title: STIM1 and calmodulin interact with Orai1 to induce Ca2+-dependent inactivation of CRAC channels.
abstract: Ca(2+)-dependent inactivation (CDI) is a key regulator and hallmark of the Ca(2+) release-activated Ca(2+) (CRAC) channel, a prototypic store-operated Ca(2+) channel. Although the roles of the endoplasmic reticulum Ca(2+) sensor STIM1 and the channel subunit Orai1 in CRAC channel activation are becoming well understood, the molecular basis of CDI remains unclear. Recently, we defined a minimal CRAC activation domain (CAD; residues 342-448) that binds directly to Orai1 to activate the channel. Surprisingly, CAD-induced CRAC currents lack fast inactivation, revealing a critical role for STIM1 in this gating process. Through truncations of full-length STIM1, we identified a short domain (residues 470-491) C-terminal to CAD that is required for CDI. This domain contains a cluster of 7 acidic amino acids between residues 475 and 483. Neutralization of aspartate or glutamate pairs in this region either reduced or enhanced CDI, whereas the combined neutralization of six acidic residues eliminated inactivation entirely. Based on bioinformatics predictions of a calmodulin (CaM) binding site on Orai1, we also investigated a role for CaM in CDI. We identified a membrane-proximal N-terminal domain of Orai1 (residues 68-91) that binds CaM in a Ca(2+)-dependent manner and mutations that eliminate CaM binding abrogate CDI. These studies identify novel structural elements of STIM1 and Orai1 that are required for CDI and support a model in which CaM acts in concert with STIM1 and the N terminus of Orai1 to evoke rapid CRAC channel inactivation.
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, CA 94305, USA.
authors.forename: Franklin M
authors.initials: FM
authors.lastname: Mullins
authors.forename: Chan Young
authors.initials: CY
authors.lastname: Park
authors.forename: Ricardo E
authors.initials: RE
authors.lastname: Dolmetsch
authors.forename: Richard S
authors.initials: RS
authors.lastname: Lewis
mesh_terms: D015220:Calcium Channels / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002460:Cell Line; D019295:Computational Biology; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004594:Electrophysiology; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D015640:Ion Channel Gating / Q000502:physiology*; D008565:Membrane Proteins / Q000378:metabolism*; D008954:Models, Biological; D016296:Mutagenesis; D009363:Neoplasm Proteins / Q000378:metabolism*; D000071740:ORAI1 Protein; D010957:Plasmids / Q000235:genetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary / Q000235:genetics; D000071737:Stromal Interaction Molecule 1; D014162:Transfection
chemical_list: D015220:Calcium Channels; D002147:Calmodulin; D017931:DNA Primers; D018076:DNA, Complementary; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D000071740:ORAI1 Protein; C510746:ORAI1 protein, human; C103530:STIM1 protein, human; D000071737:Stromal Interaction Molecule 1
references.citation: J Physiol. 2009 Jun 15;587(Pt 12):2903-18
references.pmid: 19403622
references.citation: Nature. 2006 Sep 14;443(7108):226-9
references.pmid: 16921385
references.citation: Channels (Austin). 2008 Jul-Aug;2(4):261-8
references.pmid: 18769136
references.citation: Neuron. 1999 Apr;22(4):645-8
references.pmid: 10230783
references.citation: Neuron. 2001 Sep 27;31(6):973-85
references.pmid: 11580897
references.citation: Biochim Biophys Acta. 1999 Aug 17;1433(1-2):56-67
references.pmid: 10446359
references.citation: J Cell Biol. 2006 Sep 11;174(6):815-25
references.pmid: 16966423
references.citation: Nature. 2008 Jul 24;454(7203):538-42
references.pmid: 18596693
references.citation: Annu Rev Physiol. 2002;64:289-311
references.pmid: 11826271
references.citation: Curr Biol. 2005 Jul 12;15(13):1235-41
references.pmid: 16005298
references.citation: J Physiol. 1993 Jun;465:359-86
references.pmid: 8229840
references.citation: Trends Pharmacol Sci. 2009 Mar;30(3):118-28
references.pmid: 19187978
references.citation: Cell. 2008 Oct 3;135(1):110-22
references.pmid: 18854159
references.citation: Physiol Rev. 2005 Apr;85(2):757-810
references.pmid: 15788710
references.citation: Curr Biol. 2006 Oct 24;16(20):2073-9
references.pmid: 16978865
references.citation: J Cell Biol. 2006 Sep 11;174(6):803-13
references.pmid: 16966422
references.citation: J Gen Physiol. 2003 Aug;122(2):207-23
references.pmid: 12860924
references.citation: Biochemistry. 2008 Jun 10;47(23):6089-91
references.pmid: 18484746
references.citation: J Biol Chem. 2001 Aug 17;276(33):30794-802
references.pmid: 11408490
references.citation: J Gen Physiol. 2004 Feb;123(2):167-82
references.pmid: 14744989
references.citation: Trends Biochem Sci. 1990 Feb;15(2):59-64
references.pmid: 2186516
references.citation: J Gen Physiol. 2007 Nov;130(5):525-40
references.pmid: 17968026
references.citation: J Gen Physiol. 1995 Feb;105(2):209-26
references.pmid: 7760017
references.citation: Nat Immunol. 2008 Apr;9(4):432-43
references.pmid: 18327260
references.citation: Annu Rev Immunol. 2001;19:497-521
references.pmid: 11244045
references.citation: Cell. 2009 Mar 6;136(5):876-90
references.pmid: 19249086
references.citation: J Physiol. 2004 Jul 1;558(Pt 1):85-97
references.pmid: 15226409
references.citation: J Biol Chem. 2009 Mar 27;284(13):8421-6
references.pmid: 19189966
references.citation: Nat Cell Biol. 2009 Mar;11(3):337-43
references.pmid: 19182790
references.citation: Nature. 2005 Oct 6;437(7060):902-5
references.pmid: 16208375
references.citation: J Struct Funct Genomics. 2000;1(1):8-14
references.pmid: 12836676
references.citation: J Physiol. 2004 Jan 15;554(Pt 2):255-61
references.pmid: 14500775
references.citation: Cell Calcium. 2007 Aug;42(2):163-72
references.pmid: 17499354
references.citation: Proc Natl Acad Sci U S A. 2007 May 29;104(22):9301-6
references.pmid: 17517596
references.citation: Nature. 2006 Sep 14;443(7108):230-3
references.pmid: 16921383