Journal articles: 'Orai1'

1

Eckstein, Miriam, Martin Vaeth, Francisco J. Aulestia, Veronica Costiniti, Serena N. Kassam, Timothy G. Bromage, Pal Pedersen, et al. "Differential regulation of Ca2+ influx by ORAI channels mediates enamel mineralization." Science Signaling 12, no. 578 (April 23, 2019): eaav4663. http://dx.doi.org/10.1126/scisignal.aav4663.

Full text

Abstract:

Store-operated Ca2+ entry (SOCE) channels are highly selective Ca2+ channels activated by the endoplasmic reticulum (ER) sensors STIM1 and STIM2. Their direct interaction with the pore-forming plasma membrane ORAI proteins (ORAI1, ORAI2, and ORAI3) leads to sustained Ca2+ fluxes that are critical for many cellular functions. Mutations in the human ORAI1 gene result in immunodeficiency, anhidrotic ectodermal dysplasia, and enamel defects. In our investigation of the role of ORAI proteins in enamel, we identified enamel defects in a patient with an ORAI1 null mutation. Targeted deletion of the Orai1 gene in mice showed enamel defects and reduced SOCE in isolated enamel cells. However, Orai2−/− mice showed normal enamel despite having increased SOCE in the enamel cells. Knockdown experiments in the enamel cell line LS8 suggested that ORAI2 and ORAI3 modulated ORAI1 function, with ORAI1 and ORAI2 being the main contributors to SOCE. ORAI1-deficient LS8 cells showed altered mitochondrial respiration with increased oxygen consumption rate and ATP, which was associated with altered redox status and enhanced ER Ca2+ uptake, likely due to S-glutathionylation of SERCA pumps. Our findings demonstrate an important role of ORAI1 in Ca2+ influx in enamel cells and establish a link between SOCE, mitochondrial function, and redox homeostasis.

APA, Harvard, Vancouver, ISO, and other styles

2

Jardin, Isaac, Alejandro Berna-Erro, Joel Nieto-Felipe, Alvaro Macias, Jose Sanchez-Collado, Jose J. Lopez, Gines M. Salido, and Juan A. Rosado. "Similarities and Differences between the Orai1 Variants: Orai1α and Orai1β." International Journal of Molecular Sciences 23, no. 23 (November 23, 2022): 14568. http://dx.doi.org/10.3390/ijms232314568.

Full text

Abstract:

Orai1, the first identified member of the Orai protein family, is ubiquitously expressed in the animal kingdom. Orai1 was initially characterized as the channel responsible for the store-operated calcium entry (SOCE), a major mechanism that allows cytosolic calcium concentration increments upon receptor-mediated IP3 generation, which results in intracellular Ca2+ store depletion. Furthermore, current evidence supports that abnormal Orai1 expression or function underlies several disorders. Orai1 is, together with STIM1, the key element of SOCE, conducting the Ca2+ release-activated Ca2+ (CRAC) current and, in association with TRPC1, the store-operated Ca2+ (SOC) current. Additionally, Orai1 is involved in non-capacitative pathways, as the arachidonate-regulated or LTC4-regulated Ca2+ channel (ARC/LRC), store-independent Ca2+ influx activated by the secretory pathway Ca2+-ATPase (SPCA2) and the small conductance Ca2+-activated K+ channel 3 (SK3). Furthermore, Orai1 possesses two variants, Orai1α and Orai1β, the latter lacking 63 amino acids in the N-terminus as compared to the full-length Orai1α form, which confers distinct features to each variant. Here, we review the current knowledge about the differences between Orai1α and Orai1β, the implications of the Ca2+ signals triggered by each variant, and their downstream modulatory effect within the cell.

APA, Harvard, Vancouver, ISO, and other styles

3

Liang, Ch, and F. Wu. "Reconstitution of calcium channel protein Orai3 into liposomes for functional studies." Биохимия 88, no. 9 (December 15, 2023): 1570–80. http://dx.doi.org/10.31857/s0320972523090099.

Full text

Abstract:

Store-operated calcium entry (SOCE) is the main mechanism for the Ca2+ influx in non-excitable cells. The two major components of SOCE are stromal interaction molecule 1 (STIM1) in the endoplasmic reticulum and Ca2+ release-activated Ca2+ channel (CRAC) Orai on the plasma membrane. SOCE requires interaction between STIM1 and Orai. Mammals have three Orai homologs: Orai1, Orai2, and Orai3. Although Orai1 has been widely studied and proven to be essential for numerous cellular processes, Orai3 has also attracted a significant attention recently. The gating and activation mechanisms of Orai3 have yet to be fully elucidated. Here, we expressed, purified, and reconstituted Orai3 protein into liposomes and investigated its orientation and oligomeric state in the resulting proteoliposomes. STIM1 interacted with the Orai3-containing proteoliposomes and mediated calcium release from them, suggesting that the Orai3 channel was functional and that recombinant STIM1 could directly open the Orai3 channel in vitro. The developed in vitro calcium release system could be used to study the structure, function, and pharmacology of Orai3 channel.

APA, Harvard, Vancouver, ISO, and other styles

4

Fresquez, Adriana M., and Carl White. "Extracellular cysteines C226 and C232 mediate hydrogen sulfide-dependent inhibition of Orai3-mediated store-operated calcium entry." American Journal of Physiology-Cell Physiology 322, no. 1 (January 1, 2022): C38—C48. http://dx.doi.org/10.1152/ajpcell.00490.2019.

Full text

Abstract:

The gaseous signaling molecule hydrogen sulfide (H2S) physiologically regulates store-operated Ca2+ entry (SOCE). The SOCE machinery consists of the plasma membrane-localized Orai channels (Orai1-3) and endoplasmic reticulum-localized stromal interaction molecule (STIM)1 and STIM2 proteins. H2S inhibits Orai3- but not Orai1- or Orai2-mediated SOCE. The current objective was to define the mechanism by which H2S selectively modifies Orai3. We measured SOCE and STIM1/Orai3 dynamics and interactions in HEK293 cells exogenously expressing fluorescently tagged human STIM1 and Orai3 in the presence and absence of the H2S donor GYY4137. Two cysteines (C226 and C232) are present in Orai3 that are absent in the Orai1 and Orai2. When we mutated either of these cysteines to serine, alone or in combination, SOCE inhibition by H2S was abolished. We also established that inhibition was dependent on an interaction with STIM1. To further define the effects of H2S on STIM1/Orai3 interaction, we performed a series of fluorescence recovery after photobleaching (FRAP), colocalization, and fluorescence resonance energy transfer (FRET) experiments. Treatment with H2S did not affect the mobility of Orai3 in the membrane, nor did it influence STIM1/Orai3 puncta formation or STIM1-Orai3 protein-protein interactions. These data support a model in which H2S modification of Orai3 at cysteines 226 and 232 limits SOCE evoked upon store depletion and STIM1 engagement, by a mechanism independent of the interaction between Orai3 and STIM1.

APA, Harvard, Vancouver, ISO, and other styles

5

Smyth, Jeremy T., and James W. Putney. "Regulation of store-operated calcium entry during cell division." Biochemical Society Transactions 40, no. 1 (January 19, 2012): 119–23. http://dx.doi.org/10.1042/bst20110612.

Full text

Abstract:

Store-operate Ca2+ channels gate Ca2+ entry into the cytoplasm in response to the depletion of Ca2+ from endoplasmic reticulum Ca2+ stores. The major molecular components of store-operated Ca2+ entry are STIM (stromal-interacting molecule) 1 (and in some instances STIM2) that serves as the endoplasmic reticulum Ca2+ sensor, and Orai (Orai1, Orai2 and Orai3) which function as pore-forming subunits of the store-operated channel. It has been known for some time that store-operated Ca2+ entry is shut down during cell division. Recent work has revealed complex mechanisms regulating the functions and locations of both STIM1 and Orai1 in dividing cells.

APA, Harvard, Vancouver, ISO, and other styles

6

Grimes, Derayvia, Ryan Johnson, Madeline Pashos, Celeste Cummings, Chen Kang, Georgia R. Sampedro, Eric Tycksen, et al. "ORAI1 and ORAI2 modulate murine neutrophil calcium signaling, cellular activation, and host defense." Proceedings of the National Academy of Sciences 117, no. 39 (September 14, 2020): 24403–14. http://dx.doi.org/10.1073/pnas.2008032117.

Full text

Abstract:

Calcium signals are initiated in immune cells by the process of store-operated calcium entry (SOCE), where receptor activation triggers transient calcium release from the endoplasmic reticulum, followed by opening of plasma-membrane calcium-release activated calcium (CRAC) channels. ORAI1, ORAI2, and ORAI3 are known to comprise the CRAC channel; however, the contributions of individual isoforms to neutrophil function are not well understood. Here, we show that loss of ORAI1 partially decreases calcium influx, while loss of both ORAI1 and ORAI2 completely abolishes SOCE. In other immune-cell types, loss of ORAI2 enhances SOCE. In contrast, we find that ORAI2-deficient neutrophils display decreased calcium influx, which is correlated with measurable differences in the regulation of neutrophil membrane potential via KCa3.1. Decreased SOCE in ORAI1-, ORAI2-, and ORAI1/2-deficient neutrophils impairs multiple neutrophil functions, including phagocytosis, degranulation, leukotriene, and reactive oxygen species (ROS) production, rendering ORAI1/2-deficient mice highly susceptible to staphylococcal infection. This study demonstrates that ORAI1 and ORAI2 are the primary components of the neutrophil CRAC channel and identifies subpopulations of neutrophils where cell-membrane potential functions as a rheostat to modulate the SOCE response. These findings have implications for mechanisms that modulate neutrophil function during infection, acute and chronic inflammatory conditions, and cancer.

