Статті в журналах: "ORAOV1"

1

Jiang, L., H. S. Yang, Z. Wang, Y. Zhou, M. Zhou, X. Zeng, and Q. M. Chen. "ORAOV1-A Correlates with Poor Differentiation in Oral Cancer." Journal of Dental Research 88, no. 5 (May 2009): 433–38. http://dx.doi.org/10.1177/0022034509336994.

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Oral cancer overexpressed 1 ( ORAOV1) is a crucial oncogene in oral squamous cell carcinoma (OSCC). In this study, we have identified a novel splice variant of ORAOV1, designated as ORAOV1-A. To study the potential role of ORAOV1-A in OSCC, we tested its expression in 7 OSCC cell lines, as well as in 19 normal oral tissue samples and 47 OSCC tissue samples. The expression of ORAOV1-A was detectable in 6 out of 7 OSCC cell lines tested. In OSCC tissue samples, the expression frequency of ORAOV1-A (51.1%) was much higher than that in normal samples (10.5%). Notably, an inverse correlation was found between the expression frequency of ORAOV1-A and the degree of differentiation in OSCC ( P = 0.0017). In conclusion, our results suggested that ORAOV1-A may play a functional role in the tumorigenesis of OSCC, and ORAOV1-A expression may serve as an adjunctive prognostic indicator for persons with OSCC.

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2

Xavier, Flávia Caló Aquino, Camila Oliveira Rodini, Katiúcia Batista Silva Paiva, Maria Fernanda Souza Setúbal Destro, Patricia Severino, Raquel A. Moyses, Eloiza H. Tajara, and Fabio Daumas Nunes. "ORAOV1 is amplified in oral squamous cell carcinoma." Journal of Oral Pathology & Medicine 41, no. 1 (May 28, 2011): 54–60. http://dx.doi.org/10.1111/j.1600-0714.2011.01053.x.

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3

Салеева, Д. В., В. Ф. Михайлов, Л. В. Шуленина, В. В. Виноградов, А. А. Бахтин, К. В. Акопян, М. В. Незнанова, and Г. Д. Засухина. "Activities of regulatory RNAs that affect development of tumor cells in patients with laryngeal cancer." ZHurnal «Patologicheskaia fiziologiia i eksperimental`naia terapiia», no. 4() (November 21, 2018): 67–74. http://dx.doi.org/10.25557/0031-2991.2018.04.67-74.

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Цель. Определение прогностической значимости и роли экспрессии некодирующих РНК (длинные РНК и микроРНК), и белка кодирующих генов в патогенезе рака гортани. Методика. Исследован биопсийный материал и периферическая кровь 35 пациентов с диагнозом плоскоклеточный рак гортани (ПРГ) с классификацией от T1N0M0 до T4N1M0. Контролем служили образцы близлежащей гистологически неизмененной ткани гортани тех же больных. Для оценки экспрессии генов исследовали кровь 27 здоровых доноров. Содержание мРНК генов ( р53, CCND1, ORAOV1, hPTEN ), длинных некодирующих РНК (днРНК): NEAT1, MALAT1, ROR , а также зрелых микроРНК (miR-21, miR-27a, miR-34a, miR-101, miR-124, miR-125b, miR-181а) в опухолевой ткани и крови определяли методом ПЦР в реальном времени (ПЦР-РВ). Результаты. Выявлено увеличение содержания мРНК генов CCND1, hPTEN , днРНК NEAT1, MALAT1 и miR-21, miR-27a в крови у пациентов с ПРГ. Установлено, что уровень мРНК генов CCND1, ORAOV1 был значимо выше при исследовании биоптатов у больных 3-й - 4-й стадии, чем у больных 1-й - 2-й стадии заболевания. Такая же закономерность выявлена для днРНК NEAT1, MALAT1 и для miR-101. Экспрессия miR-27a и miR-124 на более поздних стадиях болезни была ниже, чем у пациентов 1-2 стадии. Заключение. Выявлена возможность использования исследованных днРНК, микроРНК и мРНК белоккодирующих генов для индивидуального прогноза заболевания при создании панели биомаркеров. Aim. To study the role of non-coding RNA (long RNAs and microRNAs) expression and protein-coding genes in the pathogenesis of laryngeal cancer to determine their prognostic significance for oncotransformation. Methods. The expression of long non-coding RNAs, microRNAs and protein-coding genes was examined in biopsy samples (fresh frozen tissue) and peripheral blood samples from 35 patients with laryngeal squamous cell cancer (LSCC) at T1N0M0 - T4N1M0 stages. Samples of surrounding, histologically unchanged tissues collected from the same patients were used as control. Gene expression was evaluated in blood samples from 27 healthy donors. Contents of gene mRNAs ( p53, CCND1, ORAOV1, hPTEN ), long non-coding RNAs (IncRNA) ( NEAT1, MALAT1, ROR ), and mature miRNAs (miR-21, miR-27a, miR-34a, miR-101, miR-124, miR -125b, miR-181a) were measured in tissue and blood using real-time PCR. Results. Contents of CCND1 and hPTEN gene mRNAs, lncRNAs ( NEAT1, MALAT1), miR-21, and miR-27a were increased in blood of patients with LSCC. Levels of CCND1 and ORAOV1 gene mRNAs were significantly higher in biopsy samples from stage 3-4 patients compared to stage 1-2 patients. A similar expression pattern was observed for lncRNAs NEAT1 and MALAT1 and miR-101. On the other hand, expression of miR-27a and miR-124 was lower at later stages than at stages 1-2. Conclusion. The studied lncRNAs, microRNAs and protein-coding genes can be used in development of a biomarker panel for individual prognosis of the disease.

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4

Jiang, Lu, Xin Zeng, Zhi Wang, Ning Ji, Yu Zhou, Xianting Liu, and Qianming Chen. "Oral cancer overexpressed 1 (ORAOV1) regulates cell cycle and apoptosis in cervical cancer HeLa cells." Molecular Cancer 9, no. 1 (2010): 20. http://dx.doi.org/10.1186/1476-4598-9-20.

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5

Togashi, Y., T. Arao, H. Kato, K. Matsumoto, M. Terashima, H. Hayashi, Y. Fujita, T. Yasuda, H. Shiozaki, and K. Nishio. "ORAOV1 is Amplified in Esophageal Squamous Cell Cancer and Related to Tumor Growth and Poorly Differentiated Tumor." Annals of Oncology 24 (November 2013): ix60. http://dx.doi.org/10.1093/annonc/mdt459.131.

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6

Prusty, Nihar Ranjan, Francesca Camponeschi, Simone Ciofi-Baffoni, and Lucia Banci. "The human YAE1-ORAOV1 complex of the cytosolic iron-sulfur protein assembly machinery binds a [4Fe-4S] cluster." Inorganica Chimica Acta 518 (April 2021): 120252. http://dx.doi.org/10.1016/j.ica.2021.120252.

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7

Salmon Hillbertz, Nicolette H. C., Magnus Isaksson, Elinor K. Karlsson, Eva Hellmén, Gerli Rosengren Pielberg, Peter Savolainen, Claire M. Wade, et al. "Duplication of FGF3, FGF4, FGF19 and ORAOV1 causes hair ridge and predisposition to dermoid sinus in Ridgeback dogs." Nature Genetics 39, no. 11 (September 30, 2007): 1318–20. http://dx.doi.org/10.1038/ng.2007.4.

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8

Zhao, Xin, Dongjuan Liu, Lili Wang, Ruiqing Wu, Xin Zeng, Hongxia Dan, Ning Ji, Lu Jiang, Yu Zhou, and Qianming Chen. "RNAi-mediated downregulation of oral cancer overexpressed 1 (ORAOV1) inhibits vascular endothelial cell proliferation, migration, invasion, and tube formation." Journal of Oral Pathology & Medicine 45, no. 4 (October 9, 2015): 256–61. http://dx.doi.org/10.1111/jop.12371.

