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
Source: PubMed


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.

Download full-text


Available from: Ying Yu, Oct 06, 2015
33 Reads
  • Source
    • "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]. "
    [Show abstract] [Hide abstract]
    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.
    Cellular Signalling 08/2015; In press(11). DOI:10.1016/j.cellsig.2015.08.006 · 4.32 Impact Factor
  • Source
    • "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). "
    [Show abstract] [Hide abstract]
    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 · 5.43 Impact Factor
  • Source
    • "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]. "
    [Show abstract] [Hide abstract]
    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
Show more