APA, Harvard, Vancouver, ISO, and other styles

7

Tiffner, Adéla, and Isabella Derler. "Isoform-Specific Properties of Orai Homologues in Activation, Downstream Signaling, Physiology and Pathophysiology." International Journal of Molecular Sciences 22, no. 15 (July 27, 2021): 8020. http://dx.doi.org/10.3390/ijms22158020.

Full text

Abstract:

Ca2+ ion channels are critical in a variety of physiological events, including cell growth, differentiation, gene transcription and apoptosis. One such essential entry pathway for calcium into the cell is the Ca2+ release-activated Ca2+ (CRAC) channel. It consists of the Ca2+ sensing protein, stromal interaction molecule 1 (STIM1) located in the endoplasmic reticulum (ER) and a Ca2+ ion channel Orai in the plasma membrane. The Orai channel family includes three homologues Orai1, Orai2 and Orai3. While Orai1 is the “classical” Ca2+ ion channel within the CRAC channel complex and plays a universal role in the human body, there is increasing evidence that Orai2 and Orai3 are important in specific physiological and pathophysiological processes. This makes them an attractive target in drug discovery, but requires a detailed understanding of the three Orai channels and, in particular, their differences. Orai channel activation is initiated via Ca2+ store depletion, which is sensed by STIM1 proteins, and induces their conformational change and oligomerization. Upon STIM1 coupling, Orai channels activate to allow Ca2+ permeation into the cell. While this activation mechanism is comparable among the isoforms, they differ by a number of functional and structural properties due to non-conserved regions in their sequences. In this review, we summarize the knowledge as well as open questions in our current understanding of the three isoforms in terms of their structure/function relationship, downstream signaling and physiology as well as pathophysiology.

APA, Harvard, Vancouver, ISO, and other styles

8

Zhang, Xuexin, Ping Xin, Ryan E. Yoast, Scott M. Emrich, Martin T. Johnson, Trayambak Pathak, J. Cory Benson, et al. "Distinct pharmacological profiles of ORAI1, ORAI2, and ORAI3 channels." Cell Calcium 91 (November 2020): 102281. http://dx.doi.org/10.1016/j.ceca.2020.102281.

Full text

APA, Harvard, Vancouver, ISO, and other styles

9

Baryshnikov, Sergey G., Maria V. Pulina, Alessandra Zulian, Cristina I. Linde, and Vera A. Golovina. "Orai1, a critical component of store-operated Ca2+ entry, is functionally associated with Na+/Ca2+ exchanger and plasma membrane Ca2+ pump in proliferating human arterial myocytes." American Journal of Physiology-Cell Physiology 297, no. 5 (November 2009): C1103—C1112. http://dx.doi.org/10.1152/ajpcell.00283.2009.

Full text

Abstract:

Ca2+ entry through store-operated channels (SOCs) in the plasma membrane plays an important role in regulation of vascular smooth muscle contraction, tone, and cell proliferation. The C-type transient receptor potential (TRPC) channels have been proposed as major candidates for SOCs in vascular smooth muscle. Recently, two families of transmembrane proteins, Orai [also known as Ca2+ release-activated Ca2+ channel modulator (CRACM)] and stromal interacting molecule 1 (STIM1), were shown to be essential for the activation of SOCs mainly in nonexcitable cells. Here, using small interfering RNA, we show that Orai1 plays an essential role in activating store-operated Ca2+ entry (SOCE) in primary cultured proliferating human aortic smooth muscle cells (hASMCs), whereas Orai2 and Orai3 do not contribute to SOCE. Knockdown of Orai1 protein expression significantly attenuated SOCE. Moreover, inhibition of Orai1 downregulated expression of Na+/Ca2+ exchanger type 1 (NCX1) and plasma membrane Ca2+ pump isoform 1 (PMCA1). The rate of cytosolic free Ca2+ concentration decay after Ca2+ transients in Ca2+-free medium was also greatly decreased under these conditions. This reduction of Ca2+ extrusion, presumably via NCX1 and PMCA1, may be a compensation for the reduced SOCE. Immunocytochemical observations indicate that Orai1 and NCX1 are clustered in plasma membrane microdomains. Cell proliferation was attenuated in hASMCs with disrupted Orai1 expression and reduced SOCE. Thus Orai1 appears to be a critical component of SOCE in proliferating vascular smooth muscle cells, and may therefore be a key player during vascular growth and remodeling.

APA, Harvard, Vancouver, ISO, and other styles

10

Liu, Xiaoling, Tianyuan Zheng, Yan Jiang, Lei Wang, Yuchen Zhang, Qiyu Liang, and Yuejie Chen. "Molecular Mechanism Analysis of STIM1 Thermal Sensation." Cells 12, no. 22 (November 12, 2023): 2613. http://dx.doi.org/10.3390/cells12222613.

Full text

Abstract:

STIM1 has been identified as a new warm sensor, but the exact molecular mechanism remains unclear. In this study, a variety of mutants of STIM1, Orai1 and Orai3 were generated. The single–cell calcium imaging and confocal analysis were used to evaluate the thermal sensitivity of the resulting STIM mutants and the interaction between STIM1 and Orai mutants in response to temperature. Our results suggested that the CC1–SOAR of STIM1 was a direct activation domain of temperature, leading to subsequent STIM1 activation, and the transmembrane (TM) region and K domain but not EF–SAM were needed for this process. Furthermore, both the TM and SOAR domains exhibited similarities and differences between STIM1–mediated thermal sensation and store–operated calcium entry (SOCE), and the key sites of Orai1 showed similar roles in these two responses. Additionally, the TM23 (comprising TM2, loop2, and TM3) region of Orai1 was identified as the key domain determining the STIM1/Orai1 thermal response pattern, while the temperature reactive mode of STIM1/Orai3 seemed to result from a combined effect of Orai3. These findings provide important support for the specific molecular mechanism of STIM1–induced thermal response, as well as the interaction mechanism of STIM1 with Orai1 and Orai3 after being activated by temperature.

APA, Harvard, Vancouver, ISO, and other styles

11

Bisaillon, Jonathan M., Rajender K. Motiani, José C. Gonzalez-Cobos, Marie Potier, Katharine E. Halligan, Wael F. Alzawahra, Margarida Barroso, Harold A. Singer, David Jourd'heuil, and Mohamed Trebak. "Essential role for STIM1/Orai1-mediated calcium influx in PDGF-induced smooth muscle migration." American Journal of Physiology-Cell Physiology 298, no. 5 (May 2010): C993—C1005. http://dx.doi.org/10.1152/ajpcell.00325.2009.

Full text

Abstract:

We recently demonstrated that thapsigargin-induced passive store depletion activates Ca2+ entry in vascular smooth muscle cells (VSMC) through stromal interaction molecule 1 (STIM1)/Orai1, independently of transient receptor potential canonical (TRPC) channels. However, under physiological stimulations, despite the ubiquitous depletion of inositol 1,4,5-trisphosphate-sensitive stores, many VSMC PLC-coupled agonists (e.g., vasopressin and endothelin) activate various store-independent Ca2+ entry channels. Platelet-derived growth factor (PDGF) is an important VSMC promigratory agonist with an established role in vascular disease. Nevertheless, the molecular identity of the Ca2+ channels activated by PDGF in VSMC remains unknown. Here we show that inhibitors of store-operated Ca2+ entry (Gd3+ and 2-aminoethoxydiphenyl borate at concentrations as low as 5 μM) prevent PDGF-mediated Ca2+ entry in cultured rat aortic VSMC. Protein knockdown of STIM1, Orai1, and PDGF receptor-β (PDGFRβ) impaired PDGF-mediated Ca2+ influx, whereas Orai2, Orai3, TRPC1, TRPC4, and TRPC6 knockdown had no effect. Scratch wound assay showed that knockdown of STIM1, Orai1, or PDGFRβ inhibited PDGF-mediated VSMC migration, but knockdown of STIM2, Orai2, and Orai3 was without effect. STIM1, Orai1, and PDGFRβ mRNA levels were upregulated in vivo in VSMC from balloon-injured rat carotid arteries compared with noninjured control vessels. Protein levels of STIM1 and Orai1 were also upregulated in medial and neointimal VSMC from injured carotid arteries compared with noninjured vessels, as assessed by immunofluorescence microscopy. These results establish that STIM1 and Orai1 are important components for PDGF-mediated Ca2+ entry and migration in VSMC and are upregulated in vivo during vascular injury and provide insights linking PDGF to STIM1/Orai1 during neointima formation.

APA, Harvard, Vancouver, ISO, and other styles

12

Sanchez-Collado, Jose, Isaac Jardin, Jose J. López, Victor Ronco, Gines M. Salido, Charlotte Dubois, Natalia Prevarskaya, and Juan A. Rosado. "Role of Orai3 in the Pathophysiology of Cancer." International Journal of Molecular Sciences 22, no. 21 (October 22, 2021): 11426. http://dx.doi.org/10.3390/ijms222111426.

Full text

Abstract:

The mammalian exclusive Orai3 channel participates in the generation and/or modulation of two independent Ca2+ currents, the store-operated current, Icrac, involving functional interactions between the stromal interaction molecules (STIM), STIM1/STIM2, and Orai1/Orai2/Orai3, as well as the store-independent arachidonic acid (AA) (or leukotriene C4)-regulated current Iarc, which involves Orai1, Orai3 and STIM1. Overexpression of functional Orai3 has been described in different neoplastic cells and cancer tissue samples as compared to non-tumor cells or normal adjacent tissue. In these cells, Orai3 exhibits a cell-specific relevance in Ca2+ influx. In estrogen receptor-positive breast cancer cells and non-small cell lung cancer (NSCLC) cells store-operated Ca2+ entry (SOCE) is strongly dependent on Orai3 expression while in colorectal cancer and pancreatic adenocarcinoma cells Orai3 predominantly modulates SOCE. On the other hand, in prostate cancer cells Orai3 expression has been associated with the formation of Orai1/Orai3 heteromeric channels regulated by AA and reduction in SOCE, thus leading to enhanced proliferation. Orai3 overexpression is associated with supporting several cancer hallmarks, including cell cycle progression, proliferation, migration, and apoptosis resistance. This review summarizes the current knowledge concerning the functional role of Orai3 in the pathogenesis of cancer.

