Hypoxia-induced Acidosis Uncouples the STIM-Orai Calcium Signaling Complex

Mancarella, Salvatore; Wang, Youjun; Deng, Xiaoxiang; Landesberg, Gavin; Scalia, Rosario; Panettieri, Reynold A.; Mallilankaraman, Karthik; Tang, Xiang D.; Madesh, Muniswamy; Gill, Donald L.

doi:10.1074/jbc.m111.303081

Cited by 55 publications

(66 citation statements)

References 52 publications

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“…For efficient and acute hypoxia induction, 1 mM sodium dithionite (Na 2 S 2 O 4 , Sigma, 157953) was used to chemically consume the dissolved oxygen in the culture medium. [74][75][76] It is reported that Na 2 S 2 O 4 can lower pO 2 to 10 Torr compared with 30 Torr pO 2 induced by 100% N 2 . 77 After adding Na 2 S 2 O 4 , the pH of the DMEM/F-12 medium was adjusted to 7.4 and maintained by HEPES buffer considering the acidification potential of Na 2 S 2 O 4 .…”

Section: Methodsmentioning

confidence: 99%

Real-time dynamics of methyl-CpG-binding domain protein 3 and its role in DNA demethylation by fluorescence correlation spectroscopy

et al. 2013

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Section: Methodsmentioning

confidence: 99%

Real-time dynamics of methyl-CpG-binding domain protein 3 and its role in DNA demethylation by fluorescence correlation spectroscopy

et al. 2013

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“…Cells were used 18 -24 h after transfection, and then incubated with 2 M fura-2 in buffer containing: 107 mM NaCl, 7.2 mM KCl, 1.2 mM MgCl 2 , 1 mM CaCl 2 , 11.5 mM glucose, 0.1% BSA, 20 mM HEPES, pH 7.2, for 30 min at room temperature, followed by treatment with fura-2-free solution for another 30 mins. Fluorescence ratio imaging was measured utilizing the Leica DMI6000B fluorescence microscope and Hamamatsu camera ORCA-Flash 4 controlled by SlideBook 6.0 software (Intelligent Imaging Innovations; Denver, CO) as described previously (45). Consecutive excitation at 340 nm (F 340 ) and 380 nm (F 380 ) was undertaken every 2 s, and emission fluorescence was collected at 505 nm.…”

Section: Methodsmentioning

confidence: 99%

The Orai1 Store-operated Calcium Channel Functions as a Hexamer

Cai

Zhou

Nwokonko

et al. 2016

Journal of Biological Chemistry

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102

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Edited by Roger ColbranOrai channels mediate store-operated Ca 2؉ signals crucial in regulating transcription in many cell types, and implicated in numerous immunological and inflammatory disorders. Despite their central importance, controversy surrounds the basic subunit structure of Orai channels, with several biochemical and biophysical studies suggesting a tetrameric structure yet crystallographic evidence indicating a hexamer. We systematically investigated the subunit configuration of the functional Orai1 channel, generating a series of tdTomato-tagged concatenated Orai1 channel constructs (dimers to hexamers) expressed in CRISPR-derived ORAI1 knock-out HEK cells, stably expressing STIM1-YFP. Surface biotinylation demonstrated that the fulllength concatemers were surface membrane-expressed. Unexpectedly, Orai1 dimers, trimers, tetramers, pentamers, and hexamers all mediated similar and substantial store-operated Ca 2؉ entry. Moreover, each Orai1 concatemer mediated Ca 2؉ currents with inward rectification and reversal potentials almost identical to those observed with expressed Orai1 monomer. In Orai1 tetramers, subunit-specific replacement with Orai1 E106A "pore-inactive" subunits revealed that functional channels utilize only the N-terminal dimer from the tetramer. In contrast, Orai1 E106A replacement in Orai1 hexamers established that all the subunits can contribute to channel formation, indicating a hexameric channel configuration. The critical Ca 2؉ selectivity filter-forming Glu-106 residue may mediate Orai1 channel assembly around a central Ca 2؉ ion within the pore. Thus, multiple E106A substitutions in the Orai1 hexamer may promote an alternative "trimer-of-dimers" channel configuration in which the C-terminal E106A subunits are excluded from the hexameric core. Our results argue strongly against a tetrameric configuration for Orai1 channels and indicate that the Orai1 channel functions as a hexamer.

