Zhang SL, Yeromin AV, Zhang XH et al.Genome-wide RNAi screen of Ca(2+) influx identifies genes that regulate Ca(2+) release-activated Ca(2+) 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: 9.67). 07/2006; 103(24):9357-62. DOI: 10.1073/pnas.0603161103
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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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    • "channels (Nunes et al., 2012). STIM1 mediates the ubiquitous SOCE mechanism (Liou et al., 2 0 2005; Roos et al., 2005; Zhang et al., 2005) by acting as an intracellular ligand for plasma 2 1 membrane Ca 2+ -permeable channels of the Orai (Feske et al., 2006; Vig et al., 2006; Zhang et 2 2 al., 2006) and transient receptor potential canonical (TRPC) families (Huang et al., 2006), 2 3 reviewed in (Hogan et al., 2010), together referred to as store-operated Ca 2+ (SOCE) 2 4 channels. Upon ER Ca 2+ depletion, STIM1 oligomerizes and accumulates in cortical contact 2 5 sites between ER and plasma membranes (Luik et al., 2008; Lur et al., 2009; Orci et al., 2 6 2009; Wu et al., 2006) where its channel activating domain (CAD) directly interacts with 2 7 Orai1 to promote channel opening (Luik et al., 2006; Park et al., 2009; Xu et al., 2006; Zhou 2 8 et al., 2010). "
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    ABSTRACT: Local intracellular Ca(2+) elevations increase the efficiency of phagocytosis, a process essential for innate and adaptive immunity. These local Ca(2+) elevations are generated in part by the store-operated Ca(2+) entry (SOCE) sensor STIM1, which recruits ER cisternae to phagosomes and opens phagosomal Ca(2+) channels at ER-phagosome junctions. However, residual ER-phagosome contacts and periphagosomal Ca(2+) hotspots remain in Stim1(-/-) cells. Here, we tested whether junctate, a molecule that targets STIM1 to ER-plasma membrane contacts upon Ca(2+)-store depletion, could cooperate with STIM1 at phagosome junctions. Junctate expression in Stim1(-/-) and Stim1(-/-)/; Stim2(-/-) phagocytic fibroblasts increased phagocytosis and periphagosomal Ca(2+) elevations, yet with only a minimal impact on global SOCE. These Ca(2+) hotspots were only marginally reduced by the SOCE channel blocker La(3+) but were abrogated by InsP3R inhibitors 2-APB and xestospongin-C, revealing that unlike STIM1-mediated hotspots, junctate-mediated Ca(2+) ions originate predominantly from periphagosomal Ca(2+) stores. Accordingly, junctate accumulated near phagosomes and elongated ER-phagosome junctions in Stim1(-/-) cells. Thus, junctate mediates an alternative mechanism for generating localized Ca(2+) elevations within cells, promoting Ca(2+) release from internal stores recruited to phagosomes, thereby boosting phagocytosis.
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    • "During low Ca 2+ concentrations, STIM1 proteins sense the depletion of Ca 2+ stores and spatially redistribute to the ER to activate Orai proteins [31] [32] [33] [34] [35] [36], which perforate the PM and establish Ca 2+ release-activated channels (CRAC) [31] [37] (Fig. 1). The connection between ER and PM through CRAC will produce intracellular calcium transient ion flow [31,32,38–40]. "
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    ABSTRACT: Cell proliferation is orchestrated through diverse proteins related to calcium (Ca(2+)) signaling inside the cell. Cellular Ca(2+) influx occurs first by various mechanisms at the plasma membrane, is then followed by absorption of Ca(2+) ions by mitochondria and endoplasmic reticulum, and, finally, there is a connection of calcium stores to the nucleus. Experimental evidence indicates that the fluctuation of Ca(2+) from the endoplasmic reticulum provides a pivotal and physiological role for cell proliferation. Ca(2+) depletion in the endoplasmatic reticulum triggers Ca(2+) influx across the plasma membrane in an phenomenon called store-operated calcium entries (SOCE). SOCE is activated through a complex interplay between a Ca(2+) sensor, denominated STIM, localized in the endoplasmic reticulum and a Ca(2+) channel at the cell membrane, denominated Orai. The interplay between STIM and Orai proteins with cell membrane receptors and their role in cell proliferation are discussed in this review. Copyright © 2015. Published by Elsevier Inc.
    Full-text · Article · Aug 2015 · Cellular Signalling
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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.
    Full-text · Article · Mar 2015 · Journal of Cell Science
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