Mol Pharmacol. 2012 Apr;81(4):510-26. doi: 10.1124/mol.111.074658. Epub 2011 Dec 30.

The Ca(2+) sensor stromal interaction molecule 1 (STIM1) is necessary and sufficient for the store-operated Ca(2+) entry function of transient receptor potential canonical (TRPC) 1 and 4 channels in endothelial cells.External 2231691f894ba696de1310221b0a0dbbb31a7251e75115c265587c3d9d5f507c

Sundivakkam, P. C., Freichel, M., Singh, V., Yuan, J. P., Vogel, S. M., Flockerzi, V., Malik, A. B., Tiruppathi, C.,
["Department of Pharmacology and Center for Lung and Vascular Biology, College of Medicine, University of Illinois, Chicago, Illinois 60612, USA."]
We addressed the requirement for stromal interaction molecule 1 (STIM1), the endoplasmic reticulum (ER) Ca(2+)-sensor, and Orai1, a Ca(2+) selective channel, in regulating Ca(2+) entry through the store-operated channels mouse transient receptor potential canonical (TRPC) 4 or human TRPC1. Studies were made using murine and human lung endothelial cells (ECs) challenged with thrombin known to induce Ca(2+) entry via TRPC1/4. Deletion or knockdown of TRPC4 abolished Ca(2+) entry secondary to depletion of ER Ca(2+) stores, preventing the disruption of the endothelial barrier. Knockdown of STIM1 (but not of Orai1or Orai3) or expression of the dominant-negative STIM1(K684E-K685E) mutant in ECs also suppressed Ca(2+) entry secondary to store depletion. Ectopic expression of WT-STIM1 or WT-Orai1 in TRPC4(-/-)-ECs failed to rescue Ca(2+) entry; however, WT-TRPC4 expression in TRPC4(-/-)-ECs restored Ca(2+) entry indicating the requirement for TRPC4 in mediating store-operated Ca(2+) entry. Moreover, expression of the dominant-negative Orai1(R91W) mutant or Orai3(E81W) mutant in WT-ECs failed to prevent thrombin-induced Ca(2+) entry. In contrast, expression of the dominant-negative TRPC4(EE647-648KK) mutant in WT-ECs markedly reduced thrombin-induced Ca(2+) entry. In ECs expressing YFP-STIM1, ER-store Ca(2+) depletion induced formation of fluorescent membrane puncta in WT but not in TRPC4(-/-) cells, indicating that mobilization of STIM1 and engagement of its Ca(2+) sensing function required TRPC4 expression. Coimmunoprecipitation studies showed coupling of TRPC1 and TRPC4 with STIM1 on depletion of ER Ca(2+) stores. Thus, TRPC1 and TRPC4 can interact with STIM1 to form functional store-operated Ca(2+)-entry channels, which are essential for mediating Ca(2+) entry-dependent disruption of the endothelial barrier.
PMID: 22210847External 2231691f894ba696de1310221b0a0dbbb31a7251e75115c265587c3d9d5f507c
Validation: In vivo validation Toggle 893349bafcc528f8346c51dc3420151d67b0126b2c122dd1017121c03fa0f69b
  Assay with endogenous proteins Assay with overexpressed proteins Reference
Cell or tissue Cell or tissue TRP channel construct Interactor construct
TRP channel Interactor Method Species Region Species Region
TRPC1 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 TRPC4 Co-immunoprecipitation Mouse lung endothelial cell Mouse Full-length Mouse Full-length 22210847
TRPC4 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 STIM1 Co-immunoprecipitation Mouse lung endothelial cell Mouse Full-length Human Full-length 22210847
TRPC4 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 TRPC1 Co-immunoprecipitation Mouse lung endothelial cell Mouse Full-length Mouse Full-length 22210847
(Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4: click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
TRP / Interactor

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