title: Graded activation of CRAC channel by binding of different numbers of STIM1 to Orai1 subunits.
abstract: The Ca(2+) release-activated Ca(2+) (CRAC) channel pore is formed by Orai1 and gated by STIM1 after intracellular Ca(2+) store depletion. To resolve how many STIM1 molecules are required to open a CRAC channel, we fused different numbers of Orai1 subunits with functional two-tandem cytoplasmic domains of STIM1 (residues 336-485, designated as S domain). Whole-cell patch clamp recordings of these chimeric molecules revealed that CRAC current reached maximum at a stoichiometry of four Orai1 and eight S domains. Further experiments indicate that two-tandem S domains specifically interact with the C-terminus of one Orai1 subunit, and CRAC current can be gradually increased as more Orai1 subunits can interact with S domains or STIM1 proteins. Our data suggest that maximal opening of one CRAC channel requires eight STIM1 molecules, and support a model that the CRAC channel activation is not in an "all-or-none" fashion but undergoes a graded process via binding of different numbers of STIM1.
authors.affiliation: National Key Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Zhengzheng
authors.initials: Z
authors.lastname: Li
authors.forename: Lin
authors.initials: L
authors.lastname: Liu
authors.forename: Yongqiang
authors.initials: Y
authors.lastname: Deng
authors.forename: Wei
authors.initials: W
authors.lastname: Ji
authors.forename: Wen
authors.initials: W
authors.lastname: Du
authors.forename: Pingyong
authors.initials: P
authors.lastname: Xu
authors.forename: Liangyi
authors.initials: L
authors.lastname: Chen
authors.forename: Tao
authors.initials: T
authors.lastname: Xu
mesh_terms: D019943:Amino Acid Substitution; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D002460:Cell Line; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D009363:Neoplasm Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000071740:ORAI1 Protein; D018408:Patch-Clamp Techniques; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000071737:Stromal Interaction Molecule 1
chemical_list: D015220:Calcium Channels; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D000071740:ORAI1 Protein; C510746:ORAI1 protein, human; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; C103530:STIM1 protein, human; D000071737:Stromal Interaction Molecule 1; D002118:Calcium