APA, Harvard, Vancouver, ISO, and other styles

13

El Assar, Mariam, Esther García-Rojo, Alejandro Sevilleja-Ortiz, Alberto Sánchez-Ferrer, Argentina Fernández, Borja García-Gómez, Javier Romero-Otero, Leocadio Rodríguez-Mañas, and Javier Angulo. "Functional Role of STIM-1 and Orai1 in Human Microvascular Aging." Cells 11, no. 22 (November 18, 2022): 3675. http://dx.doi.org/10.3390/cells11223675.

Full text

Abstract:

The impact of aging on vascular function is heterogeneous depending on the vascular territories. Calcium regulation plays a key role in vascular function and has been implicated in aging-related hypercontractility of corpus cavernosum. We aimed to evaluate stromal interaction molecule (STIM)/Orai system involvement in aging-related vascular alterations in the human macro and microvasculature. Aortae specimens and mesenteric arteries (MA), obtained from 45 organ donors, were functionally evaluated in organ chambers and wire myographs. Subjects were divided into groups either younger or older than 65-years old. The expressions of STIM-1, Orai1, and Orai3 were determined by immunofluorescence in the aorta and MA, and by Western blot in the aorta homogenates. The inhibition of STIM/Orai with YM-58483 (20 μM) reversed adrenergic hypercontractility in MA from older subjects but did not modify aging-related hypercontractility in the aortic strips. Aging was related to an increased expression of Orai1 in human aorta, while Orai1 and STIM-1 were upregulated in MA. STIM-1 and Orai1 protein expressions were inversely correlated to endothelial function in MA. Circulating levels of Orai1 were correlated with the inflammatory factor TNF-α and with the endothelial dysfunction marker asymmetric dimethylarginine. Aging is associated with an increased expression of the STIM/Orai system in human vessels with functional relevance only in the microvascular territory, suggesting its role in aging-related microvascular dysfunction.

APA, Harvard, Vancouver, ISO, and other styles

14

Angulo, Javier, Argentina Fernández, Alejandro Sevilleja-Ortiz, Alberto Sánchez-Ferrer, Leocadio Rodríguez-Mañas, and Mariam El Assar. "Upregulation of Orai Channels Contributes to Aging-Related Vascular Alterations in Rat Coronary Arteries." International Journal of Molecular Sciences 24, no. 17 (August 29, 2023): 13402. http://dx.doi.org/10.3390/ijms241713402.

Full text

Abstract:

Vascular territories display heterogeneous sensitivity to the impacts of aging. The relevance of the STIM/Orai system to vascular function depends on the vascular bed. We aimed to evaluate the contribution of the STIM/Orai system to aging-related vascular dysfunction in rat coronary circulation. Vascular function was evaluated according to myography in coronary arteries from young (three-month-old) and older (twenty-month-old) rats. The effects of aging and STIM/Orai inhibition on the contraction and relaxation of the coronary arteries and on the protein expression of STIM-1, Orai1, and Orai3 in these vessels were determined. Aging-related hypercontractility to serotonin and endothelin-1 in arteries from male rats was reversed by STIM/Orai inhibition with YM-58483 or by specifically blocking the Orai1 channel with Synta66. The inhibitory effects of Synta66 on coronary vasoconstriction were also observed in older female rats. YM-58483 relaxed serotonin- but not KCl-contracted arteries from males. STIM/Orai inhibition improved defective endothelial vasodilations in aged arteries, even in the presence of NO synthase and cyclooxygenase inhibitors, but not in KCl-contracted segments. YM-58483 significantly enhanced relaxations to calcium-activated potassium channel stimulation in aged vessels. Increased protein expression of Orai1 and Orai3 was detected in arterial homogenates and sections from older rats. Upregulation of the Orai channel contributes to aging-related coronary dysfunction, revealing a potential target in reducing CVD risk.

APA, Harvard, Vancouver, ISO, and other styles

15

Zhang, Xuexin, Wei Zhang, José C. González-Cobos, Isaac Jardin, Christoph Romanin, Khalid Matrougui, and Mohamed Trebak. "Complex role of STIM1 in the activation of store-independent Orai1/3 channels." Journal of General Physiology 143, no. 3 (February 24, 2014): 345–59. http://dx.doi.org/10.1085/jgp.201311084.

Full text

Abstract:

Orai proteins contribute to Ca2+ entry into cells through both store-dependent, Ca2+ release–activated Ca2+ (CRAC) channels (Orai1) and store-independent, arachidonic acid (AA)-regulated Ca2+ (ARC) and leukotriene C4 (LTC4)-regulated Ca2+ (LRC) channels (Orai1/3 heteromultimers). Although activated by fundamentally different mechanisms, CRAC channels, like ARC and LRC channels, require stromal interacting molecule 1 (STIM1). The role of endoplasmic reticulum–resident STIM1 (ER-STIM1) in CRAC channel activation is widely accepted. Although ER-STIM1 is necessary and sufficient for LRC channel activation in vascular smooth muscle cells (VSMCs), the minor pool of STIM1 located at the plasma membrane (PM-STIM1) is necessary for ARC channel activation in HEK293 cells. To determine whether ARC and LRC conductances are mediated by the same or different populations of STIM1, Orai1, and Orai3 proteins, we used whole-cell and perforated patch-clamp recording to compare AA- and LTC4-activated currents in VSMCs and HEK293 cells. We found that both cell types show indistinguishable nonadditive LTC4- and AA-activated currents that require both Orai1 and Orai3, suggesting that both conductances are mediated by the same channel. Experiments using a nonmetabolizable form of AA or an inhibitor of 5-lipooxygenase suggested that ARC and LRC currents in both cell types could be activated by either LTC4 or AA, with LTC4 being more potent. Although PM-STIM1 was required for current activation by LTC4 and AA under whole-cell patch-clamp recordings in both cell types, ER-STIM1 was sufficient with perforated patch recordings. These results demonstrate that ARC and LRC currents are mediated by the same cellular populations of STIM1, Orai1, and Orai3, and suggest a complex role for both ER-STIM1 and PM-STIM1 in regulating these store-independent Orai1/3 channels.

APA, Harvard, Vancouver, ISO, and other styles

16

Spinelli, Amy M., and Mohamed Trebak. "Orai channel-mediated Ca2+ signals in vascular and airway smooth muscle." American Journal of Physiology-Cell Physiology 310, no. 6 (March 15, 2016): C402—C413. http://dx.doi.org/10.1152/ajpcell.00355.2015.

Full text

Abstract:

Orai (Orai1, Orai2, and Orai3) proteins form a family of highly Ca2+-selective plasma membrane channels that are regulated by stromal-interacting molecules (STIM1 and STIM2); STIM proteins are Ca2+ sensors located in the membrane of the endoplasmic reticulum. STIM and Orai proteins are expressed in vascular and airway smooth muscle and constitute the molecular components of the ubiquitous store-operated Ca2+ entry pathway that mediate the Ca2+ release-activated Ca2+ current. STIM/Orai proteins also encode store-independent Ca2+ entry pathways in smooth muscle. Altered expression and function of STIM/Orai proteins have been linked to vascular and airway pathologies, including restenosis, hypertension, and atopic asthma. In this review we discuss our current understanding of Orai proteins and the store-dependent and -independent signaling pathways mediated by these proteins in vascular and airway smooth muscle. We also discuss the current studies linking altered expression and function of Orai proteins with smooth muscle-related pathologies.

APA, Harvard, Vancouver, ISO, and other styles

17

Sánchez-Collado, José, José J. López, and Juan A. Rosado. "The Orai1-AC8 Interplay: How Breast Cancer Cells Escape from Orai1 Channel Inactivation." Cells 10, no. 6 (May 25, 2021): 1308. http://dx.doi.org/10.3390/cells10061308.

Full text

Abstract:

The interplay between the Ca2+-sensitive adenylyl cyclase 8 (AC8) and Orai1 channels plays an important role both in the activation of the cAMP/PKA signaling and the modulation of Orai1-dependent Ca2+ signals. AC8 interacts with a N-terminal region that is exclusive to the Orai1 long variant, Orai1α. The interaction between both proteins allows the Ca2+ that enters the cell through Orai1α to activate the generation of cAMP by AC8. Subsequent PKA activation results in Orai1α inactivation by phosphorylation at serine-34, thus shaping Orai1-mediated cellular functions. In breast cancer cells, AC8 plays a relevant role supporting a variety of cancer hallmarks, including proliferation and migration. Breast cancer cells overexpress AC8, which shifts the AC8-Orai1 stoichiometry in favor of the former and leads to the impairment of PKA-dependent Orai1α inactivation. This mechanism contributes to the enhanced SOCE observed in triple-negative breast cancer cells. This review summarizes the functional interaction between AC8 and Orai1α in normal and breast cancer cells and its relevance for different cancer features.

APA, Harvard, Vancouver, ISO, and other styles

18

Kar, Pulak, Yu-Ping Lin, Rajesh Bhardwaj, Charles J. Tucker, Gary S. Bird, Matthias A. Hediger, Carina Monico, Nader Amin, and Anant B. Parekh. "The N terminus of Orai1 couples to the AKAP79 signaling complex to drive NFAT1 activation by local Ca2+ entry." Proceedings of the National Academy of Sciences 118, no. 19 (May 3, 2021): e2012908118. http://dx.doi.org/10.1073/pnas.2012908118.