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9

Jiang, Lu, Xin Zeng, Hanshuo Yang, Zhi Wang, Jun Shen, Jingping Bai, Yuanyuan Zhang, Feng Gao, Min Zhou, and Qianming Chen. "Oral cancer overexpressed 1 (ORAOV1): A regulator for the cell growth and tumor angiogenesis in oral squamous cell carcinoma." International Journal of Cancer 123, no. 8 (October 15, 2008): 1779–86. http://dx.doi.org/10.1002/ijc.23734.

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10

Togashi, Yosuke, Tokuzo Arao, Hiroaki Kato, Kazuko Matsumoto, Masato Terashima, Hidetoshi Hayashi, Marco A. de Velasco, et al. "Frequent amplification of ORAOV1 gene in esophageal squamous cell cancer promotes an aggressive phenotype via proline metabolism and ROS production." Oncotarget 5, no. 10 (December 30, 2013): 2962–73. http://dx.doi.org/10.18632/oncotarget.1561.

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11

Zhao, X., D. Liu, L. Wang, R. Wu, X. Zeng, H. Dan, N. Ji, L. Jiang, Y. Zhou, and Q. Chen. "RNAI-mediated downregulation of oral cancer overexpressed 1 (ORAOV1) regulates proliferation, migration, invasion and tube-formation in vascular endothelial cells." International Journal of Oral and Maxillofacial Surgery 44 (October 2015): e315. http://dx.doi.org/10.1016/j.ijom.2015.08.410.

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12

Li, Man, Xiaobin Cui, Yaoyuan Shen, Hongchao Dong, Weihua Liang, Yunzhao Chen, Jianming Hu, et al. "ORAOV1 overexpression in esophageal squamous cell carcinoma and esophageal dysplasia: a possible biomarker of progression and poor prognosis in esophageal carcinoma." Human Pathology 46, no. 5 (May 2015): 707–15. http://dx.doi.org/10.1016/j.humpath.2015.01.009.

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13

Zhai, C., Y. li, C. Mascarenhas, Q. Lin, K. Li, I. Vyrides, C. M. Grant, and B. Panaretou. "The function of ORAOV1/LTO1, a gene that is overexpressed frequently in cancer: essential roles in the function and biogenesis of the ribosome." Oncogene 33, no. 4 (January 14, 2013): 484–94. http://dx.doi.org/10.1038/onc.2012.604.

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14

Das, Subal, Bigitendriya Debsharma, and Kaushik Bose. "Adiposity and Health Status among Adult Male Mundas and Oraons of Paschim Medinipur, West Bengal, India." Journal of Anthropology 2013 (May 19, 2013): 1–6. http://dx.doi.org/10.1155/2013/324264.

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The present cross-sectional study was conducted among two male tribal groups Munda (n=106) and Oraon (n=104) aged 18–73 years of Paschim Medinipur, West Bengal. Objective was to evaluate the health status based on body mass index (BMI) and percent body fat (PBF). Measurements of weight, height, circumferences, and skinfolds were recorded. Results revealed that mean age of Mundas (36.2±13.3) and Oraons (35.1±15.3) in years were similar. Significant (P<0.05) ethnic differences in mean chest circumference and anterior thigh skinfold were observed. Both Munda (50.0%) and Oraon (46.2%) males suffered from very high degree of chronic energy deficiency (CED) based on BMI. Similarly, for percent body fat (PBF), Mundas (29.3%) and Oraons (35.4%) had unhealthy (too low) PBF (i.e., ≤5%) levels. Significantly negative correlations were observed between age and BMI and positive correlations between age, waist-hip ratio (WHR), and conicity index (CI) (only Mundas) among Mundas and Oraons. In Linear regression, age had a significant impact on all derived central and overall adiposity measures. Prospective studies are required to determine the associations between health status and PBF as well as nutrition status and BMI in different indigenous ethnic groups of India and elsewhere.

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15

DASGUPTA, SANGEETA. "‘Heathen aboriginals’, ‘Christian tribes’, and ‘animistic races’: Missionary narratives on the Oraons of Chhotanagpur in colonial India." Modern Asian Studies 50, no. 2 (July 24, 2015): 437–78. http://dx.doi.org/10.1017/s0026749x15000025.

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AbstractThrough a description of the interactions of Christian missionaries in Chhotanagpur with the Oraons, this article illustrates the different ways in which the missionaries grappled with and restructured their notions of the ‘tribe’ and the ‘Oraon’ across the nineteenth and early twentieth centuries. The Oraon, I argue, was initially recognized in terms of his heathen practices, his so-called compact with the Devil, and his world of idolatry and demonology. But, by the end of the nineteenth century, he increasingly became, in missionary language, an animistic aboriginal tribe, a ‘primitive’ within an evolutionary schema. As the missionaries searched for an authentic Oraon language, for myths, traditions and histories, an array of categories—heathen, savage, race, tribe, and aboriginal—seemingly jostled with one another in their narratives. Indeed, the tension between religion and race could never be resolved in missionary narratives; this was reflected in colonial ethnographic literature that drew upon and yet eventually marginalized missionary representations. I conclude by referring to a case in the 1960s filed by Kartik Oraon against the Protestant convert David Munzni before the Election Tribunal at Ranchi, which was ultimately resolved in the Supreme Court, that raised the question whether religion or race determined tribal identity.

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16

Dey, Archita, Mahua Chanak, Kaustav Das, Koel Mukherjee, and Kaushik Bose. "Variation in lip print pattern between two ethnic groups, Oraon tribals and Bengalee Hindus, residing in West Bengal, India." Anthropological Review 82, no. 4 (December 1, 2019): 405–15. http://dx.doi.org/10.2478/anre-2019-0031.

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Abstract Lip print pattern (LPP) is unique to each individual. For decades, forensic experts have used LPP for personal identification to solve criminal cases. However, studies investigating ethnic variation in LPP are scanty. Our study wanted to investigate variation in LPP between two ethnic groups, Oraon tribals and Bengalee Hindus, residing in West Bengal, India. A total of 280 participants included 112 Oraons and168 Bengalee Hindus of both. Prints were taken using dark shaded lipstick and transparent cellophane tape and recorded into white A4 sheet. Prints were divided into four quadrants and examined by magnifying glass. For analysis of results, classification of Suzuki and Tsuchihashi was followed. A p value of 0.05 was considered to be statistically significant. It was observed that Type II pattern was dominant in first and second quadrants in both ethnic groups, irrespective of sex. Combination of Type II+III was found to be the most common pattern in males among both Oraons (16.2%) and Bengalee Hindus (12.2%) whereas in females Type II pattern (25.0%) among Oraons and Type III pattern among Bengalee Hindus (11.4%) was the most common. Chi square test showed statistically significant difference among females (p<0.05) and in third and fourth quadrants among males (p<0.01) of both ethnic groups. Our investigation clearly demonstrated sex and ethnic variations in LPP. Further studies are required to investigate ethnic variation in LPP among the various populations groups, both tribal as well as non-tribal, from different regions of India.

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17

Chowdhury, Tanaya Kundu, and Subrata K. Roy. "Prevalence of anaemia and associated factors among Oraon females of North 24 Parganas, West Bengal, India." Anthropological Review 82, no. 1 (March 1, 2019): 15–27. http://dx.doi.org/10.2478/anre-2019-0002.

