title: Molecular mechanisms underlying inhibition of STIM1-Orai1-mediated Ca2+entry induced by 2-aminoethoxydiphenyl borate.
abstract: Store-operated Ca2+entry (SOCE) mediated by STIM1 and Orai1 is crucial for Ca2+signaling and homeostasis in most cell types. 2-Aminoethoxydiphenyl borate (2-APB) is a well-described SOCE inhibitor, but its mechanisms of action remain largely elusive. Here, we show that 2-APB does not affect the dimeric state of STIM1, but enhances the intramolecular coupling between the coiled-coil 1 (CC1) and STIM-Orai-activating region (SOAR) of STIM1, with subsequent reduction in the formation of STIM1 puncta in the absence of Orai1 overexpression. 2-APB also inhibits Orai1 channels, directly inhibiting Ca2+entry through the constitutively active, STIM1-independent Orai1 mutants, Orai1-P245T and Orai1-V102A. When unbound from STIM1, the constitutively active Orai1-V102C mutant is not inhibited by 2-APB. Thus, we used Orai1-V012C as a tool to examine whether 2-APB can also inhibit the coupling between STIM1 and Orai1. We reveal that the functional coupling between STIM1 and Orai1-V102C is inhibited by 2-APB. This inhibition on coupling is indirect, arising from 2-APB's action on STIM1, and it is most likely mediated by functional channel residues in the Orai1 N-terminus. Overall, our findings on this two-site inhibition mediated by 2-APB provide new understanding on Orai1-activation by STIM1, important to future drug design.
authors.affiliation: Beijing Key Laboratory of Gene Resources and Molecular Development, College of Life Sciences, Beijing Normal University, Beijing, 100875, People's Republic of China.
authors.forename: Ming
authors.initials: M
authors.lastname: Wei
authors.affiliation: Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, PA, 17033, USA.
authors.forename: Yandong
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: Beijing Key Laboratory of Gene Resources and Molecular Development, College of Life Sciences, Beijing Normal University, Beijing, 100875, People's Republic of China.
authors.forename: Aomin
authors.initials: A
authors.lastname: Sun
authors.affiliation: Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, TX, 77030, USA.
authors.forename: Guolin
authors.initials: G
authors.lastname: Ma
authors.affiliation: Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, TX, 77030, USA.
authors.forename: Lian
authors.initials: L
authors.lastname: He
authors.affiliation: Beijing Key Laboratory of Gene Resources and Molecular Development, College of Life Sciences, Beijing Normal University, Beijing, 100875, People's Republic of China.
authors.forename: Lijuan
authors.initials: L
authors.lastname: Zhou
authors.affiliation: Beijing Key Laboratory of Gene Resources and Molecular Development, College of Life Sciences, Beijing Normal University, Beijing, 100875, People's Republic of China.
authors.forename: Shuce
authors.initials: S
authors.lastname: Zhang
authors.affiliation: Beijing Key Laboratory of Gene Resources and Molecular Development, College of Life Sciences, Beijing Normal University, Beijing, 100875, People's Republic of China.
authors.forename: Jin
authors.initials: J
authors.lastname: Liu
authors.affiliation: Department of Medical Physiology, College of Medicine, Texas A&M University Health Science Center, Temple, TX, 76504, USA.
authors.forename: Shenyuan L
authors.initials: SL
authors.lastname: Zhang
authors.affiliation: Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, PA, 17033, USA. dgill4@hmc.psu.edu.
authors.forename: Donald L
authors.initials: DL
authors.lastname: Gill
authors.affiliation: Beijing Key Laboratory of Gene Resources and Molecular Development, College of Life Sciences, Beijing Normal University, Beijing, 100875, People's Republic of China. wyoujun@bnu.edu.cn.
authors.forename: Youjun
authors.identifier: 0000-0003-0961-1716
authors.initials: Y
authors.lastname: Wang
mesh_terms: D001896:Boron Compounds / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects*; D057809:HEK293 Cells; D006801:Humans; D000071740:ORAI1 Protein / Q000378:metabolism*; D000071737:Stromal Interaction Molecule 1 / Q000378:metabolism*
chemical_list: D001896:Boron Compounds; D000071740:ORAI1 Protein; D000071737:Stromal Interaction Molecule 1; C109986:2-aminoethoxydiphenyl borate; D002118:Calcium
keywords: 2-APB; Calcium; FRET; Orai1; Puncta; SOCE; STIM1