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Orai1-Orai2 complex is involved in store-operated calcium entry in chondrocyte cell lines.
Cell Calcium. 2015 May; 57(5-6):337-47.CC

Abstract

Ca(2+) influx via store-operated Ca(2+) entry (SOCE) plays critical roles in many essential cellular functions. The Ca(2+) release-activated Ca(2+) (CRAC) channel complex, consisting of Orai and STIM, is one of the major components of store-operated Ca(2+) (SOC) channels. Our previous study demonstrated that histamine can cause sustained Ca(2+) entry through SOC channels in OUMS-27 cells derived from human chondrosarcoma. This SOCE was increased by low- and decreased by high-concentrations of 2-aminoethoxydiphenyl borate. Quantitative reverse transcription PCR and Western blot analyses revealed abundant expressions of Orai1, Orai2 and STIM1. Introduction of dominant negative mutant of Orai1, or siOrai1 knockdown significantly attenuated SOCE. Following histamine application, single molecule imaging using total internal reflection fluorescence (TIRF) microscopy demonstrated punctate Orai1-STIM1 complex formation in plasma membrane. In contrast, knockdown or over-expression of Orai2 resulted in an increase or a decrease in SOCE, respectively. Finally, TIRF imaging revealed direct coupling between Orai1 and Orai2, and suggested that Orai2 reduces Orai1 function by formation of a hetero-tetramer. These results provide substantial evidence that Orai1, Orai2 and STIM1 form functional CRAC channels in OUMS-27 cells and that these complexes are responsible for sustained Ca(2+) entry in response to agonist stimulation.

Authors+Show Affiliations

Department of Molecular & Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya 467-8603, Japan.Department of Molecular & Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya 467-8603, Japan.Department of Molecular & Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya 467-8603, Japan.Department of Molecular & Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya 467-8603, Japan.Department of Molecular & Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya 467-8603, Japan.Department of Molecular & Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya 467-8603, Japan; Department of Pharmacology, Division of Pathological Sciences, Kyoto Pharmaceutical University, Kyoto 607-8414, Japan.Faculty of Kinesiology, University of Calgary, Calgary, Alberta T2N 1N4, Canada.Department of Molecular & Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya 467-8603, Japan. Electronic address: yimaizum@phar.nagoya-cu.ac.jp.

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

25769459

Citation

Inayama, Munenori, et al. "Orai1-Orai2 Complex Is Involved in Store-operated Calcium Entry in Chondrocyte Cell Lines." Cell Calcium, vol. 57, no. 5-6, 2015, pp. 337-47.
Inayama M, Suzuki Y, Yamada S, et al. Orai1-Orai2 complex is involved in store-operated calcium entry in chondrocyte cell lines. Cell Calcium. 2015;57(5-6):337-47.
Inayama, M., Suzuki, Y., Yamada, S., Kurita, T., Yamamura, H., Ohya, S., Giles, W. R., & Imaizumi, Y. (2015). Orai1-Orai2 complex is involved in store-operated calcium entry in chondrocyte cell lines. Cell Calcium, 57(5-6), 337-47. https://doi.org/10.1016/j.ceca.2015.02.005
Inayama M, et al. Orai1-Orai2 Complex Is Involved in Store-operated Calcium Entry in Chondrocyte Cell Lines. Cell Calcium. 2015;57(5-6):337-47. PubMed PMID: 25769459.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Orai1-Orai2 complex is involved in store-operated calcium entry in chondrocyte cell lines. AU - Inayama,Munenori, AU - Suzuki,Yoshiaki, AU - Yamada,Satoshi, AU - Kurita,Takashi, AU - Yamamura,Hisao, AU - Ohya,Susumu, AU - Giles,Wayne R, AU - Imaizumi,Yuji, Y1 - 2015/02/19/ PY - 2014/08/31/received PY - 2015/02/01/revised PY - 2015/02/11/accepted PY - 2015/3/15/entrez PY - 2015/3/15/pubmed PY - 2016/1/20/medline KW - Chondrocyte KW - Orai1 KW - Orai2 KW - Store-operated calcium entry (SOCE) KW - Stromal interaction molecule 1 (STIM1) KW - Total internal reflection fluorescence (TIRF) microscopy SP - 337 EP - 47 JF - Cell calcium JO - Cell Calcium VL - 57 IS - 5-6 N2 - Ca(2+) influx via store-operated Ca(2+) entry (SOCE) plays critical roles in many essential cellular functions. The Ca(2+) release-activated Ca(2+) (CRAC) channel complex, consisting of Orai and STIM, is one of the major components of store-operated Ca(2+) (SOC) channels. Our previous study demonstrated that histamine can cause sustained Ca(2+) entry through SOC channels in OUMS-27 cells derived from human chondrosarcoma. This SOCE was increased by low- and decreased by high-concentrations of 2-aminoethoxydiphenyl borate. Quantitative reverse transcription PCR and Western blot analyses revealed abundant expressions of Orai1, Orai2 and STIM1. Introduction of dominant negative mutant of Orai1, or siOrai1 knockdown significantly attenuated SOCE. Following histamine application, single molecule imaging using total internal reflection fluorescence (TIRF) microscopy demonstrated punctate Orai1-STIM1 complex formation in plasma membrane. In contrast, knockdown or over-expression of Orai2 resulted in an increase or a decrease in SOCE, respectively. Finally, TIRF imaging revealed direct coupling between Orai1 and Orai2, and suggested that Orai2 reduces Orai1 function by formation of a hetero-tetramer. These results provide substantial evidence that Orai1, Orai2 and STIM1 form functional CRAC channels in OUMS-27 cells and that these complexes are responsible for sustained Ca(2+) entry in response to agonist stimulation. SN - 1532-1991 UR - https://www.unboundmedicine.com/medline/citation/25769459/Orai1_Orai2_complex_is_involved_in_store_operated_calcium_entry_in_chondrocyte_cell_lines_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0143-4160(15)00037-8 DB - PRIME DP - Unbound Medicine ER -