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Abstract Low haemoglobin level or anaemia is a health problem worldwide especially in developing countries like India. Anaemia is generally higher among indigenous groups compared to general population globally and females are specifically more prone to anaemia. However, studies are inadequate on indigenous groups of India. The aims of the study are to determine the prevalence of anaemia among the female Oraons of North 24 Parganas and to study the association between anaemia and concomitants like socio-demographic and food habit variables. Data have been collected on demographic, socio-economic and food habit variables using well-tested questionnaire from 309 Oraon females living in rural and urban areas of North 24 Parganas, West Bengal. Haemoglobin data were collected using standard instrument and technique. Descriptive statistics and binary logistic regression were used to analyze the data using SPSS version 16.0. Females of rural and urban areas were mostly married and non-literate, employed as labourers and had sedentary occupations. Majority of them consume fruits and vegetables but less animal protein and dairy product. Around 80% of the females were anaemic, irrespective of their habitat, socio-economic status and food habits. Anaemic status was associated with insufficient intake of animal protein, which is significantly associated with the anaemia status of the females in the present study.

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18

Järv, Risto. "Old stories in contemporary times - a collecting experience in the Orava village in Siberia." Folklore: Electronic Journal of Folklore 13 (2000): 37–65. http://dx.doi.org/10.7592/fejf2000.13.orava.

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19

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.

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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.

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20

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.

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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.

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21

Voros, Orsolya, György Panyi, and Péter Hajdu. "Immune Synapse Residency of Orai1 Alters Ca2+ Response of T Cells." International Journal of Molecular Sciences 22, no. 21 (October 26, 2021): 11514. http://dx.doi.org/10.3390/ijms222111514.

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CRAC, which plays important role in Ca2+-dependent T-lymphocyte activation, is composed of the ER-resident STIM1 and the plasma membrane Orai1 pore-forming subunit. Both accumulate at the immunological synapse (IS) between a T cell and an antigen-presenting cell (APC). We hypothesized that adapter/interacting proteins regulate Orai1 residence in the IS. We could show that mGFP-tagged Orai1-Full channels expressed in Jurkat cells had a biphasic IS-accumulation kinetics peaked at 15 min. To understand the background of Orai1 IS-redistribution we knocked down STIM1 and SAP97 (adaptor protein with a short IS-residency (15 min) and ability to bind Orai1 N-terminus): the mGFP-Orai1-Full channels kept on accumulating in the IS up to the 60th minute in the STIM1- and SAP97-lacking Jurkat cells. Deletion of Orai1 N terminus (mGFP-Orai1-Δ72) resulted in the same time course as described for STIM1/SAP97 knock-down cells. Ca2+-imaging of IS-engaged T-cells revealed that of Orai1 residency modifies the Ca2+-response: cells expressing mGFP-Orai1-Δ72 construct or mGFP-Orai1-Full in SAP-97 knock-down cells showed higher number of Ca2+-oscillation up to the 90th minute after IS formation. Overall, these data suggest that SAP97 may contribute to the short-lived IS-residency of Orai1 and binding of STIM1 to Orai1 N-terminus is necessary for SAP97-Orai1 interaction.

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22

Li, Peiyao, Yong Miao, Adish Dani та Monika Vig. "α-SNAP regulates dynamic, on-site assembly and calcium selectivity of Orai1 channels". Molecular Biology of the Cell 27, № 16 (15 серпня 2016): 2542–53. http://dx.doi.org/10.1091/mbc.e16-03-0163.

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Orai1 forms a highly calcium-selective pore of the calcium release activated channel, and α-SNAP is necessary for its function. Here we show that α-SNAP regulates on-site assembly of Orai1 dimers into calcium-selective multimers. We find that Orai1 is a dimer in resting primary mouse embryonic fibroblasts but displays variable stoichiometry in the plasma membrane of store-depleted cells. Remarkably, α-SNAP depletion induces formation of higher-order Orai1 oligomers, which permeate significant levels of sodium via Orai1 channels. Sodium permeation in α-SNAP–deficient cells cannot be corrected by tethering multiple Stim1 domains to Orai1 C-terminal tail, demonstrating that α-SNAP regulates functional assembly and calcium selectivity of Orai1 multimers independently of Stim1 levels. Fluorescence nanoscopy reveals sustained coassociation of α-SNAP with Stim1 and Orai1, and α-SNAP–depleted cells show faster and less constrained mobility of Orai1 within ER-PM junctions, suggesting Orai1 and Stim1 coentrapment without stable contacts. Furthermore, α-SNAP depletion significantly reduces fluorescence resonance energy transfer between Stim1 and Orai1 N-terminus but not C-terminus. Taken together, these data reveal a unique role of α-SNAP in the on-site functional assembly of Orai1 subunits and suggest that this process may, in part, involve enabling crucial low-affinity interactions between Orai1 N-terminus and Stim1.

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23

Gwack, Yousang, Sonal Srikanth, Masatsugu Oh-hora, Patrick G. Hogan, Edward D. Lamperti, Megumi Yamashita, Curtis Gelinas, et al. "Hair Loss and Defective T- and B-Cell Function in Mice Lacking ORAI1." Molecular and Cellular Biology 28, no. 17 (June 30, 2008): 5209–22. http://dx.doi.org/10.1128/mcb.00360-08.

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ABSTRACT ORAI1 is a pore subunit of the store-operated Ca2+ release-activated Ca2+ (CRAC) channel. To examine the physiological consequences of ORAI1 deficiency, we generated mice with targeted disruption of the Orai1 gene. The results of immunohistochemical analysis showed that ORAI1 is expressed in lymphocytes, skin, and muscle of wild-type mice and is not expressed in Orai1 −/− mice. Orai1 −/− mice with the inbred C57BL/6 background showed perinatal lethality, which was overcome by crossing them to outbred ICR mice. Orai1 −/− mice were small in size, with eyelid irritation and sporadic hair loss resembling the cyclical alopecia observed in mice with keratinocyte-specific deletion of the Cnb1 gene. T and B cells developed normally in Orai1 −/− mice, but B cells showed a substantial decrease in Ca2+ influx and cell proliferation in response to B-cell receptor stimulation. Naïve and differentiated Orai1 −/− T cells showed substantial reductions in store-operated Ca2+ entry, CRAC currents, and cytokine production. These features are consistent with the severe combined immunodeficiency and mild extraimmunological symptoms observed in a patient with a missense mutation in human ORAI1 and distinguish the ORAI1-null mice described here from a previously reported Orai1 gene-trap mutant mouse which may be a hypomorph rather than a true null.

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24

Yu, Fang, Lu Sun, and Khaled Machaca. "Constitutive recycling of the store-operated Ca2+ channel Orai1 and its internalization during meiosis." Journal of Cell Biology 191, no. 3 (November 1, 2010): 523–35. http://dx.doi.org/10.1083/jcb.201006022.

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The egg’s competency to activate at fertilization and transition to embryogenesis is dependent on its ability to generate a fertilization-specific Ca2+ transient. To endow the egg with this capacity, Ca2+ signals remodel during oocyte maturation, including inactivation of the primary Ca2+ influx pathway store-operated Ca2+ entry (SOCE). SOCE inactivation is coupled to internalization of the SOCE channel, Orai1. In this study, we show that Orai1 internalizes during meiosis through a caveolin (Cav)- and dynamin-dependent endocytic pathway. Cav binds to Orai1, and we map a Cav consensus–binding site in the Orai1 N terminus, which is required for Orai1 internalization. Furthermore, at rest, Orai1 actively recycles between an endosomal compartment and the cell membrane through a Rho-dependent endocytic pathway. A significant percentage of total Orai1 is intracellular at steady state. Store depletion completely shifts endosomal Orai1 to the cell membrane. These results define vesicular trafficking mechanisms in the oocyte that control Orai1 subcellular localization at steady state, during meiosis, and after store depletion.

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25

Son, Ga-Yeon, Krishna Prasad Subedi, Hwei Ling Ong, Lucile Noyer, Hassan Saadi, Changyu Zheng, Rajesh Bhardwaj, Stefan Feske, and Indu Suresh Ambudkar. "STIM2 targets Orai1/STIM1 to the AKAP79 signaling complex and confers coupling of Ca2+entry with NFAT1 activation." Proceedings of the National Academy of Sciences 117, no. 28 (June 29, 2020): 16638–48. http://dx.doi.org/10.1073/pnas.1915386117.

