title: MxA, a member of the dynamin superfamily, interacts with the ankyrin-like repeat domain of TRPC.
abstract: Mammalian transient receptor potential canonical channels have been proposed as the molecular entities associated with calcium entry activity in nonexcitable cells. Amino acid sequence analyses of TRPCs revealed the presence of ankyrin-like repeat domains, one of the most common protein-protein interaction motifs. Using a yeast two-hybrid interaction assay, we found that the second ankyrin-like repeat domain of TRPC6 interacted with MxA, a member of the dynamin superfamily. Using a GST pull-down and co-immunoprecipitation assay, we showed that MxA interacted with TRPC1, -3, -4, -5, -6, and -7. Overexpression of MxA in HEK293T cells slightly increased endogenous calcium entry subsequent to stimulation of G(q) protein-coupled receptors or store depletion by thapsigargin. Co-expression of MxA with TRPC6 enhanced agonist-induced or OAG-induced calcium entry activity. GTP binding-defective MxA mutants had only a minor potentiating effect on OAG-induced TRPC6 activity. However, a MxA mutant that could bind GTP but that lacked GTPase activity produced the same effect as MxA on OAG-induced TRPC6 activity. These results indicated that MxA interacted specifically with the second ankyrin-like repeat domain of TRPCs and suggested that monomeric MxA regulated the activity of TRPC6 by a mechanism requiring GTP binding. Additional results showed that an increase in the endogenous expression of MxA, induced by a treatment with interferon alpha, regulated the activity of TRPC6. The study clearly identified MxA as a new regulatory protein involved in Ca2+ signaling.
authors.affiliation: Department of Pharmacology, Université de Sherbrooke, Sherbrooke, Quebec J1H 5N4, Canada.
authors.forename: Marc P
authors.initials: MP
authors.lastname: Lussier
authors.forename: Sylvie
authors.initials: S
authors.lastname: Cayouette
authors.forename: Pascale K
authors.initials: PK
authors.lastname: Lepage
authors.forename: Cynthia L
authors.initials: CL
authors.lastname: Bernier
authors.forename: Nancy
authors.initials: N
authors.lastname: Francoeur
authors.forename: Marie
authors.initials: M
authors.lastname: St-Hilaire
authors.forename: Maxime
authors.initials: M
authors.lastname: Pinard
authors.forename: Guylain
authors.initials: G
authors.lastname: Boulay
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D017487:Ankyrins / Q000737:chemistry*; D002118:Calcium / Q000737:chemistry / Q000378:metabolism; D015220:Calcium Channels / Q000737:chemistry / Q000378:metabolism*; D027682:Cation Transport Proteins / Q000737:chemistry; D002460:Cell Line; D018076:DNA, Complementary / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism / Q000502:physiology*; D005982:Glutathione Transferase / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000737:chemistry; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D016898:Interferon-alpha / Q000378:metabolism; D007473:Ion Channels / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry; D008969:Molecular Sequence Data; D009154:Mutation; D064547:Myxovirus Resistance Proteins; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D013050:Spectrometry, Fluorescence; D050052:TRPC Cation Channels; D000074287:TRPC6 Cation Channel; D050053:TRPM Cation Channels; D019284:Thapsigargin / Q000494:pharmacology; D013997:Time Factors; D014162:Transfection; D020798:Two-Hybrid System Techniques
chemical_list: D017487:Ankyrins; D015220:Calcium Channels; D027682:Cation Transport Proteins; D018076:DNA, Complementary; D016898:Interferon-alpha; D007473:Ion Channels; C579071:MX1 protein, human; D008565:Membrane Proteins; D064547:Myxovirus Resistance Proteins; D050052:TRPC Cation Channels; C104603:TRPC3 cation channel; C489571:TRPC4 ion channel; C491780:TRPC5 protein, human; D000074287:TRPC6 Cation Channel; C117032:TRPC6 protein, human; D050053:TRPM Cation Channels; C121934:TRPM2 protein, human; C096351:transient receptor potential cation channel, subfamily C, member 1; D019284:Thapsigargin; D006160:Guanosine Triphosphate; D005982:Glutathione Transferase; D019204:GTP-Binding Proteins; D002118:Calcium