Full text

Abstract:

To avoid conflicting and deleterious outcomes, eukaryotic cells often confine second messengers to spatially restricted subcompartments. The smallest signaling unit is the Ca2+ nanodomain, which forms when Ca2+ channels open. Ca2+ nanodomains arising from store-operated Orai1 Ca2+ channels stimulate the protein phosphatase calcineurin to activate the transcription factor nuclear factor of activated T cells (NFAT). Here, we show that NFAT1 tethered directly to the scaffolding protein AKAP79 (A-kinase anchoring protein 79) is activated by local Ca2+ entry, providing a mechanism to selectively recruit a transcription factor. We identify the region on the N terminus of Orai1 that interacts with AKAP79 and demonstrate that this site is essential for physiological excitation–transcription coupling. NMR structural analysis of the AKAP binding domain reveals a compact shape with several proline-driven turns. Orai2 and Orai3, isoforms of Orai1, lack this region and therefore are less able to engage AKAP79 and activate NFAT. A shorter, naturally occurring Orai1 protein that arises from alternative translation initiation also lacks the AKAP79-interaction site and fails to activate NFAT1. Interfering with Orai1–AKAP79 interaction suppresses cytokine production, leaving other Ca2+ channel functions intact. Our results reveal the mechanistic basis for how a subtype of a widely expressed Ca2+ channel is able to activate a vital transcription pathway and identify an approach for generation of immunosuppressant drugs.

APA, Harvard, Vancouver, ISO, and other styles

19

Kilch, Tatiana, Dalia Alansary, Kathrin Dörr, Christine Peinelt, and Barbara Niemeyer. "Modification of glycan chains as a mechanism to regulate SOCE (P1156)." Journal of Immunology 190, no. 1_Supplement (May 1, 2013): 190.3. http://dx.doi.org/10.4049/jimmunol.190.supp.190.3.

Full text

Abstract:

Abstract T cell receptor mediated activation requires the influx of calcium from the extracellular medium. Here, the endoplasmic reticulum (ER) Ca2+ sensor STIM1 reports the decrease in luminal [Ca2+] caused by IP3 receptor activity, clusters and directly binds and activates Orai1 channels, providing Ca2+ influx (store-operated, SOCE) essential for immune cell activation. We are interested in the regulation and modulation of this pathway. Because alterations of glycosylation of several T cell epitopes are implicated in immunsenescence and cancer, we investigated N-glycosylation site mutants of STIM1 and Orai1 and describe a STIM1 mutant (NN/DQ) which results in a strong gain-of-function. The mutant leads to a significant increase in the number of active channels despite a concomitant decrease in Orai1 protein, uncovering a novel CRAC channel regulatory mechanism. While we have already shown that the STIM1 mutant phenotype is mainly due to an increased rate of STIM1 oligomerization, we now investigate the specificity of this effect by analysing Ca2+ influx mediated by Orai2 and Orai3 and also started delineating the mechanism of Orai1 protein reduction. In addition, we are investigating the complex glycosylation pattern of primary human T cells and its effect on SOCE. Our findings indicate that the cell type specific repertoire of N-glycoslytransferases modifies the influence of the Orai1 glycan chains on Ca2+ influx.

APA, Harvard, Vancouver, ISO, and other styles

20

Arora, Samriddhi, Jyoti Tanwar, Nutan Sharma, Suman Saurav, and Rajender K. Motiani. "Orai3 Regulates Pancreatic Cancer Metastasis by Encoding a Functional Store Operated Calcium Entry Channel." Cancers 13, no. 23 (November 25, 2021): 5937. http://dx.doi.org/10.3390/cancers13235937.

Full text

Abstract:

Store operated Ca2+ entry (SOCE) mediated by Orai1/2/3 channels is a highly regulated and ubiquitous Ca2+ influx pathway. Although the role of Orai1 channels is well studied, the significance of Orai2/3 channels is still emerging in nature. In this study, we performed extensive bioinformatic analysis of publicly available datasets and observed that Orai3 expression is inversely associated with the mean survival time of PC patients. Orai3 expression analysis in a battery of PC cell lines corroborated its differential expression profile. We then carried out thorough Ca2+ imaging experiments in six PC cell lines and found that Orai3 forms a functional SOCE channel in PC cells. Our in vitro functional assays show that Orai3 regulates PC cell cycle progression, apoptosis and migration. Most importantly, our in vivo xenograft studies demonstrate a critical role of Orai3 in PC tumor growth and secondary metastasis. Mechanistically, Orai3 controls G1 phase progression, matrix metalloproteinase expression and epithelial-mesenchymal transition in PC cells. Taken together, this study for the first-time reports that Orai3 drives aggressive phenotypes of PC cells, i.e., migration in vitro and metastasis in vivo. Considering that Orai3 overexpression leads to poor prognosis in PC patients, it appears to be a highly attractive therapeutic target.

APA, Harvard, Vancouver, ISO, and other styles

21

Ibrahim, Dakik, Vandier, Chautard, Paintaud, Mazurier, Lecomte, Guéguinou, and Raoul. "Expression Profiling of Calcium Channels and Calcium-Activated Potassium Channels in Colorectal Cancer." Cancers 11, no. 4 (April 19, 2019): 561. http://dx.doi.org/10.3390/cancers11040561.

Full text

Abstract:

Background: Colorectal cancer (CRC) is a highly devastating cancer. Ca2+-dependent channels are now considered key regulators of tumor progression. In this study, we aimed to investigate the association of non-voltage gated Ca2+ channels and Ca2+-dependent potassium channels (KCa) with CRC using the transcriptional profile of their genes. Methods: We selected a total of 35 genes covering KCa channels KCNN1–4, KCNMA1 and their subunits KCNMB1–4, endoplasmic reticulum (ER) calcium sensors STIM1 and STIM2, Ca2+ channels ORAI1–3 and the family of cation channels TRP (TRPC1–7, TRPA1, TRPV1/2,4–6 and TRPM1–8). We analyzed their expression in two public CRC datasets from The Cancer Genome Atlas (TCGA) and GSE39582. Results: KCNN4 and TRPM2 were induced while KCNMA1 and TRPM6 were downregulated in tumor tissues comparing to normal tissues. In proximal tumors, STIM2 and KCNN2 were upregulated while ORAI2 and TRPM6 were downregulated. ORAI1 decreased in lymph node metastatic tumors. TRPC1 and ORAI3 predicted poor prognosis in CRC patients. Moreover, we found that ORAI3/ORAI1 ratio is increased in CRC progression and predicted poor prognosis. Conclusions: KCa and Ca2+ channels could be important contributors to CRC initiation and progression. Our results provide new insights on KCa and Ca2+ channels remodeling in CRC.

APA, Harvard, Vancouver, ISO, and other styles

22

Zhou, Yandong, Robert M. Nwokonko, Xiangyu Cai, Natalia A. Loktionova, Raz Abdulqadir, Ping Xin, Barbara A. Niemeyer, Youjun Wang, Mohamed Trebak, and Donald L. Gill. "Cross-linking of Orai1 channels by STIM proteins." Proceedings of the National Academy of Sciences 115, no. 15 (March 26, 2018): E3398—E3407. http://dx.doi.org/10.1073/pnas.1720810115.

Full text

Abstract:

The transmembrane docking of endoplasmic reticulum (ER) Ca2+-sensing STIM proteins with plasma membrane (PM) Orai Ca2+ channels is a critical but poorly understood step in Ca2+ signal generation. STIM1 protein dimers unfold to expose a discrete STIM–Orai activating region (SOAR1) that tethers and activates Orai1 channels within discrete ER–PM junctions. We reveal that each monomer within the SOAR dimer interacts independently with single Orai1 subunits to mediate cross-linking between Orai1 channels. Superresolution imaging and mobility measured by fluorescence recovery after photobleaching reveal that SOAR dimer cross-linking leads to substantial Orai1 channel clustering, resulting in increased efficacy and cooperativity of Orai1 channel function. A concatenated SOAR1 heterodimer containing one monomer point mutated at its critical Orai1 binding residue (F394H), although fully activating Orai channels, is completely defective in cross-linking Orai1 channels. Importantly, the naturally occurring STIM2 variant, STIM2.1, has an eight-amino acid insert in its SOAR unit that renders it functionally identical to the F394H mutant in SOAR1. Contrary to earlier predictions, the SOAR1–SOAR2.1 heterodimer fully activates Orai1 channels but prevents cross-linking and clustering of channels. Interestingly, combined expression of full-length STIM1 with STIM2.1 in a 5:1 ratio causes suppression of sustained agonist-induced Ca2+ oscillations and protects cells from Ca2+ overload, resulting from high agonist-induced Ca2+ release. Thus, STIM2.1 exerts a powerful regulatory effect on signal generation likely through preventing Orai1 channel cross-linking. Overall, STIM-mediated cross-linking of Orai1 channels is a hitherto unrecognized functional paradigm that likely provides an organizational microenvironment within ER–PM junctions with important functional impact on Ca2+ signal generation.

APA, Harvard, Vancouver, ISO, and other styles

23

Azimi, Iman, Michael Milevskiy, Silke Chalmers, Kunsala Yapa, Mélanie Robitaille, Christopher Henry, Gregory Baillie, Erik Thompson, Sarah Roberts-Thomson, and Gregory Monteith. "ORAI1 and ORAI3 in Breast Cancer Molecular Subtypes and the Identification of ORAI3 as a Hypoxia Sensitive Gene and a Regulator of Hypoxia Responses." Cancers 11, no. 2 (February 11, 2019): 208. http://dx.doi.org/10.3390/cancers11020208.