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The Orai1 channel is regulated by stromal interaction molecules STIM1 and STIM2 within endoplasmic reticulum (ER)-plasma membrane (PM) contact sites. Ca2+signals generated by Orai1 activate Ca2+-dependent gene expression. When compared with STIM1, STIM2 is a weak activator of Orai1, but it has been suggested to have a unique role in nuclear factor of activated T cells 1 (NFAT1) activation triggered by Orai1-mediated Ca2+entry. In this study, we examined the contribution of STIM2 in NFAT1 activation. We report that STIM2 recruitment of Orai1/STIM1 to ER-PM junctions in response to depletion of ER-Ca2+promotes assembly of the channel with AKAP79 to form a signaling complex that couples Orai1 channel function to the activation of NFAT1. Knockdown of STIM2 expression had relatively little effect on Orai1/STIM1 clustering or local and global [Ca2+]iincreases but significantly attenuated NFAT1 activation and assembly of Orai1 with AKAP79. STIM1ΔK, which lacks the PIP2-binding polybasic domain, was recruited to ER-PM junctions following ER-Ca2+depletion by binding to Orai1 and caused local and global [Ca2+]iincreases comparable to those induced by STIM1 activation of Orai1. However, in contrast to STIM1, STIM1ΔK induced less NFAT1 activation and attenuated the association of Orai1 with STIM2 and AKAP79. Orai1-AKAP79 interaction and NFAT1 activation were recovered by coexpressing STIM2 with STIM1ΔK. Replacing the PIP2-binding domain of STIM1 with that of STIM2 eliminated the requirement of STIM2 for NFAT1 activation. Together, these data demonstrate an important role for STIM2 in coupling Orai1-mediated Ca2+influx to NFAT1 activation.

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26

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.

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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.

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Vörös, Orsolya, György Panyi, and Péter Hajdu. "Molecular background of Orai1 accumulation in the immunological synapse." Journal of Immunology 202, no. 1_Supplement (May 1, 2019): 184.3. http://dx.doi.org/10.4049/jimmunol.202.supp.184.3.

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Abstract CRAC channels have a vital role in the activation of T lymphocytes, through their calcium influx. CRAC is composed of the ER-membrane resident STIM1 (stromal interaction molecule 1) and the plasma membrane Orai1 pore-forming subunit. Both are accumulated at the interface of the immunological synapse (IS) between a T cell and an antigen-presenting cell (APC). CRAC channelopathies due to mutations in either Orai1 or STIM1 are characterized by severe combined immunodeficiency (SCID)-like disease, autoimmunity, muscular hypotonia, and ectodermal dysplasia. The down-regulation of actin-binding protein expression (e.g. HS1), which may be responsible for the immobilization of ion channels, hampers Ca2+-signaling and IS formation of T lymphocytes. We hypothesized that actin-binding proteins modulate the trafficking of the CRAC channels to the IS, which is crucial for the Ca2+-signaling. By means of GST-affinity assay we could show that HS1, which is an actin-regulatory adaptor protein, binds to the N-terminus of the Orai1. N-terminal truncated Orai1 channels (ΔN1: 1–74, ΔN2: 1–88) were created and expressed in Jurkat cells. Then Orai1 channel expressing Jurkat cells were conjugated to CD3/CD28 beads as an APC and Orai1 polarization was determined: the Orai1-ΔN1 mutant similar to the wild-type Orai1 redistributed to the IS but the Orai1-ΔN2 did not. Furthermore, the HS1 protein did not show IS-accumulation in cells expressing the Orai1-ΔN2 subunits unlike for those transfected with the Orai1-ΔN1 or the wild-type constructs. Overall, these data indicate that Orai1-HS1 interaction could contribute to the anchoring of the Orai1 in the IS and can affect the Ca2+-signaling upon T cell activation.

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Yeh, Yi-Chun, Yu-Ping Lin, Holger Kramer, and Anant B. Parekh. "Single-nucleotide polymorphisms in Orai1 associated with atopic dermatitis inhibit protein turnover, decrease calcium entry and disrupt calcium-dependent gene expression." Human Molecular Genetics 29, no. 11 (October 10, 2019): 1808–23. http://dx.doi.org/10.1093/hmg/ddz223.

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Abstract Loss-of function mutations in Orai1 Ca2+ channels lead to a form of severe combined immunodeficiency, auto-immunity, muscle hypotonia and defects in dental enamel production and sweat gland function. Two single-nucleotide polymorphisms (SNPs) in Orai1 have been found and localize to the second extracellular loop. These polymorphisms associate with atopic dermatitis but how they affect Ca2+ signalling and cell function is unknown. Here, we find that Orai1–SNPs turnover considerably more slowly than wild type Orai1 and are more abundantly expressed in the plasma membrane. We show a central role for flotillin in the endocytotic recycling of Orai1 channels and that endocytosed wild type Orai1 is trafficked to Rab 7-positive late endosomes for lysosomal degradation. Orai1–SNPs escape the degradation pathway and instead enter Rab 11-positive recycling endosomes, where they are returned to the surface membrane through Arf6-dependent exocytosis. We find that Orai1–SNPs escape late endosomes through endosomal pH regulation of interaction between the channel and flotillin. We identify a pH-sensitive electrostatic interaction between positively charged arginine in extracellular loop 2 (K210) and a negatively charged aspartate (D112) in extracellular loop 1 that helps determine Orai1 turnover. The increase in membrane Orai1–SNP leads to a mis-match in Orai1–STIM stoichiometry, resulting in inhibition of Ca2+ entry and Ca2+-dependent gene expression. Our results identify new strategies for targeting atopic dermatitis.

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Dynes, Joseph L., Anna Amcheslavsky, and Michael D. Cahalan. "Genetically targeted single-channel optical recording reveals multiple Orai1 gating states and oscillations in calcium influx." Proceedings of the National Academy of Sciences 113, no. 2 (December 28, 2015): 440–45. http://dx.doi.org/10.1073/pnas.1523410113.

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Orai1 comprises the pore-forming subunit of the Ca2+ release-activated Ca2+ (CRAC) channel. When bound and activated by stromal interacting molecule 1 (STIM1), an endoplasmic reticulum (ER)-resident calcium sensor, Orai1 channels possess high selectivity for calcium but extremely small conductance that has precluded direct recording of single-channel currents. We have developed an approach to visualize Orai1 activity by fusing Orai1 to fluorescent, genetically encoded calcium indicators (GECIs). The GECI–Orai1 probes reveal local Ca2+ influx at STIM1–Orai1 puncta. By whole cell recording, these fusions are fully functional as CRAC channels. When GECI–Orai1 and the CRAC-activating domain (CAD) of STIM1 were coexpressed at low levels and imaged using a total internal reflectance fluorescence microscope, cells exhibited sporadic fluorescence transients the size of diffraction-limited spots and the brightness of a few activated GECI proteins. Transients typically rose rapidly and fell into two classes according to duration: briefer “flickers” lasting only a few hundred milliseconds, and longer “pulses” lasting one to several seconds. The size, intensity, trace shape, frequency, distribution, physiological characteristics, and association with CAD binding together demonstrate that GECI–Orai1 fluorescence transients correspond to single-channel Orai1 responses. Single Orai1 channels gated by CAD, and small Orai1 puncta gated by STIM1, exhibit repetitive fluctuations in single-channel output. CAD binding supports a role in open state maintenance and reveals a second phase of CAD/STIM1 binding after channel opening. These first recordings of single-channel Orai1 currents reveal unexpected dynamics, and when paired with CAD association, support multiple single-channel states.

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Tiffner, Adéla, Valentina Hopl, Romana Schober, Matthias Sallinger, Herwig Grabmayr, Carmen Höglinger, Marc Fahrner, et al. "Orai1 Boosts SK3 Channel Activation." Cancers 13, no. 24 (December 17, 2021): 6357. http://dx.doi.org/10.3390/cancers13246357.

