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

Brij B Singh; Timothy P Lockwich; Bidhan C Bandyopadhyay; Xibao Liu; Sunitha Bollimuntha; So-Ching Brazer; Christian Combs; Sunit Das; A G Miriam Leenders; Zu-Hang Sheng; Mark A Knepper; Suresh V Ambudkar; Indu S Ambudkar

doi:10.1016/j.molcel.2004.07.010

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

Mol Cell. 2004 Aug 27;15(4):635-46. doi: 10.1016/j.molcel.2004.07.010.

Authors

Brij B Singh¹, Timothy P Lockwich, Bidhan C Bandyopadhyay, Xibao Liu, Sunitha Bollimuntha, So-Ching Brazer, Christian Combs, Sunit Das, A G Miriam Leenders, Zu-Hang Sheng, Mark A Knepper, Suresh V Ambudkar, Indu S Ambudkar

Affiliation

¹ Secretory Physiology Section, Gene Therapy and Therapeutics Branch, National Institute of Dental and Craniofacial Research, Bethesda, MD 20892, USA.

PMID: 15327778
DOI: 10.1016/j.molcel.2004.07.010

Abstract

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.

Publication types

Research Support, U.S. Gov't, P.H.S.

MeSH terms

Animals
Brefeldin A / metabolism
Calcium / metabolism*
Carbachol / metabolism
Carrier Proteins / metabolism
Cell Line
Cell Membrane / metabolism*
Cholinergic Agonists / metabolism
Cytoplasmic Vesicles / physiology
Exocytosis / physiology*
Fluorescence Recovery After Photobleaching
Hippocampus / cytology
Humans
Ion Channels / genetics
Ion Channels / metabolism*
Membrane Proteins / genetics
Membrane Proteins / metabolism*
Neurons / cytology
Neurons / metabolism
Protein Synthesis Inhibitors / metabolism
R-SNARE Proteins
Rats
Recombinant Fusion Proteins / genetics
Recombinant Fusion Proteins / metabolism
SNARE Proteins
Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins
TRPC Cation Channels
Tetanus Toxin / metabolism
Two-Hybrid System Techniques
Vesicular Transport Proteins / metabolism

Substances

Carrier Proteins
Cholinergic Agonists
Ion Channels
Membrane Proteins
Protein Synthesis Inhibitors
R-SNARE Proteins
Recombinant Fusion Proteins
SNARE Proteins
Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins
TRPC Cation Channels
TRPC3 cation channel
Tetanus Toxin
Vesicular Transport Proteins
Brefeldin A
Carbachol
Calcium

Abstract

Publication types

MeSH terms

Substances

Grants and funding