Full text

Abstract:

The remodeling of specific calcium-permeable ion channels is a feature of some breast cancer subtypes. ORAI1 is a protein that forms a calcium-permeable ion channel responsible for store-operated calcium entry (SOCE) in a variety of cell types. ORAI3, a related isoform, is not a regulator of SOCE in most cell types. However, ORAI3 does control SOCE in many estrogen receptor-positive breast cancer cell lines, where it also controls proliferation. ORAI1 is a well-characterized regulator of the proliferation and migration of many basal breast cancer cells; however, the role of ORAI3 in these types of breast cancer cells remains unclear. Here, we sought to define ORAI1 and ORAI3 expression in breast cancer cell lines of different molecular subtypes and assess the potential role and regulation of ORAI3 in basal breast cancer cells. Our study demonstrates that elevated ORAI1 is a feature of basal-like breast cancers, while elevated ORAI3 is a feature of luminal breast cancers. Intriguingly, we found that ORAI3 is over-expressed in the mesenchymal subtype of triple-negative breast cancer. Given this, we assessed ORAI3 levels in the presence of two inducers of the mesenchymal phenotype, hypoxia and epidermal growth factor (EGF). Hypoxia induced ORAI3 levels in basal breast cancer cell lines through a pathway involving hypoxia-inducible factor-1 alpha (HIF1α. The silencing of ORAI3 attenuated hypoxia-associated phosphorylation of the EGF receptor (EGFR) and the expression of genes associated with cell migration and inflammatory/immune responses in the MDA-MB-468 model of basal breast cancer. Although elevated ORAI3 levels were not associated with survival; basal, estrogen receptor-negative and triple-negative breast cancers with high ORAI3 and low ORAI1 levels were associated with poorer clinical outcomes. This study defines ORAI3 as a potential fine-tuner for processes relevant to the progression of basal breast cancers.

APA, Harvard, Vancouver, ISO, and other styles

24

Segin, Sebastian, Michael Berlin, Christin Richter, Rebekka Medert, Veit Flockerzi, Paul Worley, Marc Freichel, and Juan E. Camacho Londoño. "Cardiomyocyte-Specific Deletion of Orai1 Reveals Its Protective Role in Angiotensin-II-Induced Pathological Cardiac Remodeling." Cells 9, no. 5 (April 28, 2020): 1092. http://dx.doi.org/10.3390/cells9051092.

Full text

Abstract:

Pathological cardiac remodeling correlates with chronic neurohumoral stimulation and abnormal Ca2+ signaling in cardiomyocytes. Store-operated calcium entry (SOCE) has been described in adult and neonatal murine cardiomyocytes, and Orai1 proteins act as crucial ion-conducting constituents of this calcium entry pathway that can be engaged not only by passive Ca2+ store depletion but also by neurohumoral stimuli such as angiotensin-II. In this study, we, therefore, analyzed the consequences of Orai1 deletion for cardiomyocyte hypertrophy in neonatal and adult cardiomyocytes as well as for other features of pathological cardiac remodeling including cardiac contractile function in vivo. Cellular hypertrophy induced by angiotensin-II in embryonic cardiomyocytes from Orai1-deficient mice was blunted in comparison to cells from litter-matched control mice. Due to lethality of mice with ubiquitous Orai1 deficiency and to selectively analyze the role of Orai1 in adult cardiomyocytes, we generated a cardiomyocyte-specific and temporally inducible Orai1 knockout mouse line (Orai1CM–KO). Analysis of cardiac contractility by pressure-volume loops under basal conditions and of cardiac histology did not reveal differences between Orai1CM–KO mice and controls. Moreover, deletion of Orai1 in cardiomyocytes in adult mice did not protect them from angiotensin-II-induced cardiac remodeling, but cardiomyocyte cross-sectional area and cardiac fibrosis were enhanced. These alterations in the absence of Orai1 go along with blunted angiotensin-II-induced upregulation of the expression of Myoz2 and a lack of rise in angiotensin-II-induced STIM1 and Orai3 expression. In contrast to embryonic cardiomyocytes, where Orai1 contributes to the development of cellular hypertrophy, the results obtained from deletion of Orai1 in the adult myocardium reveal a protective function of Orai1 against the development of angiotensin-II-induced cardiac remodeling, possibly involving signaling via Orai3/STIM1-calcineurin-NFAT related pathways.

APA, Harvard, Vancouver, ISO, and other styles

25

Vashisht, Ayushi, Mohamed Trebak, and Rajender K. Motiani. "STIM and Orai proteins as novel targets for cancer therapy. A Review in the Theme: Cell and Molecular Processes in Cancer Metastasis." American Journal of Physiology-Cell Physiology 309, no. 7 (October 1, 2015): C457—C469. http://dx.doi.org/10.1152/ajpcell.00064.2015.

Full text

Abstract:

Calcium (Ca2+) regulates a plethora of cellular functions including hallmarks of cancer development such as cell cycle progression and cellular migration. Receptor-regulated calcium rise in nonexcitable cells occurs through store-dependent as well as store-independent Ca2+ entry pathways. Stromal interaction molecules (STIM) and Orai proteins have been identified as critical constituents of both these Ca2+ influx pathways. STIMs and Orais have emerged as targets for cancer therapeutics as their altered expression and function have been shown to contribute to tumorigenesis. Recent data demonstrate that they play a vital role in development and metastasis of a variety of tumor types including breast, prostate, cervical, colorectal, brain, and skin tumors. In this review, we will retrospect the data supporting a key role for STIM1, STIM2, Orai1, and Orai3 proteins in tumorigenesis and discuss the potential of targeting these proteins for cancer therapy.

APA, Harvard, Vancouver, ISO, and other styles

26

Kappel, Sven, Tatiana Kilch, Roland Baur, Martin Lochner, and Christine Peinelt. "The Number and Position of Orai3 Units within Heteromeric Store-Operated Ca2+ Channels Alter the Pharmacology of ICRAC." International Journal of Molecular Sciences 21, no. 7 (April 2, 2020): 2458. http://dx.doi.org/10.3390/ijms21072458.

Full text

Abstract:

Store-operated heteromeric Orai1/Orai3 channels have been discussed in the context of aging, cancer, and immune cell differentiation. In contrast to homomeric Orai1 channels, they exhibit a different pharmacology upon application of reactive oxygen species (ROS) or 2-aminoethoxydiphenyl borate (2-APB) in various cell types. In endogenous cells, subunit composition and arrangement may vary and cannot be defined precisely. In this study, we used patch-clamp electrophysiology to investigate the 2-APB profile of store-operated and store-independent homomeric Orai1 and heteromeric Orai1/Orai3 concatenated channels with defined subunit compositions. As has been shown previous, one or more Orai3 subunit(s) within the channel result(s) in decreased Ca2+ release activated Ca2+ current (ICRAC). Upon application of 50 µM 2-APB, channels with two or more Orai3 subunits exhibit large outward currents and can be activated by 2-APB independent from storedepletion and/or the presence of STIM1. The number and position of Orai3 subunits within the heteromeric store-operated channel change ion conductivity of 2-APB-activated outward current. Compared to homomeric Orai1 channels, one Orai3 subunit within the channel does not alter 2-APB pharmacology. None of the concatenated channel constructs were able to exactly simulate the complex 2-APB pharmacology observed in prostate cancer cells. However, 2-APB profiles of prostate cancer cells are similar to those of concatenated channels with Orai3 subunit(s). Considering the presented and previous results, this indicates that distinct subtypes of heteromeric SOCE channels may be selectively activated or blocked. In the future, targeting distinct heteromeric SOCE channel subtypes may be the key to tailored SOCE-based therapies.

APA, Harvard, Vancouver, ISO, and other styles

27

Robitaille, Mélanie, Shao Ming Chan, Amelia A. Peters, Limin Dai, Choon Leng So, Alice H. L. Bong, Francisco Sadras, Sarah J. Roberts-Thomson, and Gregory R. Monteith. "ORAI1-Regulated Gene Expression in Breast Cancer Cells: Roles for STIM1 Binding, Calcium Influx and Transcription Factor Translocation." International Journal of Molecular Sciences 23, no. 11 (May 24, 2022): 5867. http://dx.doi.org/10.3390/ijms23115867.

Full text

Abstract:

A remodeling of calcium homeostasis, including calcium influx via store-operated calcium entry (SOCE), is a feature of breast cancers. SOCE is critical to maintain calcium balance in the endoplasmic reticulum calcium store and is an important mechanism for calcium signaling in a variety of cell types, including breast cancer cells. The canonical mechanism of SOCE is stromal interacting molecule 1 (STIM1)-mediated activation of ORAI. Elevated ORAI1 expression is a feature of basal breast cancer cells. However, the role of ORAI1 in the regulation of transcription in breast cancer cells of the basal molecular subtype is still unclear. Using CRISPR-Cas9 gene editing, ORAI1 protein expression was disrupted in MDA-MB-231 and MDA-MB-468 basal breast cancer cells. The ORAI1 wild-type and mutants were reintroduced into ORAI1 knockout cells to study the role of ORAI1 in gene transcriptional regulation. In the absence of calcium store depletion, ORAI1 regulated PTGS2 in MDA-MB-231 cells, and this was dependent on ORAI1 pore function and STIM1 binding. The activation of SOCE by thapsigargin resulted in ORAI1-dependent increases in IL6 transcription in MDA-MB-468 cells; this was also dependent on ORAI1 pore function and STIM1 binding and was associated with the translocation of NFAT1. Given the upregulation of ORAI1 in basal breast cancer cells, our results provide further evidence that ORAI1 may contribute to cancer progression through regulation of gene expression.

APA, Harvard, Vancouver, ISO, and other styles

28

DeHaven, Wayne I., Jeremy T. Smyth, Rebecca R. Boyles, and James W. Putney. "Calcium Inhibition and Calcium Potentiation of Orai1, Orai2, and Orai3 Calcium Release-activated Calcium Channels." Journal of Biological Chemistry 282, no. 24 (April 23, 2007): 17548–56. http://dx.doi.org/10.1074/jbc.m611374200.

Full text

APA, Harvard, Vancouver, ISO, and other styles

29

Perni, Stefano, Joseph L. Dynes, Andriy V. Yeromin, Michael D. Cahalan, and Clara Franzini-Armstrong. "Nanoscale patterning of STIM1 and Orai1 during store-operated Ca2+ entry." Proceedings of the National Academy of Sciences 112, no. 40 (September 8, 2015): E5533—E5542. http://dx.doi.org/10.1073/pnas.1515606112.

