J Neurochem. 2007 Dec;103(6):2391-400. Epub 2007 Oct 17.
Direct binding of alpha-actinin enhances TRPP3 channel activity.
Li, Q., Dai, X. Q., Shen, P. Y., Wu, Y., Long, W., Chen, C. X., Hussain, Z., Wang, S., Chen, X. Z.,
["Membrane Protein Research Group, Department of Physiology, University of Alberta, Edmonton, Alberta, Canada."]
["Membrane Protein Research Group, Department of Physiology, University of Alberta, Edmonton, Alberta, Canada."]
Transient receptor potential (TRP) polycystin 2 and 3 (TRPP2 and 3) are homologous members of the TRP superfamily of cation channels but have different physiological functions. TRPP2 is part of a flow sensor, and is defective in autosomal dominant polycystic kidney disease and implicated in left-right asymmetry development. TRPP3 is reported to implicate in sour tasting in bipolar cells of taste buds of the tongue and in the regulation of pH-sensitive action potential in neurons surrounding the central canal of spinal cord. TRPP3 is present in both excitable and non-excitable cells in various tissues, such as retina, brain, heart, testis, and kidney, but its common and cell type-specific functional characteristics remain largely unknown. In this study, we investigated physical and functional interactions between TRPP3 and alpha-actinin, an actin-bundling protein known to regulate several types of ion channels. We employed planer lipid bilayer electrophysiology system to study the function of TRPP3 channel that was affinity-purified from Madin-Darby canine kidney cells. Upon reconstitution in bilayer, TRPP3 exhibited cation channel activities that were substantially augmented by alpha-actinin. The TRPP3-alpha-actinin association was documented by co-immunoprecipitation using native cells and tissues, yeast two-hybrid, and in vitro binding assays. Further, TRPP3 was abundantly present in mouse brain where it associates with alpha-actinin-2. Taken together, alpha-actinin not only attaches TRPP3 to the cytoskeleton but also up-regulates TRPP3 channel function. It remains to be determined whether the TRPP3-alpha-actinin interaction is relevant to acid sensing and other functions in neuronal and non-neuronal cells.
PMID: 17944866

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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 | |||
TRPP2 |
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ơ-actinin | Inference | Prediction | 17944866 | |||||
TRPP2 |
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ơ-actinin-1 | Inference | Prediction | 17944866 | |||||
TRPP2 |
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ơ-actinin-2 | Inference | Prediction | 17944866 |
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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 | ||
TRPP2 |
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ơ-actinin | Fusion protein-pull down assay | Human | 1-97 | E. coli | Chicken | Full-length | In vitro translation | 17944866 |
TRPP2 |
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ơ-actinin | Fusion protein-pull down assay | Human | 562-805 | E. coli | Chicken | Full-length | In vitro translation | 17944866 |
TRPP2 |
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ơ-actinin-1 | Yeast two-hybrid | Human | 1-97 | Yeast | Human | 334-892 | Yeast | 17944866 |
TRPP2 |
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ơ-actinin-1 | Yeast two-hybrid | Human | 562-805 | Yeast | Human | 334-892 | Yeast | 17944866 |
TRPP2 |
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ơ-actinin-2 | Yeast two-hybrid | Human | 1-97 | Yeast | Human | 341-894 | Yeast | 17944866 |
TRPP2 |
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ơ-actinin-2 | Yeast two-hybrid | Human | 562-805 | Yeast | Human | 341-894 | Yeast | 17944866 |
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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 | ||||
TRPP2 |
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ơ-actinin-1 | Co-immunoprecipitation | HEK293 | 17944866 | |||||
TRPP2 |
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ơ-actinin-2 | Co-immunoprecipitation | Mouse brain lysates | 17944866 |
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click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)

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Characterization
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Binding region mapping | Stoichiometry | Affinity (Kd) | Reference | |||||||
TRP channel | Interactor | |||||||||
TRP channel | Interactor | Method | Species | Region | Species | Region | ||||
TRPP2 |
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ơ-actinin-2 | Yeast two-hybrid | Human | 1-97 | Human | 596-728 | 17944866 |
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click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)

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Functional consequence
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TRP channel | Interactor | Method | Post-translational modification | Subcellular trafficking | Activity | Reference | ||||||
TRPP2 |
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ơ-actinin | Patch clamp | Activation | 17944866 |
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click the arrow icon to show interactions only between the corresponding TRP channel and the interactor)