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The interplay of SK3, a Ca2+ sensitive K+ ion channel, with Orai1, a Ca2+ ion channel, has been reported to increase cytosolic Ca2+ levels, thereby triggering proliferation of breast and colon cancer cells, although a molecular mechanism has remained elusive to date. We show in the current study, via heterologous protein expression, that Orai1 can enhance SK3 K+ currents, in addition to constitutively bound calmodulin (CaM). At low cytosolic Ca2+ levels that decrease SK3 K+ permeation, co-expressed Orai1 potentiates SK3 currents. This positive feedback mechanism of SK3 and Orai1 is enabled by their close co-localization. Remarkably, we discovered that loss of SK3 channel activity due to overexpressed CaM mutants could be restored by Orai1, likely via its interplay with the SK3–CaM binding site. Mapping for interaction sites within Orai1, we identified that the cytosolic strands and pore residues are critical for a functional communication with SK3. Moreover, STIM1 has a bimodal role in SK3–Orai1 regulation. Under physiological ionic conditions, STIM1 is able to impede SK3–Orai1 interplay by significantly decreasing their co-localization. Forced STIM1–Orai1 activity and associated Ca2+ influx promote SK3 K+ currents. The dynamic regulation of Orai1 to boost endogenous SK3 channels was also determined in the human prostate cancer cell line LNCaP.

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Robinson, Lisa J., Salvatore Mancarella, Irina L. Tourkova, John B. Barnett, Harry C. Blair, and Jonathan Soboloff. "Critical Role for the Calcium-Release Activated Calcium Channel Orai1 In RANKL-Stimulated Osteoclast Formation From Monocytic Cells." Blood 116, no. 21 (November 19, 2010): 928. http://dx.doi.org/10.1182/blood.v116.21.928.928.

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Abstract Abstract 928 Calcium signals are major regulators of human osteoclast formation and function, and the molecular mechanisms underlying calcium effects are of interest as possible targets for pharmacologic regulation of bone resorption. IP3-receptor regulated release of calcium stores is linked to NFATc1 activation, which stimulates expression of key osteoclast genes in precursors, but the roles of other calcium channels in osteoclastogenesis are not clear. In particular, the identity of the channel(s) mediating extracellular calcium influx triggered by release of calcium stores remains uncertain. In lymphoid cells, a major mediator of this extracellular calcium influx is the Calcium-Release Activated Calcium (CRAC) channel consisting of Orai1, a plasma-membrane calcium channel, and the calcium-sensitive regulatory protein, STIM1. Calcium released from intracellular stores binds to a low affinity EF-hand in STIM1 causing a conformational change in STIM1 that permits binding to Orai1, aggregation in microscopically distinct puncta at the cell membrane, and opening of the Orai1 channel with consequent influx of extracellular calcium. Targeted deletion of Orai1 or Stim1 in mice results in severe immunodeficiency and early death; this has limited the assessment of Orai1 effects in other tissues. To evaluate the specific role of Orai1 in human osteoclasts, we used peripheral blood monocytes which form multinucleated osteoclasts capable of bone resorption when treated with CSF1 and RANKL. We confirmed Orai1 expression in human monocytes using Western blot and quantitative PCR assays, and found that the protein was down-regulated in mature osteoclasts. We used fura-2 to measure store-dependent and -independent changes in intracellular calcium during osteoclastic differentiation of monocytes over 10–14 days in RANKL and CSF1. RANKL-associated calcium oscillations were detected throughout differentiation, but calcium-release activated influx of extracellular calcium was markedly lower in the mature osteoclasts compared to precursors, paralleling their expression of Orai1. Human monocytes, transfected with Orai1-specific siRNA producing an 80% reduction in Orai1 protein compared to control siRNA treated cells, showed inhibition of store-regulated calcium influx during osteoclastogenesis. Furthermore, monocytes deficient in Orai1 showed impaired osteoclast formation; in particular, multinucleation resulting from osteoclast precursor cell fusion was markedly reduced, impairing bone resorption. Orai1 deficiency in T-cells inhibits activation of NFATc1, but this did not appear prominent in our cells: we found no significant inhibition of NFATc1 regulated gene expression in Orai1 siRNA-transfected cells compared to control siRNA-transfected cells, despite the marked difference in Orai1 protein. Other calcium channels may mediate calcium dependent NFATc1 activation in osteoclast precursors; alternatively, the low level of Orai1 protein remaining in Orai1 siRNA treated monocytes may be sufficient for NFATc1 activation. To define the effects of complete Orai1 deficiency, we examined osteoclast formation and in vivo skeletal development in mice with targeted deletion of the Orai1 gene (Gwack et al. Mol Cell Biol 28 (2008) 5209-22). Consistent with our in vitro results, multinucleated osteoclasts were nearly absent from Orai1-/- mice, but mononuclear cells expressing osteoclast markers such as TRAcP, were seen. Surprisingly, the knock-out mice did not show the osteopetrotic phenotype typical of osteoclast deficiency. Retention of fetal cartilage was seen, indicating defective osteoclastic function in Orai1-/- mice, but marked inhibition of bone formation was also present. Using micro-computed tomography we found significant reductions in both cortical ossification and trabecular bone formation in Orai1-/- mice. This raised the possibility of a previously unrecognized role for Orai1 in osteoblasts, or the osteoblast defect might simply reflect abnormalities of Orai1-/- osteoclasts and/or lymphocytes, since both cell types have regulatory effects on osteoblast formation and function. In summary, our studies identify a requirement for CRAC channel mediated calcium influx, and specifically the Orai1 channel, for normal formation and activity of human osteoclasts; these results are confirmed in an Orai1 knock-out mouse which also shows defective bone formation. Disclosures: No relevant conflicts of interest to declare.

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Peckys, Diana B., Daniel Gaa, Dalia Alansary, Barbara A. Niemeyer, and Niels de Jonge. "Supra-Molecular Assemblies of ORAI1 at Rest Precede Local Accumulation into Puncta after Activation." International Journal of Molecular Sciences 22, no. 2 (January 14, 2021): 799. http://dx.doi.org/10.3390/ijms22020799.

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The Ca2+ selective channel ORAI1 and endoplasmic reticulum (ER)-resident STIM proteins form the core of the channel complex mediating store operated Ca2+ entry (SOCE). Using liquid phase electron microscopy (LPEM), the distribution of ORAI1 proteins was examined at rest and after SOCE-activation at nanoscale resolution. The analysis of over seven hundred thousand ORAI1 positions revealed a number of ORAI1 channels had formed STIM-independent distinct supra-molecular clusters. Upon SOCE activation and in the presence of STIM proteins, a fraction of ORAI1 assembled in micron-sized two-dimensional structures, such as the known puncta at the ER plasma membrane contact zones, but also in divergent structures such as strands, and ring-like shapes. Our results thus question the hypothesis that stochastically migrating single ORAI1 channels are trapped at regions containing activated STIM, and we propose instead that supra-molecular ORAI1 clusters fulfill an amplifying function for creating dense ORAI1 accumulations upon SOCE-activation.

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Shawer, Heba, Chew W. Cheng, and Marc A. Bailey. "Absence of association between host genetic mutations in the ORAI1 gene and COVID-19 fatality." PLOS ONE 17, no. 2 (February 3, 2022): e0263303. http://dx.doi.org/10.1371/journal.pone.0263303.

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The calcium ion channel ORAI1 has emerged as a promising therapeutic target for the Coronavirus Disease 19 (COVID-19)-associated pneumonia, and a pharmacological inhibitor of ORAI1 has now reached clinical trials for severe COVID-19 pneumonia. Whether ORAI1 itself is associated with an increased risk for severe COVID-19 presentation is still unknown. Here, we employed genetic association analysis to investigate the potential association of host genetic polymorphisms of ORAI1 with the risk of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection and its associated COVID-19 fatality in UK Biobank participants from white British background. The analysis showed no significant association between ORAI1 variants and COVID-19 positivity or fatality, despite the well-established roles of ORAI1 in immune response and inflammation and the success of ORAI1 inhibition in clinical trials. Our results suggest that the host genetic polymorphisms of ORAI1 are unlikely to be implicated in the broad variability in symptoms severity among afflicted patients.

