Mol Cell. 2004 Aug 27;15(4):635-46.

VAMP2-dependent exocytosis regulates plasma membrane insertion of TRPC3 channels and contributes to agonist-stimulated Ca2+ influx.External 2231691f894ba696de1310221b0a0dbbb31a7251e75115c265587c3d9d5f507c

Singh, B. B., Lockwich, T. P., Bandyopadhyay, B. C., Liu, X., Bollimuntha, S., Brazer, S. C., Combs, C., Das, S., Leenders, A. G., Sheng, Z. H., Knepper, M. A., Ambudkar, S. V., Ambudkar, I. S.,
["Secretory Physiology Section, Gene Therapy and Therapeutics Branch, National Institute of Dental and Craniofacial Research, Bethesda, MD 20892, USA."]
The mechanism(s) involved in agonist-stimulation of TRPC3 channels is not yet known. Here we demonstrate that TRPC3-N terminus interacts with VAMP2 and alphaSNAP. Further, endogenous and exogenously expressed TRPC3 colocalized and coimmunoprecipitated with SNARE proteins in neuronal and epithelial cells. Imaging of GFP-TRPC3 revealed its localization in the plasma membrane region and in mobile intracellular vesicles. Recovery of TRPC3-GFP fluorescence after photobleaching of the plasma membrane region was decreased by brefeldin-A or BAPTA-AM. Cleavage of VAMP2 with tetanus toxin (TeNT) did not prevent delivery of TRPC3 to the plasma membrane region but reduced its surface expression. TeNT also decreased carbachol and OAG, but not thapsigargin, stimulated Ca2+ influx. Importantly, carbachol, not thapsigargin, increased surface expression of TRPC3 that was attenuated by TeNT and not by BAPTA. In aggregate, these data suggest that VAMP2-dependent exocytosis regulates plasma membrane insertion of TRPC3 channels and contributes to carbachol-stimulation of Ca2+ influx.
PMID: 15327778External 2231691f894ba696de1310221b0a0dbbb31a7251e75115c265587c3d9d5f507c
Screening Toggle 893349bafcc528f8346c51dc3420151d67b0126b2c122dd1017121c03fa0f69b
  Experimental screening Non-experimental screening Reference
TRP channel construct Interactor source
TRP channel Interactor Method Species Region Species Organ/tissue Sample type
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 ơSNAP Yeast two-hybrid Human 1-341 Rat Brain cDNA library 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 SNAP23 Inference Prediction 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 Syntaxin3 Inference Prediction 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 VAMP2 Yeast two-hybrid Human 1-341 Rat Brain cDNA library 15327778
(Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4: click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
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
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 ơSNAP Co-immunoprecipitation HEK293 Human Full-length Not used 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 ơSNAP Co-immunofluorescence staining Rat hippocampal neuron 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 ơSNAP Co-immunofluorescence staining HSG cell 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 ơSNAP Co-immunofluorescence staining HEK293 Human Full-length Not used 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 SNAP23 Co-immunoprecipitation HEK293 Human Full-length Not used 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 SNAP23 Co-immunofluorescence staining Rat hippocampal neuron 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 SNAP23 Co-immunofluorescence staining HEK293 Human Full-length Not used 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 Syntaxin3 Co-immunoprecipitation Rat hippocampal neuron 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 Syntaxin3 Co-immunoprecipitation HEK293 Human Full-length Not used 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 Syntaxin3 Co-immunofluorescence staining Rat hippocampal neuron 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 Syntaxin3 Co-immunofluorescence staining HEK293 Human Full-length Not used 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 VAMP2 Co-immunoprecipitation Rat hippocampal neuron 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 VAMP2 Co-immunoprecipitation HEK293 Human Full-length Not used 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 VAMP2 Co-immunofluorescence staining Rat hippocampal neuron 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 VAMP2 Co-immunofluorescence staining HSG cell 15327778
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 VAMP2 Co-immunofluorescence staining HEK293 Human Full-length Not used 15327778
(Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4: click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
Characterization Toggle 893349bafcc528f8346c51dc3420151d67b0126b2c122dd1017121c03fa0f69b
  Binding region mapping Stoichiometry Affinity (Kd) Reference
TRP channel Interactor
TRP channel Interactor Method Species Region Species Region
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 VAMP2 Yeast two-hybrid Human 1-123 Rat Not determined 15327778
(Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4: click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
TRP / Interactor

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