J Biol Chem. 2005 May 13;280(19):19393-400. Epub 2005 Mar 9.

MxA, a member of the dynamin superfamily, interacts with the ankyrin-like repeat domain of TRPC.External 2231691f894ba696de1310221b0a0dbbb31a7251e75115c265587c3d9d5f507c

Lussier, M. P., Cayouette, S., Lepage, P. K., Bernier, C. L., Francoeur, N., St-Hilaire, M., Pinard, M., Boulay, G.,
["Department of Pharmacology, Universite de Sherbrooke, Sherbrooke, Quebec J1H 5N4, Canada."]
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.
PMID: 15757897External 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
TRPC1 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Inference Prediction 15757897
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Inference Prediction 15757897
TRPC4 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Inference Prediction 15757897
TRPC5 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Inference Prediction 15757897
TRPC6 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Yeast two-hybrid Human 3-259 Human Fetal brain cDNA library 15757897
TRPC7 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Inference Prediction 15757897
(Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4: click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
Validation: In vitro validation Toggle 893349bafcc528f8346c51dc3420151d67b0126b2c122dd1017121c03fa0f69b
  Assay with recombinant proteins Reference
TRP channel construct Interactor construct
TRP channel Interactor Method Species Region Expression system Species Region Expression system
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Fusion protein-pull down assay Human 1-201 In vitro translation Human 572-662 E. coli 15757897
TRPC4 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Fusion protein-pull down assay Human 3-183 In vitro translation Human 572-662 E. coli 15757897
TRPC5 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Fusion protein-pull down assay Human 1-183 In vitro translation Human 572-662 E. coli 15757897
TRPC6 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Fusion protein-pull down assay Human 95-243 In vitro translation Human 572-662 E. coli 15757897
TRPC7 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Fusion protein-pull down assay Human 3-195 In vitro translation Human 572-662 E. coli 15757897
(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
TRPC1 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Co-immunoprecipitation HEK293T Human Full-length Human Full-length 15757897
TRPC3 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Co-immunoprecipitation HEK293T Human Full-length Human Full-length 15757897
TRPC4 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Co-immunoprecipitation HEK293T Human Full-length Human Full-length 15757897
TRPC6 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Co-immunoprecipitation HEK293T Human Full-length Human Full-length 15757897
(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 MxA Fusion protein-pull down assay Human 1-201 Human 572-662 15757897
TRPC4 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Fusion protein-pull down assay Human 3-183 Human 572-662 15757897
TRPC5 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Fusion protein-pull down assay Human 1-183 Human 572-662 15757897
TRPC6 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Yeast two-hybrid Human 3-259 Human 542-662 15757897
TRPC6 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Fusion protein-pull down assay Human 95-243 Human 572-662 15757897
TRPC7 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Fusion protein-pull down assay Human 3-195 Human 572-662 15757897
(Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4: click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
Functional consequence Toggle 893349bafcc528f8346c51dc3420151d67b0126b2c122dd1017121c03fa0f69b
TRP channel Interactor Method Post-translational modification Subcellular trafficking Activity Reference
TRPC6 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 MxA Calcium measurement Activation 15757897
(Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4: click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
TRP / Interactor

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