and inhibits Ca2+ extrusion to sustain cytosolic Ca2+ signaling. However, how Ca2+ uptake by the mitochondrial Ca2+ uniporter (MCU) shapes receptor-evoked interorganellar Ca2+ signaling is unknown. Here, we generated several cell lines with MCU-knockout (MCU-KO) as well as tissue-specific MCU-knockdown mice. We show that mitochondrial depolarization, but not MCU-KO, inhibits store-operated Ca2+ entry (SOCE). Paradoxically, despite enhancing Ca2+ extrusion and promoting CRAC channel CDI, MCU-KO increased cytosolic Ca2+ in response to store depletion. Further, physiological agonist stimulation in MCU-KO cells led to enhanced frequency of cytosolic Ca2+ oscillations, endoplasmic reticulum Ca2+ refilling, NFAT nuclear translocation and proliferation. However, MCU-KO did not affect inositol-1,4,5-trisphosphate receptor activity. Mathematical modeling supports that MCU-KO enhances cytosolic Ca2+, despite limiting CRAC channel activity.

scholarly journals Orai1α, but not Orai1β, co-localizes with TRPC1 and is required for its plasma membrane location and activation in HeLa cells

2022 ◽  
Vol 79 (1) ◽  
Author(s):  
Jose Sanchez-Collado ◽  
Jose J. Lopez ◽  
Isaac Jardin ◽  
Alejandro Berna-Erro ◽  
Pedro J. Camello ◽  
...  
Keyword(s):  
Amino Acids ◽  
Plasma Membrane ◽  
Hela Cells ◽  
Mammalian Cells ◽  
Store Depletion ◽  
Agonist Stimulation ◽  
Dependent Inactivation ◽  
Crac Channel ◽  
Membrane Location

AbstractThe identification of two variants of the canonical pore-forming subunit of the Ca2+ release-activated Ca2+ (CRAC) channel Orai1, Orai1α and Orai1β, in mammalian cells arises the question whether they exhibit different functional characteristics. Orai1α and Orai1β differ in the N-terminal 63 amino acids, exclusive of Orai1α, and show different sensitivities to Ca2+-dependent inactivation, as well as distinct ability to form arachidonate-regulated channels. We have evaluated the role of both Orai1 variants in the activation of TRPC1 in HeLa cells. We found that Orai1α and Orai1β are required for the maintenance of regenerative Ca2+ oscillations, while TRPC1 plays a role in agonist-induced Ca2+ influx but is not essential for Ca2+ oscillations. Using APEX2 proximity labeling, co-immunoprecipitation and the fluorescence of G-GECO1.2 fused to Orai1α our results indicate that agonist stimulation and Ca2+ store depletion enhance Orai1α–TRPC1 interaction. Orai1α is essential for TRPC1 plasma membrane location and activation. Thus, TRPC1 function in HeLa cells depends on Ca2+ influx through Orai1α exclusively.

Glu106 in the Orai1 pore contributes to fast Ca2+-dependent inactivation and pH dependence of Ca2+ release-activated Ca2+ (CRAC) current

2011 ◽  
Vol 441 (2) ◽  
pp. 743-753 ◽  
Author(s):  
Nathan R. Scrimgeour ◽  
David P. Wilson ◽  
Grigori Y. Rychkov
Keyword(s):  
Binding Site ◽  
Voltage Dependence ◽  
Ph Dependence ◽  
Low Ph ◽  
Wild Type ◽  
Channel Pore ◽  
Crac Channels ◽  
Dependent Inactivation ◽  
Crac Channel ◽  
Ca2 Channels

FCDI (fast Ca2+-dependent inactivation) is a mechanism that limits Ca2+ entry through Ca2+ channels, including CRAC (Ca2+ release-activated Ca2+) channels. This phenomenon occurs when the Ca2+ concentration rises beyond a certain level in the vicinity of the intracellular mouth of the channel pore. In CRAC channels, several regions of the pore-forming protein Orai1, and STIM1 (stromal interaction molecule 1), the sarcoplasmic/endoplasmic reticulum Ca2+ sensor that communicates the Ca2+ load of the intracellular stores to Orai1, have been shown to regulate fast Ca2+-dependent inactivation. Although significant advances in unravelling the mechanisms of CRAC channel gating have occurred, the mechanisms regulating fast Ca2+-dependent inactivation in this channel are not well understood. We have identified that a pore mutation, E106D Orai1, changes the kinetics and voltage dependence of the ICRAC (CRAC current), and the selectivity of the Ca2+-binding site that regulates fast Ca2+-dependent inactivation, whereas the V102I and E190Q mutants when expressed at appropriate ratios with STIM1 have fast Ca2+-dependent inactivation similar to that of WT (wild-type) Orai1. Unexpectedly, the E106D mutation also changes the pH dependence of ICRAC. Unlike WT ICRAC, E106D-mediated current is not inhibited at low pH, but instead the block of Na+ permeation through the E106D Orai1 pore by Ca2+ is diminished. These results suggest that Glu106 inside the CRAC channel pore is involved in co-ordinating the Ca2+-binding site that mediates fast Ca2+-dependent inactivation.

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