J Biol Chem. 2013 Jun 21;288(25):18407-20. doi: 10.1074/jbc.M113.463059. Epub 2013 May 3.

Gain-of-function mutations in transient receptor potential C6 (TRPC6) activate extracellular signal-regulated kinases 1/2 (ERK1/2).External 2231691f894ba696de1310221b0a0dbbb31a7251e75115c265587c3d9d5f507c

Chiluiza, D., Krishna, S., Schumacher, V. A., Schlondorff, J.,
["Division of Nephrology, Department of Medicine, Beth Israel Deaconess Medical Center, Boston, Massachusetts 02215, USA."]
Gain-of-function mutations in the canonical transient receptor potential 6 (TRPC6) gene are a cause of autosomal dominant focal segmental glomerulosclerosis (FSGS). The mechanisms whereby abnormal TRPC6 activity results in proteinuria remain unknown. The ERK1/2 MAPKs are activated in glomeruli and podocytes in several proteinuric disease models. We therefore examined whether FSGS-associated mutations in TRPC6 result in activation of these kinases. In 293T cells and cultured podocytes, overexpression of gain-of-function TRPC6 mutants resulted in increased ERK1/2 phosphorylation, an effect dependent upon channel function. Pharmacologic inhibitor studies implicated several signaling mediators, including calmodulin and calcineurin, supporting the importance of TRPC6-mediated calcium influx in this process. Through medium transfer experiments, we uncovered two distinct mechanisms for ERK activation by mutant TRPC6, a cell-autonomous, EGF receptor-independent mechanism and a non-cell-autonomous mechanism involving metalloprotease-mediated release of a presumed EGF receptor ligand. The inhibitors KN-92 and H89 were able to block both pathways in mutant TRPC6 expressing cells as well as the prolonged elevation of intracellular calcium levels upon carbachol stimulation seen in these cells. However, these effects appear to be independent of their effects on calcium/calmodulin-dependent protein kinase II and PKA, respectively. Phosphorylation of Thr-70, Ser-282, and Tyr-31/285 were not necessary for ERK activation by mutant TRPC6, although a phosphomimetic TRPC6 S282E mutant was capable of ERK activation. Taken together, these results identify two pathways downstream of mutant TRPC6 leading to ERK activation that may play a role in the development of FSGS.
PMID: 23645677External 2231691f894ba696de1310221b0a0dbbb31a7251e75115c265587c3d9d5f507c
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
TRPC6 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 PKA Co-immunoprecipitation HEK293T Human Full-length Not specified Not specified 23645677
(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 PKA In vitro PTM assay Phosphorylation (Thr-70) 23645677
TRPC6 Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4 PKA In vivo PTM assay Phosphorylation (Thr-70) 23645677
(Link 2bd4d11adb659cddf58197a94e201f0a44c55d8d7cb427c624971b42e122c0a4: click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
TRP / Interactor

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