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Lin, Lin, Jinjin Wei, Zheng Chen, Xinyue Tang, Fei Dai, and Guangbin Sun. "Intervention of Orai1 Influences the Response of Nuocytes From Allergic Rhinitis Mice to IL-33." Annals of Otology, Rhinology & Laryngology 128, no. 9 (May 2019): 838–47. http://dx.doi.org/10.1177/0003489419846142.

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Objective: Nuocytes are essential in innate type-2 immunity and contribute to the exacerbation of allergic rhinitis (AR). This study aimed to evaluate the intervention of Orai1 on the response of nuocytes from AR mice to interleukin (IL)-33. Methods: We established a murine model of AR. Nuocytes were obtained from the mouse nasal-associated lymphoid tissue. Then, we assessed expressions of Orai1, Ca2+ mean fluorescence intensity (MFI) in nuocytes, and their cellular response to mouse recombinant (rm) IL-33. After that, we administered rmlentivirus vectors (lenti) that encoded small hairpin RNA (shRNA) against ORAI1 (lenti-ORAI1) into nuocytes cultures and again evaluated Orai1 and Ca2+ MFI in nuocytes and their response to rmIL-33. Finally, we adoptively transferred nuocytes alone or nuocytes transfected by lenti or lenti-ORAI1 to AR models to investigate their roles during allergic inflammation. Results: We showed that Orai1 and Ca2+ MFI were upregulated in AR mice nuocytes. These cells were induced to produce more IL-5 and IL-13 by rmIL-33. However, the intervention of Orai1 by lenti-ORAI1 in nuocytes decreased Orai1 and Ca2+ MFI and reduced productions of aforementioned cytokines even after the administration of rmIL-33. Numbers of sneezing, nasal rubbing, and counts of eosinophils were all enhanced after the adoptive transfer of nuocytes. Concentrations of IL-5, IL-13, and IL-33 in the nasal lavage fluid (NLF) of allergic mice were also increased. However, the adoptive transfer of nuocytes transfected by lenti-ORAI1 decreased aforementioned parameters. Conclusion: These findings show that the intervention of Orai1 in nuocytes influences the response of nuocytes to rmIL-33.

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Radoslavova, Silviya, Antoine Folcher, Thibaut Lefebvre, Kateryna Kondratska, Stéphanie Guénin, Isabelle Dhennin-Duthille, Mathieu Gautier, Natalia Prevarskaya та Halima Ouadid-Ahidouch. "Orai1 Channel Regulates Human-Activated Pancreatic Stellate Cell Proliferation and TGFβ1 Secretion through the AKT Signaling Pathway". Cancers 13, № 10 (15 травня 2021): 2395. http://dx.doi.org/10.3390/cancers13102395.

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Activated pancreatic stellate cells (aPSCs), the crucial mediator of pancreatic desmoplasia, are characterized, among others, by high proliferative potential and abundant transforming growth factor β1 (TGFβ1) secretion. Over the past years, the involvement of Ca2+ channels in PSC pathophysiology has attracted great interest in pancreatic cancer research. We, thus, aimed to investigate the role of the Orai1 Ca2+ channel in these two PSC activation processes. Using the siRNA approach, we invalided Orai1 expression and assessed the channel functionality by Ca2+ imaging, the effect on aPSC proliferation, and TGFβ1 secretion. We demonstrated the functional expression of the Orai1 channel in human aPSCs and its implication in the store-operated Ca2+ entry (SOCE). Orai1 silencing led to a decrease in aPSC proliferation, TGFβ1 secretion, and AKT activation. Interestingly, TGFβ1 induced a higher SOCE response by increasing Orai1 mRNAs and proteins and promoted both AKT phosphorylation and cell proliferation, abolished by Orai1 silencing. Together, our results highlight the role of Orai1-mediated Ca2+ entry in human aPSC pathophysiology by controlling cell proliferation and TGFβ1 secretion through the AKT signaling pathway. Moreover, we showed a TGFβ1-induced autocrine positive feedback loop by promoting the Orai1/AKT-dependent proliferation via the stimulation of Orai1 expression and function.

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Huang, Ping-Chun, Tai-Yu Chiu, Li-Chun Wang, Hsiao-Chuan Teng, Fu-Jen Kao, and De-Ming Yang. "Visualization of the Orai1 Homodimer and the Functional Coupling of Orai1-STIM1 by Live-Cell Fluorescence Lifetime Imaging." Microscopy and Microanalysis 16, no. 3 (April 9, 2010): 313–26. http://dx.doi.org/10.1017/s1431927610000188.

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AbstractThe Orai1-STIM1 constructed store-operated Ca2+ channels (SOCs) have been found to exert several essential Ca2+ entry/signaling cascades, e.g., the generation of immune response in T lymphocytes. Although biochemical and novel imaging evidence appear to indicate that Orai1 and STIM1 interact with each other to achieve store-operated Ca2+ entry (SOCE), the detailed mechanism of functional SOCE in situ has yet to be fully understood. In this study, green fluorescence protein (EGFP as donor) targeted to either the N- or C-terminal of Orai1 (wild type or ▵1-90+▵267-301 double deletion type) and mOrange (as acceptor) tagged STIM1 were used to comprise a fluorescence resonance energy transfer (FRET) pair within living PC12 cells. The fluorescence lifetime map and histogram/distribution of each single cell, determined by one-photon excitation fluorescence lifetime imaging microscopy (FLIM), was used to visualize FRET and show the Orai1 homodimer and Orai1-STIM1 binding. Both the color-coded lifetime map and the distribution of EGFP-tagged Orai1 significantly changed after the administration of thapsigargin, the SOCE stimulating agent. The FRET efficiency from each experimental set was also calculated and compared using double exponential analysis. In summary, we show the detailed interactions Orai1-Orai1 and Orai1-STIM1 within intact living cells by using the FLIM-FRET technique.

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Yu, Tao, Shangbin Chen, Jingying Pan, Conglin Su, and Jun He. "Optical investigations reveal the effects of 2-aminoethyldiphenyl borate on STIM1 puncta formation." Journal of Innovative Optical Health Sciences 11, no. 02 (February 19, 2018): 1850003. http://dx.doi.org/10.1142/s1793545818500037.

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2-Aminoethyldiphenyl borate (2-APB) is the most commonly used pharmacological agent in the study of calcium release-activated channels (CRACs); however, its inhibitory mechanism to CRACs remains unclear. To address this issue, we systematically employed confocal imaging, dual-wavelength excitation photometry and FRET to examine the effects of 2-APB on the dynamic activities and function of STIM1 and Orai1, two key components of CRACs. Imaging results support that there are two signaling pathways (Orai1-independent and Orai1-dependent) for the formation of STIM1 puncta. 2-APB could dose dependently block Orai1-independent but not Orai1-dependent STIM1 puncta formation, despite its obvious inhibition effect on store-operated Ca[Formula: see text] entry (SOCE). In addition, we found that although 2-APB could not visibly alter near plasma membrane CAD-eYFP localization, it could completely block CAD-YFP-induced constitutive Ca[Formula: see text] entry and promote the interaction between Orai1 and CAD by FRET measurements. Therefore, we proposed that inhibitory action of 2-APB on SOCE might attribute to its direct inhibitory effects on Orai1 channel itself, but not the interference on puncta formation between STIM1 and Orai1.

