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The action of selective CRAC channel blockers is affected by the Orai pore geometry.
Cell Calcium. 2013 Feb; 53(2):139-51.CC

Abstract

As the molecular composition of calcium-release activated calcium (CRAC) channels has been unknown for two decades, elucidation of selective inhibitors has been considerably hampered. By the identification of the two key components of CRAC channels, STIM1 and Orai1 have emerged as promising targets for CRAC blockers. The aim of this study was to thoroughly characterize the effects of two selective CRAC channel blockers on currents derived from STIM1/Orai heterologoulsy expressed in HEK293 cells. The novel compounds GSK-7975A and GSK-5503A were tested for effects on STIM1 mediated Orai1 or Orai3 currents by whole-cell patch-clamp recordings and for the effects on STIM1 oligomerisation or STIM1/Orai coupling by FRET microscopy. To investigate their site of action, inhibitory effects of these molecules were explored using Orai pore mutants. The GSK blockers inhibited Orai1 and Orai3 currents with an IC(50) of approximately 4μM and exhibited a substantially slower rate of onset than the typical pore blocker La(3+), together with almost no current recovery upon wash-out over 4min. For the less Ca(2+)-selective Orai1 E106D pore mutant, I(CRAC) inhibition was significantly reduced. FRET experiments indicated that neither STIM1-STIM1 oligomerization nor STIM1-Orai1 coupling was affected by these compounds. These CRAC channel blockers are acting downstream of STIM1 oligomerization and STIM1/Orai1 interaction, potentially via an allosteric effect on the selectivity filter of Orai. The elucidation of these CRAC current blockers represents a significant step toward the identification of CRAC channel-selective drug compounds.

Authors+Show Affiliations

Institute of Biophysics, University of Linz, 4040 Linz, Austria.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

23218667

Citation

Derler, Isabella, et al. "The Action of Selective CRAC Channel Blockers Is Affected By the Orai Pore Geometry." Cell Calcium, vol. 53, no. 2, 2013, pp. 139-51.
Derler I, Schindl R, Fritsch R, et al. The action of selective CRAC channel blockers is affected by the Orai pore geometry. Cell Calcium. 2013;53(2):139-51.
Derler, I., Schindl, R., Fritsch, R., Heftberger, P., Riedl, M. C., Begg, M., House, D., & Romanin, C. (2013). The action of selective CRAC channel blockers is affected by the Orai pore geometry. Cell Calcium, 53(2), 139-51. https://doi.org/10.1016/j.ceca.2012.11.005
Derler I, et al. The Action of Selective CRAC Channel Blockers Is Affected By the Orai Pore Geometry. Cell Calcium. 2013;53(2):139-51. PubMed PMID: 23218667.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - The action of selective CRAC channel blockers is affected by the Orai pore geometry. AU - Derler,Isabella, AU - Schindl,Rainer, AU - Fritsch,Reinhard, AU - Heftberger,Peter, AU - Riedl,Maria Christine, AU - Begg,Malcolm, AU - House,David, AU - Romanin,Christoph, Y1 - 2012/12/05/ PY - 2012/09/03/received PY - 2012/11/02/revised PY - 2012/11/03/accepted PY - 2012/12/11/entrez PY - 2012/12/12/pubmed PY - 2013/8/16/medline SP - 139 EP - 51 JF - Cell calcium JO - Cell Calcium VL - 53 IS - 2 N2 - As the molecular composition of calcium-release activated calcium (CRAC) channels has been unknown for two decades, elucidation of selective inhibitors has been considerably hampered. By the identification of the two key components of CRAC channels, STIM1 and Orai1 have emerged as promising targets for CRAC blockers. The aim of this study was to thoroughly characterize the effects of two selective CRAC channel blockers on currents derived from STIM1/Orai heterologoulsy expressed in HEK293 cells. The novel compounds GSK-7975A and GSK-5503A were tested for effects on STIM1 mediated Orai1 or Orai3 currents by whole-cell patch-clamp recordings and for the effects on STIM1 oligomerisation or STIM1/Orai coupling by FRET microscopy. To investigate their site of action, inhibitory effects of these molecules were explored using Orai pore mutants. The GSK blockers inhibited Orai1 and Orai3 currents with an IC(50) of approximately 4μM and exhibited a substantially slower rate of onset than the typical pore blocker La(3+), together with almost no current recovery upon wash-out over 4min. For the less Ca(2+)-selective Orai1 E106D pore mutant, I(CRAC) inhibition was significantly reduced. FRET experiments indicated that neither STIM1-STIM1 oligomerization nor STIM1-Orai1 coupling was affected by these compounds. These CRAC channel blockers are acting downstream of STIM1 oligomerization and STIM1/Orai1 interaction, potentially via an allosteric effect on the selectivity filter of Orai. The elucidation of these CRAC current blockers represents a significant step toward the identification of CRAC channel-selective drug compounds. SN - 1532-1991 UR - https://www.unboundmedicine.com/medline/citation/23218667/The_action_of_selective_CRAC_channel_blockers_is_affected_by_the_Orai_pore_geometry_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0143-4160(12)00183-2 DB - PRIME DP - Unbound Medicine ER -