Full text

Abstract:

Stromal interacting molecule (STIM) and Orai proteins constitute the core machinery of store-operated calcium entry. We used transmission and freeze–fracture electron microscopy to visualize STIM1 and Orai1 at endoplasmic reticulum (ER)–plasma membrane (PM) junctions in HEK 293 cells. Compared with control cells, thin sections of STIM1-transfected cells possessed far more ER elements, which took the form of complex stackable cisternae and labyrinthine structures adjoining the PM at junctional couplings (JCs). JC formation required STIM1 expression but not store depletion, induced here by thapsigargin (TG). Extended molecules, indicative of STIM1, decorated the cytoplasmic surface of ER, bridged a 12-nm ER-PM gap, and showed clear rearrangement into small clusters following TG treatment. Freeze–fracture replicas of the PM of Orai1-transfected cells showed extensive domains packed with characteristic “particles”; TG treatment led to aggregation of these particles into sharply delimited “puncta” positioned upon raised membrane subdomains. The size and spacing of Orai1 channels were consistent with the Orai crystal structure, and stoichiometry was unchanged by store depletion, coexpression with STIM1, or an Orai1 mutation (L273D) affecting STIM1 association. Although the arrangement of Orai1 channels in puncta was substantially unstructured, a portion of channels were spaced at ∼15 nm. Monte Carlo analysis supported a nonrandom distribution for a portion of channels spaced at ∼15 nm. These images offer dramatic, direct views of STIM1 aggregation and Orai1 clustering in store-depleted cells and provide evidence for the interaction of a single Orai1 channel with small clusters of STIM1 molecules.

APA, Harvard, Vancouver, ISO, and other styles

30

Zhu, D., R. He, W. Yu, C. Li, H. Cheng, B. Zhu, and J. Yan. "ORAI3 contributes to hypoxia-inducible factor 1/2α-sensitive colon cell migration." Physiology International 108, no. 2 (July 9, 2021): 221–37. http://dx.doi.org/10.1556/2060.2021.00137.

Full text

Abstract:

AbstractBackgroundHypoxia is a pivotal initiator of tumor angiogenesis and growth through the stabilization of hypoxia-inducible factors (HIFs). This study set out to examine the involvement of HIF-1α and HIF-2α in colon cancer and ascertained whether ORAI3 was involved in the pathway.Materials and methodsPatients and murine models as well as human colorectal adenocarcinoma tumor (CW2) cells were included to examine the levels of ORAI1/3 and HIF-1/2α levels. Calcium imaging was utilized to ascertain the activity of calcium channel. Scratch assay was used to assess the migration capacity of the cells.ResultsTumors from murine colon cancer xenograft models and patients with colon cancer displayed high ORAI1/3 and HIF-1/2α levels. Hypoxia treatment, mimicking the tumor microenvironment in vitro, increased ORAI1/3 and HIF-1/2α expression as well as store-operated Ca2+ entry (SOCE). Of note is that HIF-1/2α silencing decreased SOCE, and HIF-1/2α overexpression facilitated SOCE. Furthermore, ORAI3 rather than ORAI1 expression was inhibited by HIF-1/2α silencing while increased by ML228. Luciferase assay also confirmed that ORAI3 was elevated in the presence of ML228, indicating the linkage between HIF-1/2α and ORAI3. Additionally, colony-forming potential and cell migration capacity were decreased in siHIF-1α and siHIF-2α as well as siORAI3 cells, and the facilitating effect of ML228 on cell migration and colony-forming potential was also decreased in siORAI3 CW-2 cells, which points out the importance of ORAI3 in HIF1/2α pathway.ConclusionOur findings allow to conclude that both HIF-1α and HIF-2α facilitate ORAI3 expression, hence enhancing colon cancer progression.

APA, Harvard, Vancouver, ISO, and other styles

31

Qin, Ying, Qinggang Meng, Qunhua Wang, Mingzhu Wu, Yan Fang, Chengcheng Tu, Xinyang Hu, Bing Shen, Hongbo Chen, and Xiaohong Xu. "Pregnancy-Specific Glycoprotein 9 Enhances Store-Operated Calcium Entry and Nitric Oxide Release in Human Umbilical Vein Endothelial Cells." Diagnostics 13, no. 6 (March 16, 2023): 1134. http://dx.doi.org/10.3390/diagnostics13061134.

Full text

Abstract:

We explored changes in pregnancy-specific glycoprotein 9 (PSG9) levels in the serum of patients with preeclampsia and the effects and underlying mechanisms of PSG9 effects on calcium (Ca2+) homeostasis and nitric oxide (NO) release in human umbilical vein endothelial cells (HUVECs). Western blotting was used to detect protein expression levels, and an NO fluorescence probe was used to examine NO production. Intracellular Ca2+ concentrations were measured using a Ca2+-sensitive fluorescent dye under a fluorescence microscope. Compared with those in healthy pregnant women, serum PSG9 levels were significantly decreased in patients with preeclampsia. PSG9 (0.1 μg/mL) treatment of HUVECs significantly enhanced the expression levels of store-operated calcium entry (SOCE) channel proteins Orai1 and Orai2, but not Orai3, and of endothelial nitric oxide synthase (eNOS) and NO production. Pretreatment with an inhibitor of SOCE (BTP2) abolished PSG9-enhanced Orai1, Orai2, and eNOS expression levels and NO production in HUVECs. The mechanisms underlying SOCE that were PSG9 enhanced in HUVECs appear to involve the Ca2+/eNOS/NO signaling pathway. These findings suggest that serum PSG9 levels may be a potential biomarker for monitoring the occurrence or development of preeclampsia in pregnancy and that PSG9 may be a potential therapeutic target for the treatment of preeclampsia.

APA, Harvard, Vancouver, ISO, and other styles

32

Bokhobza, Alexandre, Nathalie Ziental-Gelus, Laurent Allart, Oksana Iamshanova, and Fabien Vanden Abeele. "Impact of SOCE Abolition by ORAI1 Knockout on the Proliferation, Adhesion, and Migration of HEK-293 Cells." Cells 10, no. 11 (November 4, 2021): 3016. http://dx.doi.org/10.3390/cells10113016.

Full text

Abstract:

Store-operated calcium entry (SOCE) provided through channels formed by ORAI proteins is a major regulator of several cellular processes. In immune cells, it controls fundamental processes such as proliferation, cell adhesion, and migration, while in cancer, SOCE and ORAI1 gene expression are dysregulated and lead to abnormal migration and/or cell proliferation. In the present study, we used the CRISPR/Cas9 technique to delete the ORAI1 gene and to identify its role in proliferative and migrative properties of the model cell line HEK-293. We showed that ORAI1 deletion greatly reduced SOCE. Thereby, we found that this decrease and the absence of ORAI1 protein did not affect HEK-293 proliferation. In addition, we determined that ORAI1 suppression did not affect adhesive properties but had a limited impact on HEK-293 migration. Overall, we showed that ORAI1 and SOCE are largely dispensable for cellular proliferation, migration, and cellular adhesion of HEK-293 cells. Thus, despite its importance in providing Ca2+ entry in non-excitable cells, our results indicate that the lack of SOCE does not deeply impact HEK-293 cells. This finding suggests the existence of compensatory mechanism enabling the maintenance of their physiological function.

APA, Harvard, Vancouver, ISO, and other styles

33

Schindl, R., I. Frischauf, J. Bergsmann, M. Muik, I. Derler, B. Lackner, K. Groschner, and C. Romanin. "Plasticity in Ca2+ selectivity of Orai1/Orai3 heteromeric channel." Proceedings of the National Academy of Sciences 106, no. 46 (November 3, 2009): 19623–28. http://dx.doi.org/10.1073/pnas.0907714106.

Full text

APA, Harvard, Vancouver, ISO, and other styles

34

Yeromin, Andriy V., Olga Safrina, and Michael D. Cahalan. "Ph Dependence of Orai1 and Orai3 Store-Operated Current." Biophysical Journal 106, no. 2 (January 2014): 315a. http://dx.doi.org/10.1016/j.bpj.2013.11.1824.

Full text

APA, Harvard, Vancouver, ISO, and other styles

35

Sanchez-Collado, Lopez, Jardin, Camello, Falcon, Regodon, Salido, Smani, and Rosado. "Adenylyl Cyclase Type 8 Overexpression Impairs Phosphorylation-Dependent Orai1 Inactivation and Promotes Migration in MDA-MB-231 Breast Cancer Cells." Cancers 11, no. 11 (October 23, 2019): 1624. http://dx.doi.org/10.3390/cancers11111624.

Full text

Abstract:

Orai1 plays a major role in store-operated Ca2+ entry (SOCE) in triple-negative breast cancer (TNBC) cells. This channel is inactivated via different mechanisms, including protein kinase C (PKC) and protein kinase A (PKA)-dependent phosphorylation at Ser-27 and Ser-30 or Ser-34, respectively, which shapes the Ca2+ responses to agonists. The Ca2+ calmodulin-activated adenylyl cyclase type 8 (AC8) was reported to interact directly with Orai1, thus mediating a dynamic interplay between the Ca2+- and cyclic adenosine monophosphate (cAMP)-dependent signaling pathways. Here, we show that the breast cancer cell lines MCF7 and MDA-MB-231 exhibit enhanced expression of Orai1 and AC8 as compared to the non-tumoral breast epithelial MCF10A cell line. In these cells, AC8 interacts with the Orai1α variant in a manner that is not regulated by Orai1 phosphorylation. AC8 knockdown in MDA-MB-231 cells, using two different small interfering RNAs (siRNAs), attenuates thapsigargin (TG)-induced Ca2+ entry and also Ca2+ influx mediated by co-expression of Orai1 and the Orai1-activating small fragment (OASF) of STIM1 (stromal interaction molecule-1). Conversely, AC8 overexpression enhances SOCE, as well as Ca2+ entry, in cells co-expressing Orai1 and OASF. In MDA-MB-231 cells, we found that AC8 overexpression reduces the Orai1 phosphoserine content, thus suggesting that AC8 interferes with Orai1 serine phosphorylation, which takes place at residues located in the AC8-binding site. Consistent with this, the subset of Orai1 associated with AC8 in naïve MDA-MB-231 cells is not phosphorylated in serine residues in contrast to the AC8-independent Orai1 subset. AC8 expression knockdown attenuates migration of MCF7 and MDA-MB-231 cells, while this maneuver has no effect in the MCF10A cell line, which is likely attributed to the low expression of AC8 in these cells. We found that AC8 is required for FAK (focal adhesion kinase) phosphorylation in MDA-MB-231 cells, which might explain its role in cell migration. Finally, we found that AC8 is required for TNBC cell proliferation. These findings indicate that overexpression of AC8 in breast cancer MDA-MB-231 cells impairs the phosphorylation-dependent Orai1 inactivation, a mechanism that might support the enhanced ability of these cells to migrate.

