title: Potent functional uncoupling between STIM1 and Orai1 by dimeric 2-aminodiphenyl borinate analogs.
abstract: The coupling of ER Ca(2+)-sensing STIM proteins and PM Orai Ca(2+) entry channels generates "store-operated" Ca(2+) signals crucial in controlling responses in many cell types. The dimeric derivative of 2-aminoethoxydiphenyl borinate (2-APB), DPB162-AE, blocks functional coupling between STIM1 and Orai1 with an IC50 (200 nM) 100-fold lower than 2-APB. Unlike 2-APB, DPB162-AE does not affect L-type or TRPC channels or Ca(2+) pumps at maximal STIM1-Orai1 blocking levels. DPB162-AE blocks STIM1-induced Orai1 or Orai2, but does not block Orai3 or STIM2-mediated effects. We narrowed the DPB162-AE site of action to the STIM-Orai activating region (SOAR) of STIM1. DPB162-AE does not prevent the SOAR-Orai1 interaction but potently blocks SOAR-mediated Orai1 channel activation, yet its action is not as an Orai1 channel pore blocker. Using the SOAR-F394H mutant which prevents both physical and functional coupling to Orai1, we reveal DPB162-AE rapidly restores SOAR-Orai binding but only slowly restores Orai1 channel-mediated Ca(2+) entry. With the same SOAR mutant, 2-APB induces rapid physical and functional coupling to Orai1, but channel activation is transient. We infer that the actions of both 2-APB and DPB162-AE are directed toward the STIM1-Orai1 coupling interface. Compared to 2-APB, DPB162-AE is a much more potent and specific STIM1/Orai1 functional uncoupler. DPB162-AE provides an important pharmacological tool and a useful mechanistic probe for the function and coupling between STIM1 and Orai1 channels.
authors.affiliation: Department of Biochemistry, Temple University School of Medicine, Philadelphia, PA 19140, United States.
authors.forename: Eunan
authors.initials: E
authors.lastname: Hendron
authors.affiliation: Department of Biochemistry, Temple University School of Medicine, Philadelphia, PA 19140, United States; Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, PA 17033, United States.
authors.forename: Xizhuo
authors.initials: X
authors.lastname: Wang
authors.affiliation: Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, PA 17033, United States.
authors.forename: Yandong
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, PA 17033, United States.
authors.forename: Xiangyu
authors.initials: X
authors.lastname: Cai
authors.affiliation: Department of Physiology, Yamagata University School of Medicine, Yamagata 990-9585, Japan.
authors.forename: Jun-ichi
authors.initials: J
authors.lastname: Goto
authors.affiliation: Laboratory for Developmental Neurobiology, RIKEN Brain Science Institute, Saitama 351-0198, Japan.
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Mikoshiba
authors.affiliation: Laboratory for Lymphocyte Differentiation, WPI Immunology Frontier Research Center, Osaka University, Suita, Osaka 565-0871, Japan; Laboratory for Lymphocyte Differentiation, RIKEN Center for Integrative Medical Sciences, Yokohama, Kanagawa 230-0045, Japan.
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Baba
authors.affiliation: Laboratory for Lymphocyte Differentiation, WPI Immunology Frontier Research Center, Osaka University, Suita, Osaka 565-0871, Japan; Laboratory for Lymphocyte Differentiation, RIKEN Center for Integrative Medical Sciences, Yokohama, Kanagawa 230-0045, Japan.
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Kurosaki
authors.affiliation: Beijing Key Laboratory of Gene Resources and Molecular Development College of Life Sciences, Beijing Normal University, Beijing 100875, PR China. Electronic address: wyoujun@bnu.edu.cn.
authors.forename: Youjun
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, PA 17033, United States. Electronic address: dongill@psu.edu.
authors.forename: Donald L
authors.initials: DL
authors.lastname: Gill
mesh_terms: D000818:Animals; D001896:Boron Compounds / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000187:drug effects* / Q000378:metabolism; D002460:Cell Line; D031541:Fluorescence Resonance Energy Transfer; D057809:HEK293 Cells; D006801:Humans; D019169:Jurkat Cells; D015471:Leukemia, Basophilic, Acute; D008562:Membrane Glycoproteins / Q000187:drug effects*; D008565:Membrane Proteins / Q000187:drug effects*; D009363:Neoplasm Proteins / Q000187:drug effects*; D000071740:ORAI1 Protein; D051381:Rats; D000071737:Stromal Interaction Molecule 1; D014475:Uncoupling Agents / Q000494:pharmacology*
chemical_list: D001896:Boron Compounds; D015220:Calcium Channels; C000599936:DPB162-AE; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D000071740:ORAI1 Protein; C510746:ORAI1 protein, human; C532493:Orai1 protein, rat; C103530:STIM1 protein, human; C518857:Stim1 protein, rat; D000071737:Stromal Interaction Molecule 1; D014475:Uncoupling Agents; C109986:2-aminoethoxydiphenyl borate; D002118:Calcium
keywords: 2-APB; Calcium; Orai1; Orai2; Orai3; STIM1; STIM2