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Kim, Kyun-Do, Sonal Srikanth, Yossan-Var Tan, Ma-Khin Yee, Marcus Jew, Robert Damoiseaux, Michael E. Jung, et al. "Small molecule blockers reveal an important role for Orai1-NFAT-orphan receptor pathway in Th17 differentiation (P5169)." Journal of Immunology 190, no. 1_Supplement (May 1, 2013): 68.12. http://dx.doi.org/10.4049/jimmunol.190.supp.68.12.

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Abstract Orai1 is the pore subunit of the Calcium Release Activated Channels (CRAC) that activate downstream calcineurin-NFAT pathway. Loss of function mutation in Orai1 causes lethality due to severe defects in immune system, which can be rescued by bone marrow transplantation. Using a high throughput chemical library screening, we have identified a class of small molecule blockers that inhibit Orai1. One of the small molecules, compound 5D blocked the activity of a constitutively active mutant of Orai1, suggesting a direct block of ion permeation via Orai1. Primary murine Th17 cells showed highest sensitivity to block by this compound. Treatment with compound 5D not only suppressed cytokine production, but also inhibited differentiation of Th17 cells by suppression of the retinoic-acid-receptor-related orphan receptors RORα and RORγt. Exogenous expression of RORα, RORγt or a constitutive active mutant of NFAT (CA-NFAT) rescued the blocking effect, identifying the Orai1-NFAT- ROR signaling pathway to be important for Th17 differentiation. The defects in cytokine production and differentiation were recapitulated in Th17 cells from Orai1-deficient mice suggesting Orai1 as a specific target of the blockers. In vivo administration of Orai1 blockers effectively delayed the onset and reduced the severity of autoimmune disease in mice. These data indicate that derivatives of compound 5D can be used as chemical templates for development of therapeutic agents to alleviate autoimmune diseases.

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Jiang, Hui, Shubiao Zou, Sarika Chaudhari, and Rong Ma. "Short-term high-glucose treatment decreased abundance of Orai1 protein through posttranslational mechanisms in rat mesangial cells." American Journal of Physiology-Renal Physiology 314, no. 5 (May 1, 2018): F855—F863. http://dx.doi.org/10.1152/ajprenal.00513.2017.

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The short-term effect of high-glucose (HG) treatment on store-operated Ca2+ entry in mesangial cells (MCs) is not well-known. The aim of the present study was to determine whether and how HG treatment for a short period altered protein abundance of Orai1, the channel mediating store-operated Ca2+ entry in MCs. Rat and human MCs were exposed to HG (25 mM) for 2, 4, 8, and 24 h, and the abundance of Orai1 protein was significantly decreased at the time points of 8 and 16 h. Consistently, HG treatment for 8 h significantly reduced store-operated Ca2+ entry in rat MCs. However, HG treatment for the same time periods did not alter the levels of Orai1 transcript. Cycloheximide, a protein synthesis inhibitor, did not affect the HG-induced decrease of Orai1 protein, suggesting a posttranslational mechanism was involved. However, the HG effect on Orai1 protein was significantly attenuated by MG132 (a ubiquitin-proteasome inhibitor) and NH4Cl (a lysosomal pathway inhibitor). Furthermore, HG treatment for 8 h stimulated ubiquitination of Orai1 protein. We further found that polyethylene glycol-catalase, an antioxidant, significantly blunted the HG-induced reduction of Orai1 protein. In support of involvement of reactive oxygen species in the HG effects, hydrogen peroxide (H2O2) itself significantly decreased abundance of Orai1 protein and increased the level of ubiquitinated Orai1. Taken together, these results suggest that a short-term HG treatment decreased abundance of Orai1 protein in MCs by promoting the protein degradation through the ubiquitination-proteasome and –lysosome mechanisms. This HG-stimulated posttranslational mechanism was mediated by H2O2.

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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.

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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.

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41

Bartoli, Fiona, Marc A. Bailey, Baptiste Rode, Philippe Mateo, Fabrice Antigny, Kaveen Bedouet, Pascale Gerbaud, et al. "Orai1 Channel Inhibition Preserves Left Ventricular Systolic Function and Normal Ca 2+ Handling After Pressure Overload." Circulation 141, no. 3 (January 21, 2020): 199–216. http://dx.doi.org/10.1161/circulationaha.118.038891.

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Background: Orai1 is a critical ion channel subunit, best recognized as a mediator of store-operated Ca 2+ entry (SOCE) in nonexcitable cells. SOCE has recently emerged as a key contributor of cardiac hypertrophy and heart failure but the relevance of Orai1 is still unclear. Methods: To test the role of these Orai1 channels in the cardiac pathophysiology, a transgenic mouse was generated with cardiomyocyte-specific expression of an ion pore-disruptive Orai1 R91W mutant (C-dnO1). Synthetic chemistry and channel screening strategies were used to develop 4-(2,5-dimethoxyphenyl)-N-[(pyridin-4-yl)methyl]aniline (hereafter referred to as JPIII), a small-molecule Orai1 channel inhibitor suitable for in vivo delivery. Results: Adult mice subjected to transverse aortic constriction (TAC) developed cardiac hypertrophy and reduced ventricular function associated with increased Orai1 expression and Orai1-dependent SOCE (assessed by Mn 2+ influx). C-dnO1 mice displayed normal cardiac electromechanical function and cellular excitation-contraction coupling despite reduced Orai1-dependent SOCE. Five weeks after TAC, C-dnO1 mice were protected from systolic dysfunction (assessed by preserved left ventricular fractional shortening and ejection fraction) even if increased cardiac mass and prohypertrophic markers induction were observed. This is correlated with a protection from TAC-induced cellular Ca 2+ signaling alterations (increased SOCE, decreased [Ca 2+ ] i transients amplitude and decay rate, lower SR Ca 2+ load and depressed cellular contractility) and SERCA2a downregulation in ventricular cardiomyocytes from C-dnO1 mice, associated with blunted Pyk2 signaling. There was also less fibrosis in heart sections from C-dnO1 mice after TAC. Moreover, 3 weeks treatment with JPIII following 5 weeks of TAC confirmed the translational relevance of an Orai1 inhibition strategy during hypertrophic insult. Conclusions: The findings suggest a key role of cardiac Orai1 channels and the potential for Orai1 channel inhibitors as inotropic therapies for maintaining contractility reserve after hypertrophic stress.

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42

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.

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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.

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43

Wang, Lei, Jiaqi Hao, Yijian Zhang, Ziyi Yang, Yang Cao, Wei Lu, Yijun Shu, et al. "Orai1 mediates tumor-promoting store-operated Ca2+ entry in human gastrointestinal stromal tumors via c-KIT and the extracellular signal–regulated kinase pathway." Tumor Biology 39, no. 2 (February 2017): 101042831769142. http://dx.doi.org/10.1177/1010428317691426.

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Gastrointestinal stromal tumors originate from interstitial cells of Cajal, the pacemaker cells of the gut. Ca2+ regulates the pacemaker activity of interstitial cells of Cajal. Store-operated Ca2+ entry mediates the majority of Ca2+ entry in most cancer cells and may be a factor in regulating intracellular Ca2+ in interstitial cells of Cajal and gastrointestinal stromal tumors. Therefore, a blockade of this mechanism may affect the progression of gastrointestinal stromal tumors. Orai1 is the pore subunit of store-operated Ca2+ channels. Here, we reported that Orai1 was overexpressed in gastrointestinal stromal tumor tissues and was positively correlated with a high-risk grade in gastrointestinal stromal tumor patients. Furthermore, upon Orai1 silencing, the functional store-operated Ca2+ entry in gastrointestinal stromal tumor cells was decreased, indicating that the function of store-operated Ca2+ entry was mediated by Orai1. Inhibition of Orai1-mediated store-operated Ca2+ entry by Orai1 silencing or store-operated Ca2+ entry blockers (SKF-96365 and 2-aminoethyl diphenylborate) induced obvious cell proliferation suppression, cell-cycle distribution, and apoptosis stimulation in GIST-T1 cells. Conversely, Orai1 overexpression increased store-operated Ca2+ entry and cell proliferation in GIST882 cells. In addition, we found that activation of c-KIT and the extracellular signal–regulated kinase pathway participated in the oncogenic functions of Orai1-mediated store-operated Ca2+ entry in gastrointestinal stromal tumor cells. These results revealed that Orai1-mediated store-operated Ca2+ entry is critical for gastrointestinal stromal tumor cell proliferation via c-KIT and ERK signaling pathway activation. Orai1-mediated store-operated Ca2+ entry plays an oncogenic role and may be a novel prognostic factor and therapeutic target for patients with gastrointestinal stromal tumors.

