Zhang, S. L. et al. Genome-wide RNAi screen of Ca2+ influx identifies genes that regulate Ca2+ release-activated Ca2+ channel activity. Proc. Natl Acad. Sci. USA 103, 9357-9362

Department of Physiology and Biophysics, University of California, Irvine, CA 92697, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.81). 07/2006; 103(24):9357-62. DOI: 10.1073/pnas.0603161103
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ABSTRACT Recent studies by our group and others demonstrated a required and conserved role of Stim in store-operated Ca(2+) influx and Ca(2+) release-activated Ca(2+) (CRAC) channel activity. By using an unbiased genome-wide RNA interference screen in Drosophila S2 cells, we now identify 75 hits that strongly inhibited Ca(2+) influx upon store emptying by thapsigargin. Among these hits are 11 predicted transmembrane proteins, including Stim, and one, olf186-F, that upon RNA interference-mediated knockdown exhibited a profound reduction of thapsigargin-evoked Ca(2+) entry and CRAC current, and upon overexpression a 3-fold augmentation of CRAC current. CRAC currents were further increased to 8-fold higher than control and developed more rapidly when olf186-F was cotransfected with Stim. olf186-F is a member of a highly conserved family of four-transmembrane spanning proteins with homologs from Caenorhabditis elegans to human. The endoplasmic reticulum (ER) Ca(2+) pump sarco-/ER calcium ATPase (SERCA) and the single transmembrane-soluble N-ethylmaleimide-sensitive (NSF) attachment receptor (SNARE) protein Syntaxin5 also were required for CRAC channel activity, consistent with a signaling pathway in which Stim senses Ca(2+) depletion within the ER, translocates to the plasma membrane, and interacts with olf186-F to trigger CRAC channel activity.

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Available from: Ying Yu, Aug 30, 2015
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    • "Store-operated Ca 2+ entry (SOCE) is an evolutionarily conserved signaling mechanism induced by the Ca 2+ depletion of the endoplasmic reticulum (ER) that sustains long-lasting cytosolic Ca 2+ signals required for transcription, cell proliferation and effector function (Hogan et al., 2010; Parekh, 2010). SOCE is mediated by the ER Ca 2+ sensors stromal interaction molecules (STIMs) STIM1 and STIM2 (Liou et al., 2005; Roos et al., 2005; Zhang et al., 2005) and the plasma membrane Ca 2+ -permeable channels Orai1, Orai2 and Orai3 (Feske et al., 2006; Vig et al., 2006; Zhang et al., 2006). STIM1 is a single-pass transmembrane ER protein bearing a luminal Ca 2+ -binding EF-hand domain (Liou et al., 2005; Stathopulos et al., 2008; Zhang et al., 2005) and a cytosolic channel activation domain (CAD) that mediates the trapping and gating of Orai channels (Kawasaki et al., 2009; Park et al., 2009; Yuan et al., 2009). "
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    ABSTRACT: STIM proteins populate and expand cortical ER sheets to mediate store-operated Ca(2+) entry (SOCE) by trapping and gating Orai channels in ER-PM clusters. A longer splice variant, STIM1L, forms permanent ER-PM clusters and mediates rapid influx in muscle. Here, we used electron microscopy, TIRF, and Ca(2+) imaging to establish the trafficking and signaling properties of the two STIM1 isoforms in Stim1(-/-)/Stim2(-/-) fibroblasts. Unlike STIM1, STIM1L was poorly recruited into ER-PM clusters and did not mediate store-dependent expansion of cortical ER cisternae. Removal of the STIM1 lysine-rich tail prevented store-dependent cluster enlargement, while inhibition of cytosolic Ca(2+) elevations or removal of the STIM1L actin binding domain had no impact on cluster expansion. Finally, STIM1L restored robust, but not accelerated SOCE and clustered with Orai1 channels more slowly than STIM1 following store depletion. These results indicate that STIM1L does not mediate rapid SOCE but can trap and gate Orai1 channels efficiently without remodeling cortical ER cisternae. The ability of STIM proteins to induce cortical ER formation is dispensable for SOCE and requires the lysine-rich tail of STIM1 involved in binding to phosphoinositides.
