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STIM1, an essential and conserved component of store-operated Ca²⁺ channel function

¹ Torrey Pines Therapeutics, Inc., La Jolla, CA 92037
² Department of Physiology and Biophysics and Center for Immunology, University of California, Irvine, CA 92697

Correspondence to K.A. Stauderman: kstauderman{at}torreypinestherapeutics.com; or M.D. Cahalan: mcahalan{at}uci.edu

Store-operated Ca²⁺ (SOC) channels regulate many cellular processes, but the underlying molecular components are not well defined. Using an RNA interference (RNAi)-based screen to identify genes that alter thapsigargin (TG)-dependent Ca²⁺ entry, we discovered a required and conserved role of Stim in SOC influx. RNAi-mediated knockdown of Stim in Drosophila S2 cells significantly reduced TG-dependent Ca²⁺ entry. Patch-clamp recording revealed nearly complete suppression of the Drosophila Ca²⁺ release-activated Ca²⁺ (CRAC) current that has biophysical characteristics similar to CRAC current in human T cells. Similarly, knockdown of the human homologue STIM1 significantly reduced CRAC channel activity in Jurkat T cells. RNAi-mediated knockdown of STIM1 inhibited TG- or agonist-dependent Ca²⁺ entry in HEK293 or SH-SY5Y cells. Conversely, overexpression of STIM1 in HEK293 cells modestly enhanced TG-induced Ca²⁺ entry. We propose that STIM1, a ubiquitously expressed protein that is conserved from Drosophila to mammalian cells, plays an essential role in SOC influx and may be a common component of SOC and CRAC channels.

Abbreviations used in this paper: [Ca²⁺]_i, intracellular free Ca²⁺ concentration; CRAC, Ca²⁺ release-activated Ca²⁺; dsRNA, double-stranded RNA; iPLA₂, Ca²⁺-independent PLA₂; RFU, relative fluorescence unit; RNAi, RNA interference; SOC, store-operated Ca²⁺; SOCE, SOC entry; TG, thapsigargin.

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• Smyth, J. T., DeHaven, W. I., Bird, G. S., Putney, J. W. Jr (2007). Role of the microtubule cytoskeleton in the function of the store-operated Ca2+ channel activator STIM1. J. Cell Sci. 120: 3762-3771 [Abstract] [Full Text]  
• Liu, X., Cheng, K. T., Bandyopadhyay, B. C., Pani, B., Dietrich, A., Paria, B. C., Swaim, W. D., Beech, D., Yildrim, E., Singh, B. B., Birnbaumer, L., Ambudkar, I. S. (2007). Attenuation of store-operated Ca2+ current impairs salivary gland fluid secretion in TRPC1( / ) mice. Proc. Natl. Acad. Sci. USA 104: 17542-17547 [Abstract] [Full Text]  
• Draberova, L., Shaik, G. M., Volna, P., Heneberg, P., Tumova, M., Lebduska, P., Korb, J., Draber, P. (2007). Regulation of Ca2+ Signaling in Mast Cells by Tyrosine-Phosphorylated and Unphosphorylated Non-T Cell Activation Linker. J. Immunol. 179: 5169-5180 [Abstract] [Full Text]  
• Varnai, P., Toth, B., Toth, D. J., Hunyady, L., Balla, T. (2007). Visualization and Manipulation of Plasma Membrane-Endoplasmic Reticulum Contact Sites Indicates the Presence of Additional Molecular Components within the STIM1-Orai1 Complex. J. Biol. Chem. 282: 29678-29690 [Abstract] [Full Text]  
• Li, Z., Lu, J., Xu, P., Xie, X., Chen, L., Xu, T. (2007). Mapping the Interacting Domains of STIM1 and Orai1 in Ca2+ Release-activated Ca2+ Channel Activation. J. Biol. Chem. 282: 29448-29456 [Abstract] [Full Text]  
• Hong, M., Ross, W. N. (2007). Priming of intracellular calcium stores in rat CA1 pyramidal neurons. J. Physiol. 584: 75-87 [Abstract] [Full Text]  
• Nishida, M., Onohara, N., Sato, Y., Suda, R., Ogushi, M., Tanabe, S., Inoue, R., Mori, Y., Kurose, H. (2007). G{alpha}12/13-mediated Up-regulation of TRPC6 Negatively Regulates Endothelin-1-induced Cardiac Myofibroblast Formation and Collagen Synthesis through Nuclear Factor of Activated T Cells Activation. J. Biol. Chem. 282: 23117-23128 [Abstract] [Full Text]  
• Kaznacheyeva, E., Glushankova, L., Bugaj, V., Zimina, O., Skopin, A., Alexeenko, V., Tsiokas, L., Bezprozvanny, I., Mozhayeva, G. N. (2007). Suppression of TRPC3 Leads to Disappearance of Store-operated Channels and Formation of a New Type of Store-independent Channels in A431 Cells. J. Biol. Chem. 282: 23655-23662 [Abstract] [Full Text]  
• Hirota, S., Helli, P., Janssen, L. J. (2007). Ionic mechanisms and Ca2+ handling in airway smooth muscle. Eur Respir J 30: 114-133 [Abstract] [Full Text]  
• DeHaven, W. I., Smyth, J. T., Boyles, R. R., Putney, J. W. Jr. (2007). Calcium Inhibition and Calcium Potentiation of Orai1, Orai2, and Orai3 Calcium Release-activated Calcium Channels. J. Biol. Chem. 282: 17548-17556 [Abstract] [Full Text]  
• Putney, J. W. Jr (2007). New molecular players in capacitative Ca2+ entry. J. Cell Sci. 120: 1959-1965 [Abstract] [Full Text]  
• Gwack, Y., Srikanth, S., Feske, S., Cruz-Guilloty, F., Oh-hora, M., Neems, D. S., Hogan, P. G., Rao, A. (2007). Biochemical and Functional Characterization of Orai Proteins. J. Biol. Chem. 282: 16232-16243 [Abstract] [Full Text]  
• Liou, J., Fivaz, M., Inoue, T., Meyer, T. (2007). Live-cell imaging reveals sequential oligomerization and local plasma membrane targeting of stromal interaction molecule 1 after Ca2+ store depletion. Proc. Natl. Acad. Sci. USA 104: 9301-9306 [Abstract] [Full Text]  
• Chung, S. C., Limnander, A., Kurosaki, T., Weiss, A., Korenbrot, J. I. (2007). Coupling Ca2+ store release to Icrac channel activation in B lymphocytes requires the activity of Lyn and Syk kinases. JCB 177: 317-328 [Abstract] [Full Text]  
• Ong, H. L., Liu, X., Tsaneva-Atanasova, K., Singh, B. B., Bandyopadhyay, B. C., Swaim, W. D., Russell, J. T., Hegde, R. S., Sherman, A., Ambudkar, I. S. (2007). Relocalization of STIM1 for Activation of Store-operated Ca2+ Entry Is Determined by the Depletion of Subplasma Membrane Endoplasmic Reticulum Ca2+ Store. J. Biol. Chem. 282: 12176-12185 [Abstract] [Full Text]  

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