Mol Pharmacol. 2009 Mar;75(3):466-77. doi: 10.1124/mol.108.051912. Epub 2008 Dec 3.
Canonical transient receptor potential channel (TRPC)3 and TRPC6 associate with large-conductance Ca2+-activated K+ (BKCa) channels: role in BKCa trafficking to the surface of cultured podocytes.
Kim, E. Y., Alvarez-Baron, C. P., Dryer, S. E.,
["Department of Biology and Biochemistry, University of Houston, Houston, TX 77204-5001, USA."]
["Department of Biology and Biochemistry, University of Houston, Houston, TX 77204-5001, USA."]
Large-conductance (BK(Ca) type) Ca(2+)-activated K(+) channels encoded by the Slo1 gene and various canonical transient receptor potential channels (TRPCs) are coexpressed in many cell types, including podocytes (visceral epithelial cells) of the renal glomerulus. In this study, we show by coimmunoprecipitation and GST pull-down assays that BK(Ca) channels can associate with endogenous TRPC3 and TRPC6 channels in differentiated cells of a podocyte cell line. Both types of TRPC channels colocalize with Slo1 in podocytes and in human embryonic kidney (HEK) 293T cells transiently coexpressing the TRPC channels with Slo1. In HEK293T cells, coexpression of TRPC6 increased surface expression of a Slo1 subunit splice variant (Slo1(VEDEC)) that is typically retained in intracellular compartments, as assessed by cell-surface biotinylation assays and confocal microscopy. Corresponding currents through BK(Ca) channels were also increased with TRPC6 coexpression, as assessed by whole-cell and excised inside-out patch recordings. By contrast, coexpression of TRPC3 had no effect on the surface expression of BK(Ca) channels in HEK293T cells or on the amplitudes of currents in whole cells or excised patches. In podocytes, small interfering RNA knockdown of endogenous TRPC6 reduced steady-state surface expression of endogenous Slo1 channels, but knockdown of TRPC3 had no effect. TRPC6, but not TRPC3 knockdown also reduced voltage-evoked outward current through podocyte BK(Ca) channels. These data indicate that TRPC6 and TRPC3 channels can bind to Slo1, and this colocalization may allow them to serve as a source of Ca(2+) for the activation of BK(Ca) channels. TRPC6 channels also play a role in the regulation of surface expression of a subset of podocyte BK(Ca) channels.
PMID: 19052171

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Screening
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Experimental screening | Non-experimental screening | Reference | ||||||||
TRP channel construct | Interactor source | |||||||||
TRP channel | Interactor | Method | Species | Region | Species | Organ/tissue | Sample type | |||
TRPC3 |
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BKca | Inference | Prediction | 19052171 | |||||
TRPC6 |
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BKca | Inference | Prediction | 19052171 |
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click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)

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Validation: In vitro validation
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Assay with recombinant proteins | Reference | |||||||||
TRP channel construct | Interactor construct | |||||||||
TRP channel | Interactor | Method | Species | Region | Expression system | Species | Region | Expression system | ||
TRPC3 |
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BKca | Fusion protein-pull down assay | Not used | Mouse podocyte lysates | Chicken | Full-length | E. coli | 19052171 | |
TRPC6 |
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BKca | Fusion protein-pull down assay | Not used | Mouse podocyte lysates | Chicken | Full-length | E. coli | 19052171 |
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click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)

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Validation: In vivo validation
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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 | ||||
TRPC3 |
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BKca | Co-immunoprecipitation | Mouse podocyte lysates | 19052171 | |||||
TRPC3 |
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BKca | Co-immunoprecipitation | HEK293T | Not specified | Full-length | Not specified | Full-length | 19052171 | |
TRPC3 |
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BKca | Co-immunofluorescence staining | HEK293T | Not specified | Full-length | Not specified | Full-length | 19052171 | |
TRPC6 |
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BKca | Co-immunoprecipitation | Mouse podocyte lysates | 19052171 | |||||
TRPC6 |
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BKca | Co-immunoprecipitation | HEK293T | Not specified | Full-length | Not specified | Full-length | 19052171 | |
TRPC6 |
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BKca | Co-immunofluorescence staining | HEK293T | Not specified | Full-length | Not specified | Full-length | 19052171 |
(
:
click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
