title: Complex actions of 2-aminoethyldiphenyl borate on store-operated calcium entry.
abstract: Store-operated Ca2+ entry (SOCE) is likely the most common mode of regulated influx of Ca2+ into cells. However, only a limited number of pharmacological agents have been shown to modulate this process. 2-Aminoethyldiphenyl borate (2-APB) is a widely used experimental tool that activates and then inhibits SOCE and the underlying calcium release-activated Ca2+ current (I CRAC). The mechanism by which depleted stores activates SOCE involves complex cellular movements of an endoplasmic reticulum Ca2+ sensor, STIM1, which redistributes to puncta near the plasma membrane and, in some manner, activates plasma membrane channels comprising Orai1, -2, and -3 subunits. We show here that 2-APB blocks puncta formation of fluorescently tagged STIM1 in HEK293 cells. Accordingly, 2-APB also inhibited SOCE and I(CRAC)-like currents in cells co-expressing STIM1 with the CRAC channel subunit, Orai1, with similar potency. However, 2-APB inhibited STIM1 puncta formation less well in cells co-expressing Orai1, indicating that the inhibitory effects of 2-APB are not solely dependent upon STIM1 reversal. Further, 2-APB only partially inhibited SOCE and current in cells co-expressing STIM1 and Orai2 and activated sustained currents in HEK293 cells expressing Orai3 and STIM1. Interestingly, the Orai3-dependent currents activated by 2-APB showed large outward currents at potentials greater than +50 mV. Finally, Orai3, and to a lesser extent Orai1, could be directly activated by 2-APB, independently of internal Ca2+ stores and STIM1. These data reveal novel and complex actions of 2-APB effects on SOCE that can be attributed to effects on both STIM1 as well as Orai channel subunits.
authors.affiliation: Laboratory of Signal Transduction, NIEHS, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, North Carolina 27709, USA.
authors.forename: Wayne I
authors.initials: WI
authors.lastname: DeHaven
authors.forename: Jeremy T
authors.initials: JT
authors.lastname: Smyth
authors.forename: Rebecca R
authors.initials: RR
authors.lastname: Boyles
authors.forename: Gary S
authors.initials: GS
authors.lastname: Bird
authors.forename: James W
authors.initials: JW
authors.lastname: Putney
mesh_terms: D001896:Boron Compounds / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism*; D002120:Calcium Channel Agonists / Q000494:pharmacology*; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D006801:Humans; D008564:Membrane Potentials / Q000187:drug effects; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D000071740:ORAI1 Protein; D000071741:ORAI2 Protein; D000071737:Stromal Interaction Molecule 1
chemical_list: D001896:Boron Compounds; D002120:Calcium Channel Agonists; D015220:Calcium Channels; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D000071740:ORAI1 Protein; C510746:ORAI1 protein, human; D000071741:ORAI2 Protein; C514506:ORAI2 protein, human; C522867:Orai3 protein, human; C103530:STIM1 protein, human; D000071737:Stromal Interaction Molecule 1; C109986:2-aminoethoxydiphenyl borate; D002118:Calcium