APA, Harvard, Vancouver, ISO, and other styles

36

Wu, Minnie M., Elizabeth D. Covington, and Richard S. Lewis. "Single-molecule analysis of diffusion and trapping of STIM1 and Orai1 at endoplasmic reticulum–plasma membrane junctions." Molecular Biology of the Cell 25, no. 22 (November 5, 2014): 3672–85. http://dx.doi.org/10.1091/mbc.e14-06-1107.

Full text

Abstract:

Following endoplasmic reticulum (ER) Ca2+ depletion, STIM1 and Orai1 complexes assemble autonomously at ER–plasma membrane (PM) junctions to trigger store-operated Ca2+ influx. One hypothesis to explain this process is a diffusion trap in which activated STIM1 diffusing in the ER becomes trapped at junctions through interactions with the PM, and STIM1 then traps Orai1 in the PM through binding of its calcium release-activated calcium activation domain. We tested this model by analyzing STIM1 and Orai1 diffusion using single-particle tracking, photoactivation of protein ensembles, and Monte Carlo simulations. In resting cells, STIM1 diffusion is Brownian, while Orai1 is slightly subdiffusive. After store depletion, both proteins slow to the same speeds, consistent with complex formation, and are confined to a corral similar in size to ER–PM junctions. While the escape probability at high STIM:Orai expression ratios is <1%, it is significantly increased by reducing the affinity of STIM1 for Orai1 or by expressing the two proteins at comparable levels. Our results provide direct evidence that STIM-Orai complexes are trapped by their physical connections across the junctional gap, but also reveal that the complexes are surprisingly dynamic, suggesting that readily reversible binding reactions generate free STIM1 and Orai1, which engage in constant diffusional exchange with extrajunctional pools.

APA, Harvard, Vancouver, ISO, and other styles

37

Cai, Xiangyu, Yandong Zhou, Xianming Wang, Natalia Loktionova, Robert Nwokonko, Mohamed Trebak, and Donald Gill. "Orai1 Concatemers Reveal a Hexameric Orai1 Channel Assembly." Biophysical Journal 110, no. 3 (February 2016): 264a—265a. http://dx.doi.org/10.1016/j.bpj.2015.11.1444.

Full text

APA, Harvard, Vancouver, ISO, and other styles

38

Schaff, Ulrich Y., Neha Dixit, Emily Procyk, Itsukyo Yamayoshi, Tiffany Tse, and Scott I. Simon. "Orai1 regulates intracellular calcium, arrest, and shape polarization during neutrophil recruitment in shear flow." Blood 115, no. 3 (January 21, 2010): 657–66. http://dx.doi.org/10.1182/blood-2009-05-224659.

Full text

Abstract:

Abstract Orai1 was reported to function as a calcium channel subunit that facilitates store operated calcium entry (SOCE) in T cells and is necessary for formation of the immune synapse. We reasoned that SOCE via Orai1 might regulate PMNs activation during recruitment to inflamed endothelium. Orai1 function was assessed by real-time imaging of calcium transients as PMNs were stimulated to roll, arrest, and migrate on E-selectin and ICAM-1 in shear flow. Calcium entry was significantly reduced when Orai1 function was impaired by heterozygous knockout in a mouse model or by siRNA knockdown in HL-60 cells. Reduced Orai-1 expression correlated with the delayed onset of arrest and reduced ability to transition to a polarized migratory phenotype. Inhibition of SOCE by treatment with 2-APB, or blocking phospholipase C (PLC) mediated calcium store release with U73122, abrogated formyl peptide induced calcium elevation, and delayed subsequent cell arrest and polarization. These results suggest that calcium entry via Orai1 is the predominant SOCE that cooperates with cytoplasmic calcium store release in coordinating integrin-dependent PMN arrest and migration in the acute response to inflammation.

APA, Harvard, Vancouver, ISO, and other styles

39

Rana, Anshul, Michelle Yen, Amir Masoud Sadaghiani, Seth Malmersjö, Chan Young Park, Ricardo E. Dolmetsch, and Richard S. Lewis. "Alternative splicing converts STIM2 from an activator to an inhibitor of store-operated calcium channels." Journal of Cell Biology 209, no. 5 (June 1, 2015): 653–70. http://dx.doi.org/10.1083/jcb.201412060.

Full text

Abstract:

Store-operated calcium entry (SOCE) regulates a wide variety of essential cellular functions. SOCE is mediated by STIM1 and STIM2, which sense depletion of ER Ca2+ stores and activate Orai channels in the plasma membrane. Although the amplitude and dynamics of SOCE are considered important determinants of Ca2+-dependent responses, the underlying modulatory mechanisms are unclear. In this paper, we identify STIM2β, a highly conserved alternatively spliced isoform of STIM2, which, in contrast to all known STIM isoforms, is a potent inhibitor of SOCE. Although STIM2β does not by itself strongly bind Orai1, it is recruited to Orai1 channels by forming heterodimers with other STIM isoforms. Analysis of STIM2β mutants and Orai1-STIM2β chimeras suggested that it actively inhibits SOCE through a sequence-specific allosteric interaction with Orai1. Our results reveal a previously unrecognized functional flexibility in the STIM protein family by which alternative splicing creates negative and positive regulators of SOCE to shape the amplitude and dynamics of Ca2+ signals.

APA, Harvard, Vancouver, ISO, and other styles

40

Alansary, Dalia, Ivan Bogeski, and Barbara A. Niemeyer. "Facilitation of Orai3 targeting and store-operated function by Orai1." Biochimica et Biophysica Acta (BBA) - Molecular Cell Research 1853, no. 7 (July 2015): 1541–50. http://dx.doi.org/10.1016/j.bbamcr.2015.03.007.

Full text

APA, Harvard, Vancouver, ISO, and other styles

41

Rychkov, Grigori Y., Fiona H. Zhou, Melissa K. Adams, Stuart M. Brierley, Linlin Ma, and Greg J. Barritt. "Orai1‐ and Orai2‐, but not Orai3‐mediated I _CRAC is regulated by intracellular pH." Journal of Physiology 600, no. 3 (December 25, 2021): 623–43. http://dx.doi.org/10.1113/jp282502.

Full text

APA, Harvard, Vancouver, ISO, and other styles

42

Ramanagoudr-Bhojappa, Ramanagouda, Yong Miao, and Monika Vig. "High affinity associations with α-SNAP enable calcium entry via Orai1 channels." PLOS ONE 16, no. 10 (October 15, 2021): e0258670. http://dx.doi.org/10.1371/journal.pone.0258670.

Full text

Abstract:

Molecular steps that activate store-operated calcium entry (SOCE) via Orai channel supramolecular complex remain incompletely defined. We have earlier shown that α-SNAP regulates the on-site functional assembly and calcium selectivity of Orai1 channels. Here we investigate the molecular basis of its association with Orai, Stim and find that the affinity of α-SNAP for Orai and Stim is substantially higher than previously reported affinities between Stim and Orai sub-domains. α-SNAP binds the coiled-coil 3 (CC3) sub-domain of Stim1. Mutations of Tryptophan 430 in Stim1-CC3 disrupted α-SNAP association and SOCE, demonstrating a novel α-SNAP dependent function for this crucial subdomain. Further, α-SNAP binds the hinge region near the C-terminus of Orai1 and an additional broad region near the N-terminus and Valine 262 and Leucine 74 were necessary for these respective interactions, but not Orai, Stim co-clustering. Thus, high affinity interactions with α-SNAP are necessary for imparting functionality to Stim, Orai clusters and induction of SOCE.

APA, Harvard, Vancouver, ISO, and other styles

43

Quintana, Ariel, Vangipurapu Rajanikanth, Suzette Farber-Katz, Aparna Gudlur, Chen Zhang, Ji Jing, Yubin Zhou, Anjana Rao, and Patrick G. Hogan. "TMEM110 regulates the maintenance and remodeling of mammalian ER–plasma membrane junctions competent for STIM–ORAI signaling." Proceedings of the National Academy of Sciences 112, no. 51 (December 7, 2015): E7083—E7092. http://dx.doi.org/10.1073/pnas.1521924112.

Full text

Abstract:

The stromal interaction molecule (STIM)–ORAI calcium release-activated calcium modulator (ORAI) pathway controls store-dependent calcium entry, a major mechanism of physiological calcium signaling in mammalian cells. The core elements of the pathway are the regulatory protein STIM1, located in the endoplasmic reticulum (ER) membrane, the calcium channel ORAI1 in the plasma membrane, and sites of close contact between the ER and the plasma membrane that permit the two proteins to interact. Research on calcium signaling has centered on STIM1, ORAI1, and a few proteins that directly modulate STIM–ORAI function. However, little is known about proteins that organize ER–plasma membrane junctions for STIM–ORAI-dependent calcium signaling. Here, we report that an ER-resident membrane protein identified in a previous genome-wide RNAi screen, transmembrane protein 110 (TMEM110), regulates the long-term maintenance of ER–plasma membrane junctions and the short-term physiological remodeling of the junctions during store-dependent calcium signaling.