    Journal of Cell Science 03/2015; 128(8). DOI:10.1242/jcs.164228 · 5.33 Impact Factor
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    • "Two types of STIM-regulated SOCs have been described: the Orai channels [43], [44], [45] and transient receptor potential canonical (TRPC) channels [46], [47]. Upon store depletion, STIM proteins and SOCs translocate and cluster at the PM-ER junctions, leading to the formation of the STIM-Orai [44], [48] and STIM-TRPC [49] complexes and SOC activation. Activated SOCs mediateCa2+ influx to refill depleted intracellular stores and regulate cellular processes [50], [51]. "
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    ABSTRACT: The mechanisms of release, depletion, and refilling of endoplasmic reticulum (ER) Ca2+ were investigated in type I horizontal cells of the carp retina using a fluo-3-based Ca2+ imaging technique. Exogenous application of caffeine, a ryanodine receptor agonist, induced oscillatory intracellular free Ca2+ concentration ([Ca2+]i) responses in a duration- and concentration-dependent manner. In Ca2+-free Ringer's solution, [Ca2+]i transients could also be induced by a brief caffeine application, whereas subsequent caffeine application induced no [Ca2+]i increase, which implied that extracellular Ca2+ was required for ER refilling, confirming the necessity of a Ca2+ influx pathway for ER refilling. Depletion of ER Ca2+ by thapsigargin triggered a Ca2+ influx which could be blocked by the store-operated channel inhibitor 2-APB, which proved the existence of the store-operated Ca2+ entry pathway. Taken together, these results suggested that after being depleted by caffeine, the ER was replenished by Ca2+ influx via store-operated channels. These results reveal the fine modulation of ER Ca2+ signaling, and the activation of the store-operated Ca2+ entry pathway guarantees the replenishment of the ER so that the cell can be ready for response to the subsequent stimulus.
    PLoS ONE 06/2014; 9(6):e100095. DOI:10.1371/journal.pone.0100095 · 3.23 Impact Factor
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    • "Among the three IP 3 R paralogs (IP 3 R1–IP 3 R3), IP 3 R2 and IP 3 R3 are the major isoforms in secretory epithelial cells [16]. ER Ca 2+ release is frequently followed by activation of the store-operated channels (SOCs) in the plasma membrane, such as the Orai [17] [18] [19] and transient receptor potential cation (TRPC) channels [20] [21] [22]. In response to the depletion of Ca 2+ stores, the ER Ca 2+ sensor, stromal interaction molecule 1 (STIM1), clusters with these SOCs to activate them [23] [24]. "
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    ABSTRACT: Transepithelial bicarbonate secretion plays a key role in the maintenance of fluid and protein secretion from epithelial cells and the protection of the epithelial cell surface from various pathogens. Epithelial bicarbonate secretion is mainly under the control of cAMP and calcium signaling. While the physiological roles and molecular mechanisms of cAMP-induced bicarbonate secretion are relatively well defined, those induced by calcium signaling remain poorly understood in most epithelia. The present review summarizes the current status of knowledge on the role of calcium signaling in epithelial bicarbonate secretion. Specifically, this review introduces how cytosolic calcium signaling can increase bicarbonate secretion by regulating membrane transport proteins and how it synergizes with cAMP-induced mechanisms in epithelial cells. In addition, tissue-specific variations in the pancreas, salivary glands, intestines, bile ducts, and airways are discussed. We hope that the present report will stimulate further research into this important topic. These studies will provide the basis for future medicines for a wide spectrum of epithelial disorders including cystic fibrosis, Sjögren's syndrome, and chronic pancreatitis.
    Cell calcium 06/2014; 55(6). DOI:10.1016/j.ceca.2014.02.002 · 4.21 Impact Factor
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