Ca2+ store depletion causes STIM1 to accumulate in ER regions closely associated with the plasma membrane

doi:10.1083/jcb.200604014

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Published 11 September 2006. doi:10.1083/jcb.200604014
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JCB, Volume 174, Number 6, 803-813

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Ca²⁺ store depletion causes STIM1 to accumulate in ER regions closely associated with the plasma membrane

Minnie M. Wu, JoAnn Buchanan, Riina M. Luik, and Richard S. Lewis

Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305

Correspondence to Richard S. Lewis: rslewis{at}stanford.edu

Stromal interacting molecule 1 (STIM1), reported to be an endoplasmicreticulum (ER) Ca²⁺ sensor controlling store-operated Ca²⁺ entry,redistributes from a diffuse ER localization into puncta atthe cell periphery after store depletion. STIM1 redistributionis proposed to be necessary for Ca²⁺ release–activatedCa²⁺ (CRAC) channel activation, but it is unclear whether redistributionis rapid enough to play a causal role. Furthermore, the locationof STIM1 puncta is uncertain, with recent reports supportingretention in the ER as well as insertion into the plasma membrane(PM). Using total internal reflection fluorescence (TIRF) microscopyand patch-clamp recording from single Jurkat cells, we showthat STIM1 puncta form several seconds before CRAC channelsopen, supporting a causal role in channel activation. Fluorescencequenching and electron microscopy analysis reveal that punctacorrespond to STIM1 accumulation in discrete subregions of junctionalER located 10–25 nm from the PM, without detectable insertionof STIM1 into the PM. Roughly one third of these ER–PMcontacts form in response to store depletion. These studiesidentify an ER structure underlying store-operated Ca²⁺ entry,whose extreme proximity to the PM may enable STIM1 to interactwith CRAC channels or associated proteins.

Abbreviations used in this paper: CCD, charge-coupled device;CRAC, Ca²⁺ release–activated Ca²⁺; GPI, glycosylphosphatidylinositol;I_CRAC, CRAC current; IP₃, inositol 1,4,5-trisphosphate; SOC,store-operated channel; STIM1, stromal interacting molecule1; TCR, T cell receptor; TG, thapsigargin; TIRF, total internalreflection fluorescence.

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