Intramolecular shielding maintains the ER Ca2+ sensor stim1 in an inactive conformation

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Abstract

Store-operated calcium entry (SOCE) represents a major calcium influx pathway in non-excitable cells and is central to many physiological processes such as T cell activationand mast cell degranulation. SOCE is activated through intricate coordination between theCa2+ sensor on the ER membrane (stromal interaction molecule 1, STIM1) and the plasma membrane channel Orai1. When Ca2+ stores are depleted, STIM1 oligomerizes and physically interacts with Orai1 through its SOAR/CAD domain, resulting in Orai1 gating and Ca2+ influx. Here, we describe novel inter- and intramolecular FRET sensors in the context of the full-length membrane-anchored STIM1, and show that STIM1 undergoes a conformational change in response to store depletion to adopt a stretched 'open' conformation that exposes SOAR/CAD and allows it to interact with Orai1. Mutational analyses reveal that electrostatic interactions between the predicted first and third coiled-coil domains of STIM1 are not involved in maintaining the 'closed' inactive conformation. In addition, the results argue that an amphipathic a-helix between residues 317 and 336 in the so-called inhibitory domain is important to maintain STIM1 in a closed conformation at rest. Indeed, mutations that alter the amphipathic properties of this helix result in a STIM1 variant that is unable to respond to store depletion in terms of forming puncta, translocation to the cortical ER or activating Orai1.

Original languageEnglish
Pages (from-to)2401-2410
Number of pages10
JournalJournal of Cell Science
Volume126
Issue number11
DOIs
Publication statusPublished - 1 Jun 2013

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Calcium
Physiological Phenomena
Cell Degranulation
Membranes
Stromal Interaction Molecule 1
Static Electricity
Ion Channels
Mast Cells
Cell Membrane
T-Lymphocytes
Mutation

Keywords

  • Conformational change
  • Fret
  • Orai1
  • Stim1
  • Store-operated calcium entry

ASJC Scopus subject areas

  • Cell Biology

Cite this

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title = "Intramolecular shielding maintains the ER Ca2+ sensor stim1 in an inactive conformation",
abstract = "Store-operated calcium entry (SOCE) represents a major calcium influx pathway in non-excitable cells and is central to many physiological processes such as T cell activationand mast cell degranulation. SOCE is activated through intricate coordination between theCa2+ sensor on the ER membrane (stromal interaction molecule 1, STIM1) and the plasma membrane channel Orai1. When Ca2+ stores are depleted, STIM1 oligomerizes and physically interacts with Orai1 through its SOAR/CAD domain, resulting in Orai1 gating and Ca2+ influx. Here, we describe novel inter- and intramolecular FRET sensors in the context of the full-length membrane-anchored STIM1, and show that STIM1 undergoes a conformational change in response to store depletion to adopt a stretched 'open' conformation that exposes SOAR/CAD and allows it to interact with Orai1. Mutational analyses reveal that electrostatic interactions between the predicted first and third coiled-coil domains of STIM1 are not involved in maintaining the 'closed' inactive conformation. In addition, the results argue that an amphipathic a-helix between residues 317 and 336 in the so-called inhibitory domain is important to maintain STIM1 in a closed conformation at rest. Indeed, mutations that alter the amphipathic properties of this helix result in a STIM1 variant that is unable to respond to store depletion in terms of forming puncta, translocation to the cortical ER or activating Orai1.",
keywords = "Conformational change, Fret, Orai1, Stim1, Store-operated calcium entry",
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AU - Yu, Fang

AU - Sun, Lu

AU - Hubrack, Satanay Zuhair

AU - Selvaraj, Senthil

AU - Machaca, Khaled

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N2 - Store-operated calcium entry (SOCE) represents a major calcium influx pathway in non-excitable cells and is central to many physiological processes such as T cell activationand mast cell degranulation. SOCE is activated through intricate coordination between theCa2+ sensor on the ER membrane (stromal interaction molecule 1, STIM1) and the plasma membrane channel Orai1. When Ca2+ stores are depleted, STIM1 oligomerizes and physically interacts with Orai1 through its SOAR/CAD domain, resulting in Orai1 gating and Ca2+ influx. Here, we describe novel inter- and intramolecular FRET sensors in the context of the full-length membrane-anchored STIM1, and show that STIM1 undergoes a conformational change in response to store depletion to adopt a stretched 'open' conformation that exposes SOAR/CAD and allows it to interact with Orai1. Mutational analyses reveal that electrostatic interactions between the predicted first and third coiled-coil domains of STIM1 are not involved in maintaining the 'closed' inactive conformation. In addition, the results argue that an amphipathic a-helix between residues 317 and 336 in the so-called inhibitory domain is important to maintain STIM1 in a closed conformation at rest. Indeed, mutations that alter the amphipathic properties of this helix result in a STIM1 variant that is unable to respond to store depletion in terms of forming puncta, translocation to the cortical ER or activating Orai1.

AB - Store-operated calcium entry (SOCE) represents a major calcium influx pathway in non-excitable cells and is central to many physiological processes such as T cell activationand mast cell degranulation. SOCE is activated through intricate coordination between theCa2+ sensor on the ER membrane (stromal interaction molecule 1, STIM1) and the plasma membrane channel Orai1. When Ca2+ stores are depleted, STIM1 oligomerizes and physically interacts with Orai1 through its SOAR/CAD domain, resulting in Orai1 gating and Ca2+ influx. Here, we describe novel inter- and intramolecular FRET sensors in the context of the full-length membrane-anchored STIM1, and show that STIM1 undergoes a conformational change in response to store depletion to adopt a stretched 'open' conformation that exposes SOAR/CAD and allows it to interact with Orai1. Mutational analyses reveal that electrostatic interactions between the predicted first and third coiled-coil domains of STIM1 are not involved in maintaining the 'closed' inactive conformation. In addition, the results argue that an amphipathic a-helix between residues 317 and 336 in the so-called inhibitory domain is important to maintain STIM1 in a closed conformation at rest. Indeed, mutations that alter the amphipathic properties of this helix result in a STIM1 variant that is unable to respond to store depletion in terms of forming puncta, translocation to the cortical ER or activating Orai1.

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