APA, Harvard, Vancouver, ISO, and other styles

44

Shen, Yihan, Nagendra Babu Thillaiappan, and Colin W. Taylor. "The store-operated Ca2+ entry complex comprises a small cluster of STIM1 associated with one Orai1 channel." Proceedings of the National Academy of Sciences 118, no. 10 (March 1, 2021): e2010789118. http://dx.doi.org/10.1073/pnas.2010789118.

Full text

Abstract:

Increases in cytosolic Ca2+ concentration regulate diverse cellular activities and are usually evoked by opening of Ca2+ channels in intracellular Ca2+ stores and the plasma membrane (PM). For the many signals that evoke formation of inositol 1,4,5-trisphosphate (IP3), IP3 receptors coordinate the contributions of these two Ca2+ sources by mediating Ca2+ release from the endoplasmic reticulum (ER). Loss of Ca2+ from the ER then activates store-operated Ca2+ entry (SOCE) by causing dimers of STIM1 to cluster and unfurl cytosolic domains that interact with the PM Ca2+ channel, Orai1, causing its pore to open. The relative concentrations of STIM1 and Orai1 are important, but most analyses of their interactions use overexpressed proteins that perturb the stoichiometry. We tagged endogenous STIM1 with EGFP using CRISPR/Cas9. SOCE evoked by loss of ER Ca2+ was unaffected by the tag. Step-photobleaching analysis of cells with empty Ca2+ stores revealed an average of 14.5 STIM1 molecules within each sub-PM punctum. The fluorescence intensity distributions of immunostained Orai1 puncta were minimally affected by store depletion, and similar for Orai1 colocalized with STIM1 puncta or remote from them. We conclude that each native SOCE complex is likely to include only a few STIM1 dimers associated with a single Orai1 channel. Our results, demonstrating that STIM1 does not assemble clusters of interacting Orai channels, suggest mechanisms for digital regulation of SOCE by local depletion of the ER.

APA, Harvard, Vancouver, ISO, and other styles

45

Yeung, Priscilla S. W., Megumi Yamashita, Christopher E. Ing, Régis Pomès, Douglas M. Freymann, and Murali Prakriya. "Mapping the functional anatomy of Orai1 transmembrane domains for CRAC channel gating." Proceedings of the National Academy of Sciences 115, no. 22 (May 14, 2018): E5193—E5202. http://dx.doi.org/10.1073/pnas.1718373115.

Full text

Abstract:

Store-operated Orai1 channels are activated through a unique inside-out mechanism involving binding of the endoplasmic reticulum Ca2+ sensor STIM1 to cytoplasmic sites on Orai1. Although atomic-level details of Orai structure, including the pore and putative ligand binding domains, are resolved, how the gating signal is communicated to the pore and opens the gate is unknown. To address this issue, we used scanning mutagenesis to identify 15 residues in transmembrane domains (TMs) 1–4 whose perturbation activates Orai1 channels independently of STIM1. Cysteine accessibility analysis and molecular-dynamics simulations indicated that constitutive activation of the most robust variant, H134S, arises from a pore conformational change that opens a hydrophobic gate to augment pore hydration, similar to gating evoked by STIM1. Mutational analysis of this locus suggests that H134 acts as steric brake to stabilize the closed state of the channel. In addition, atomic packing analysis revealed distinct functional contacts between the TM1 pore helix and the surrounding TM2/3 helices, including one set mediated by a cluster of interdigitating hydrophobic residues and another by alternative ridges of polar and hydrophobic residues. Perturbing these contacts via mutagenesis destabilizes STIM1-mediated Orai1 channel gating, indicating that these bridges between TM1 and the surrounding TM2/3 ring are critical for conveying the gating signal to the pore. These findings help develop a framework for understanding the global conformational changes and allosteric interactions between topologically distinct domains that are essential for activation of Orai1 channels.

APA, Harvard, Vancouver, ISO, and other styles

46

Prabakaran, Sudhakaran. "Milk needs Orai1." Science Signaling 8, no. 377 (May 19, 2015): ec127-ec127. http://dx.doi.org/10.1126/scisignal.aac5307.

Full text

APA, Harvard, Vancouver, ISO, and other styles

47

Schindl, Rainer, Irene Frischauf, Judith Bergsmann, Martin Muik, Isabella Derler, and Christoph Romanin. "Heteromeric channel assembly of Orai1 and Orai3 exhibits altered Ca2+ selectivity." Biophysical Journal 96, no. 3 (February 2009): 559a—560a. http://dx.doi.org/10.1016/j.bpj.2008.12.3672.

Full text

APA, Harvard, Vancouver, ISO, and other styles

48

Hooper, Robert, Xuexin Zhang, Marie Webster, Christina Go, Joseph Kedra, Katie Marchbank, Donald L. Gill, Ashani T. Weeraratna, Mohamed Trebak, and Jonathan Soboloff. "Novel Protein Kinase C-Mediated Control of Orai1 Function in Invasive Melanoma." Molecular and Cellular Biology 35, no. 16 (June 8, 2015): 2790–98. http://dx.doi.org/10.1128/mcb.01500-14.

Full text

Abstract:

The incidence of malignant melanoma, a cancer of the melanocyte cell lineage, has nearly doubled in the past 20 years. Wnt5A, a key driver of melanoma invasiveness, induces Ca2+signals. To understand how store-operated calcium entry (SOCE) contributes to Wnt5A-induced malignancy in melanoma models, we examined the expression and function of STIM1 and Orai1 in patient-derived malignant melanoma cells, previously characterized as either highly invasive (metastatic) or noninvasive. Using both fluorescence microscopy and electrophysiological approaches, we show that SOCE is greatly diminished in invasive melanoma compared to its level in noninvasive cell types. However, no loss of expression of any members of the STIM and Orai families was observed in invasive melanoma cells. Moreover, overexpressed wild-type STIM1 and Orai1 failed to restore SOCE in invasive melanoma cells, and we observed no defects in their localization before or after store depletion in any of the invasive cell lines. Importantly, however, we determined that SOCE was restored by inhibition of protein kinase C, a known downstream target of Wnt5A. Furthermore, coexpression of STIM1 with an Orai1 mutant insensitive to protein kinase C-mediated phosphorylation fully restored SOCE in invasive melanoma. These findings reveal a level of control for STIM/Orai function in invasive melanoma not previously reported.

APA, Harvard, Vancouver, ISO, and other styles

49

Yu, Tao, Xi Li, Qianqian Luo, Huajing Liu, Jing Jin, Shengjie Li, and Jun He. "S417 in the CC3 region of STIM1 is critical for STIM1-Orai1 binding and CRAC channel activation." Life Science Alliance 6, no. 4 (January 23, 2023): e202201623. http://dx.doi.org/10.26508/lsa.202201623.

Full text

Abstract:

Store-operated Ca2+entry (SOCE) is a universal Ca2+influx pathway that is important for the function of many cell types. SOCE is controlled by the interaction of the ER Ca2+sensor STIM1 with the plasma membrane Ca2+channel Orai1. S417 is located in the third coiled-coil (CC3) domain of the C-terminus of STIM1. We found that single-point mutation of this residue (S417G) abolished STIM1 C-terminus interactions with Orai1. Mutation of S417 also abolished CAD-Orai1 binding and Orai1 channel activation, eliminated STIM1 puncta formation, and co-localization with Orai1 and SOCE. 2-APB was found to restore the binding of the STIM1 C-terminus mutant (S417G) to Orai1 and dose-dependently activate Orai1 channel. Both CBD and NBD of Orai1 are required for 2-APB–induced coupling between the Orai1 and STIM1 C-terminus mutant (S417G) and CRAC channel activation. We also demonstrated that 2-APB led to delayed activation of Orai1-K85E channel, although Orai1-K85E obviously impairs 2-APB–induced STIM1 C-terminus mutant (S417G)–Orai1 coupling. Our results suggest S417 in the CC3 domain of STIM1 is essential for STIM1–Orai1 binding and CRAC channel activation.

APA, Harvard, Vancouver, ISO, and other styles

50

Hodeify, Rawad, Manjula Nandakumar, Maryam Own, Raphael J. Courjaret, Johannes Graumann, Satanay Z. Hubrack, and Khaled Machaca. "The CCT chaperonin is a novel regulator of Ca2+ signaling through modulation of Orai1 trafficking." Science Advances 4, no. 9 (September 2018): eaau1935. http://dx.doi.org/10.1126/sciadv.aau1935.

Full text

Abstract:

Store-operated Ca2+ entry (SOCE) encodes a range of cellular responses downstream of Ca2+ influx through the SOCE channel Orai1. Orai1 recycles at the plasma membrane (PM), with ~40% of the total Orai1 pool residing at the PM at steady state. The mechanisms regulating Orai1 recycling remain poorly understood. We map the domains in Orai1 that are required for its trafficking to and recycling at the PM. We further identify, using biochemical and proteomic approaches, the CCT [chaperonin-containing TCP-1 (T-complex protein 1)] chaperonin complex as a novel regulator of Orai1 recycling by primarily regulating Orai1 endocytosis. We show that Orai1 interacts with CCT through its intracellular loop and that inhibition of CCT-Orai1 interaction increases Orai1 PM residence. This increased residence is functionally significant as it results in prolonged Ca2+ signaling, early formation of STIM1-Orai1 puncta, and more rapid activation of NFAT (nuclear factor of activated T cells) downstream of SOCE. Therefore, the CCT chaperonin is a novel regulator of Orai1 trafficking and, as such, a modulator of Ca2+ signaling and effector activation kinetics.

APA, Harvard, Vancouver, ISO, and other styles

Journal articles on the topic 'Orai1'

Create a spot-on reference in APA, MLA, Chicago, Harvard, and other styles