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44

Tiffner, Adéla, Lena Maltan, Sarah Weiß, and Isabella Derler. "The Orai Pore Opening Mechanism." International Journal of Molecular Sciences 22, no. 2 (January 7, 2021): 533. http://dx.doi.org/10.3390/ijms22020533.

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Cell survival and normal cell function require a highly coordinated and precise regulation of basal cytosolic Ca2+ concentrations. The primary source of Ca2+ entry into the cell is mediated by the Ca2+ release-activated Ca2+ (CRAC) channel. Its action is stimulated in response to internal Ca2+ store depletion. The fundamental constituents of CRAC channels are the Ca2+ sensor, stromal interaction molecule 1 (STIM1) anchored in the endoplasmic reticulum, and a highly Ca2+-selective pore-forming subunit Orai1 in the plasma membrane. The precise nature of the Orai1 pore opening is currently a topic of intensive research. This review describes how Orai1 gating checkpoints in the middle and cytosolic extended transmembrane regions act together in a concerted manner to ensure an opening-permissive Orai1 channel conformation. In this context, we highlight the effects of the currently known multitude of Orai1 mutations, which led to the identification of a series of gating checkpoints and the determination of their role in diverse steps of the Orai1 activation cascade. The synergistic action of these gating checkpoints maintains an intact pore geometry, settles STIM1 coupling, and governs pore opening. We describe the current knowledge on Orai1 channel gating mechanisms and summarize still open questions of the STIM1–Orai1 machinery.

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45

Silva-Rojas, Roberto, Laura Pérez-Guàrdia, Emma Lafabrie, David Moulaert, Jocelyn Laporte, and Johann Böhm. "Silencing of the Ca2+ Channel ORAI1 Improves the Multi-Systemic Phenotype of Tubular Aggregate Myopathy (TAM) and Stormorken Syndrome (STRMK) in Mice." International Journal of Molecular Sciences 23, no. 13 (June 23, 2022): 6968. http://dx.doi.org/10.3390/ijms23136968.

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Tubular aggregate myopathy (TAM) and Stormorken syndrome (STRMK) form a clinical continuum associating progressive muscle weakness with additional multi-systemic anomalies of the bones, skin, spleen, and platelets. TAM/STRMK arises from excessive extracellular Ca2+ entry due to gain-of-function mutations in the Ca2+ sensor STIM1 or the Ca2+ channel ORAI1. Currently, no treatment is available. Here we assessed the therapeutic potential of ORAI1 downregulation to anticipate and reverse disease development in a faithful mouse model carrying the most common TAM/STRMK mutation and recapitulating the main signs of the human disorder. To this aim, we crossed Stim1R304W/+ mice with Orai1+/− mice expressing 50% of ORAI1. Systematic phenotyping of the offspring revealed that the Stim1R304W/+Orai1+/− mice were born with a normalized ratio and showed improved postnatal growth, bone architecture, and partly ameliorated muscle function and structure compared with their Stim1R304W/+ littermates. We also produced AAV particles containing Orai1-specific shRNAs, and intramuscular injections of Stim1R304W/+ mice improved the skeletal muscle contraction and relaxation properties, while muscle histology remained unchanged. Altogether, we provide the proof-of-concept that Orai1 silencing partially prevents the development of the multi-systemic TAM/STRMK phenotype in mice, and we also established an approach to target Orai1 expression in postnatal tissues.

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46

Tiffner, Adéla, Lena Maltan, Sarah Weiß, and Isabella Derler. "The Orai Pore Opening Mechanism." International Journal of Molecular Sciences 22, no. 2 (January 7, 2021): 533. http://dx.doi.org/10.3390/ijms22020533.

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Анотація:

Cell survival and normal cell function require a highly coordinated and precise regulation of basal cytosolic Ca2+ concentrations. The primary source of Ca2+ entry into the cell is mediated by the Ca2+ release-activated Ca2+ (CRAC) channel. Its action is stimulated in response to internal Ca2+ store depletion. The fundamental constituents of CRAC channels are the Ca2+ sensor, stromal interaction molecule 1 (STIM1) anchored in the endoplasmic reticulum, and a highly Ca2+-selective pore-forming subunit Orai1 in the plasma membrane. The precise nature of the Orai1 pore opening is currently a topic of intensive research. This review describes how Orai1 gating checkpoints in the middle and cytosolic extended transmembrane regions act together in a concerted manner to ensure an opening-permissive Orai1 channel conformation. In this context, we highlight the effects of the currently known multitude of Orai1 mutations, which led to the identification of a series of gating checkpoints and the determination of their role in diverse steps of the Orai1 activation cascade. The synergistic action of these gating checkpoints maintains an intact pore geometry, settles STIM1 coupling, and governs pore opening. We describe the current knowledge on Orai1 channel gating mechanisms and summarize still open questions of the STIM1–Orai1 machinery.

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47

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.

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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.

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48

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.

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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.

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49

Ng, Lih Chyuan, Deepa Ramduny, Judith A. Airey, Cherie A. Singer, Phillip S. Keller, Xiao-Ming Shen, Honglin Tian, Maria Valencik, and Joseph R. Hume. "Orai1 interacts with STIM1 and mediates capacitative Ca2+ entry in mouse pulmonary arterial smooth muscle cells." American Journal of Physiology-Cell Physiology 299, no. 5 (November 2010): C1079—C1090. http://dx.doi.org/10.1152/ajpcell.00548.2009.

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Previous studies in mouse pulmonary arterial smooth muscle cells (PASMCs) showed that cannonical transient receptor potential channel TRPC1 and stromal interaction molecule 1 (STIM1) mediate the sustained component of capacitative Ca2+ entry (CCE), but the molecular candidate(s) that mediate the transient component of CCE remain unknown. The aim of the present study was to examine whether Orai1 mediates the transient component of CCE through activation of STIM1 in mouse PASMCs. In primary cultured mouse PASMCs loaded with fura-2, cyclopiazonic acid (CPA) caused a transient followed by a sustained rise in intracellular Ca2+ concentration ([Ca2+]i). The transient but not the sustained rise in [Ca2+]i was partially inhibited by nifedipine. The nifedipine-insensitive transient rise in [Ca2+]i and the increase in Mn2+ quench of fura-2 fluorescence caused by CPA were both reduced in cells treated with Orai1 siRNA. These responses to CPA were further reduced in cells treated with Orai1 and STIM1 small interfering (si)RNA. Moreover, overexpression of STIM1 enhanced the rise in [Ca2+]i and the increase in Mn2+ quench of fura-2 fluorescence caused by CPA, and these responses were reduced in cells treated with Orai1 siRNA. RT-PCR revealed Orai1 and STIM1 mRNAs, and Western blot analysis identified Orai1 and STIM1 proteins in mouse PASMCs. Furthermore, Orai1 was found to coimmunoprecipitate with STIM1, and the precipitation level of Orai1 was increased in cells subjected to store-depletion. Immunostaining revealed colocalization of Orai1 and STIM1 proteins, and the colocalization of these proteins was more apparent after store-depletion. These data provide direct evidence that the transient component of CCE is mediated by Orai1 channel as a result of STIM1 activation in mouse PASMCs.

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50

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.

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