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“…In support of this concept, following ischemia in the isolated perfused heart, SR Ca 2ϩ levels were observed to decrease rapidly at reperfusion (124), which is of course a key step in the activation of SOCE. Furthermore, Mancarella et al (61) reported that hypoxia stimulated STIM1 accumulation at ER-plasma membrane junctions, whereas the accompanying acidosis uncoupled the STIM1-Orai1 complex, thereby preventing Ca 2ϩ overload. Although not in cardiomyocytes, this study suggests that ischemia, which is also characterized by hypoxia and acidosis, could be a trigger for the molecular machinery required for SOCE to be positioned for rapid activation upon reperfusion when the acidosis is quickly resolved.…”

Section: Potential Role For Stim1-mediated Soce In Ischemia/reperfusimentioning

confidence: 99%

“…Also as noted earlier, increased Review H452 STORE-OPERATED Ca 2ϩ ENTRY IN THE HEART Orai-mediated SOCE has been implicated in initiation of atrial and ventricular arrhythmias (132,137) and we reported that in the isolated perfused heart glucosamine, which inhibits SOCE, decreased the incidence of arrhythmias following I/R (25). Therefore, with hypoxia activating STIM1 accumulation in plasma membrane junctions (61), combined with a rapid decrease in SR Ca 2ϩ levels at reperfusion (124), the stage is set for uncontrolled SOCE contributing to Ca 2ϩ overload, a key mediator of early I/R injury, and a potential trigger for arrhythmogenesis. This is supported by observations in a range of different model systems demonstrating that pharmacological inhibition of SOCE blunts Ca 2ϩ overload and attenuates I/Rinduced arrhythmias.…”

Section: Potential Role For Stim1-mediated Soce In Ischemia/reperfusimentioning

confidence: 99%

STIM1/Orai1-mediated SOCE: current perspectives and potential roles in cardiac function and pathology

Collins

Zhu-Mauldin

Marchase

et al. 2013

American Journal of Physiology-Heart and Circulatory Physiology

110

102

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Collins HE, Zhu-Mauldin X, Marchase RB, Chatham JC. STIM1/Orai1-mediated SOCE: current perspectives and potential roles in cardiac function and pathology. Am J Physiol Heart Circ Physiol 305: H446 -H458, 2013. First published June 21, 2013 doi:10.1152/ajpheart.00104.2013.-Store-operated Ca 2ϩ entry (SOCE) is critical for Ca 2ϩ signaling in nonexcitable cells; however, its role in the regulation of cardiomyocyte Ca 2ϩ homeostasis has only recently been investigated. The increased understanding of the role of stromal interaction molecule 1 (STIM1) in regulating SOCE combined with recent studies demonstrating the presence of STIM1 in cardiomyocytes provides support that this pathway co-exists in the heart with the more widely recognized Ca 2ϩ handling pathways associated with excitation-contraction coupling. There is now substantial evidence that STIM1-mediated SOCE plays a key role in mediating cardiomyocyte hypertrophy, both in vitro and in vivo, and there is growing support for the contribution of SOCE to Ca 2ϩ overload associated with ischemia/reperfusion injury. Here, we provide an overview of our current understanding of the molecular regulation of SOCE and discuss the evidence supporting the role of STIM1/Orai1-mediated SOCE in regulating cardiomyocyte function.store-operated Ca 2ϩ entry; stromal interaction molecule 1; orai1; cardiomyocytes STORE-OPERATED CA 2ϩ ENTRY (SOCE), also known as capacitative calcium entry (CCE), is the major mechanism of Ca 2ϩ entry in nearly all nonexcitable cells (97, 99). In addition, it is increasingly recognized to co-exist with voltage-gated Ca 2ϩ channels in excitable cells, including neurons, skeletal muscle cells, and cardiomyocytes (1,38,45,83,121). In response to inositol 1,4,5-triphosphate (IP 3 )-(43) or ryanodine receptor (RyR)-mediated (3) Ca 2ϩ release from ER/SR stores, SOCE facilitates the influx of Ca 2ϩ from the extracellular space, resulting in a sustained increase in cytosolic Ca 2ϩ levels. Thus, although one of the roles of SOCE is to rapidly refill the depleted ER/SR stores, the subsequent sustained increase in cytosolic Ca 2ϩ also regulates numerous gene transcription pathways (33,50,151). Indeed, aberrant SOCE has been observed in a growing number of diseases including severe combined immunodeficiency, acute pancreatitis, and Alzheimer's disease (86).The integration of SOCE into well-established models of Ca 2ϩ homeostasis was hampered for many years by the lack of specific molecular mediators; even as recently as 2004 there was considerable controversy as to the specific mechanisms regulating SOCE (90, 113). However, in 2005, stromal interaction molecule 1 (STIM1) was found to localize to the ER/SR membrane and shown to function as a primary mediator of SOCE (54, 102). The putative physiological and pathophysiological roles of SOCE and STIM1 that have been identified to date are summarized in Table 1. A number of studies have recently reported the presence of STIM1 in adult cardiomyocytes (37,59,153), and consistent with earlier evidence supporting a rol...

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Hypoxia-induced Acidosis Uncouples the STIM-Orai Calcium Signaling Complex

Cited by 55 publications

References 52 publications

Real-time dynamics of methyl-CpG-binding domain protein 3 and its role in DNA demethylation by fluorescence correlation spectroscopy

Real-time dynamics of methyl-CpG-binding domain protein 3 and its role in DNA demethylation by fluorescence correlation spectroscopy

The Orai1 Store-operated Calcium Channel Functions as a Hexamer

STIM1/Orai1-mediated SOCE: current perspectives and potential roles in cardiac function and pathology

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