title: Identification of two domains involved in the assembly of transient receptor potential canonical channels.
abstract: Transient receptor potential canonical (TRPC) channels are associated with calcium entry activity in nonexcitable cells. TRPCs can form homo- or heterotetrameric channels, in which case they can assemble together within a subfamily groups. TRPC1, 4, and 5 represent one group, and TRPC3, 6, and 7 represent the other. The molecular determinants involved in promoting subunit tetramerization are not known. To identify them, we generated chimeras by swapping the different domains of TRPC4 with the same regions in TRPC6. We showed that TRPC4 coimmunoprecipitated with the chimeras containing the ankyrin repeats and coiled-coil domains of TRPC4 into TRPC6. However, chimeras containing only the ankyrin repeats or only the coiled-coil domain of TRPC4 did not coimmunoprecipitate with TRPC4. We also showed that a second domain of interaction composed of the pore region and the C-terminal tail is involved in the oligomerization of TRPC4. However, chimeras containing only the pore region or only the C-terminal tail of TRPC4 did not coimmunoprecipitate with TRPC4. Furthermore, we showed that the N terminus of TRPC6 coimmunoprecipitated with the C terminus of TRPC6. Overexpression in HEK293T cells of chimeras that contained an N terminus and a C terminus from different subfamily groups increased intracellular calcium entry subsequent to stimulation of G(q) protein-coupled receptors. These results suggest that two types of interactions are involved in the assembly of the four subunits of the TRPC channel. The first interaction occurs between the N termini and involves two regions. The second interaction occurs between the N terminus and the C terminus and does not appear to be necessary for the activity of TRPCs.
authors.affiliation: Department of Pharmacology, Université de Sherbrooke, Sherbrooke, Quebec J1H 5N4, Canada.
authors.forename: Pascale K
authors.initials: PK
authors.lastname: Lepage
authors.forename: Marc P
authors.initials: MP
authors.lastname: Lussier
authors.forename: Hector
authors.initials: H
authors.lastname: Barajas-Martinez
authors.forename: Simon M
authors.initials: SM
authors.lastname: Bousquet
authors.forename: Alexandre P
authors.initials: AP
authors.lastname: Blanchard
authors.forename: Nancy
authors.initials: N
authors.lastname: Francoeur
authors.forename: Robert
authors.initials: R
authors.lastname: Dumaine
authors.forename: Guylain
authors.initials: G
authors.lastname: Boulay
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D003600:Cytosol / Q000378:metabolism; D004594:Electrophysiology; D019286:Endopeptidase K / Q000737:chemistry; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D051379:Mice; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D014162:Transfection; D050051:Transient Receptor Potential Channels / Q000737:chemistry* / Q000378:metabolism
chemical_list: D011993:Recombinant Fusion Proteins; D050051:Transient Receptor Potential Channels; D005982:Glutathione Transferase; D019286:Endopeptidase K; D002118:Calcium