Journal article
The adaptor protein 14-3-3 binds to the calcium-sensing receptor and attenuates receptor-mediated Rho kinase signalling
Biochemical Journal, Vol.441(3), pp.995-1006
2012
Abstract
A yeast two-hybrid screen performed to identify binding partners of the CaR (calcium-sensing receptor) intracellular tail identified the adaptor protein 14-3-3θ as a novel binding partner that bound to the proximal membrane region important for CaR expression and signalling. The 14-3-3θ protein directly interacted with the CaR tail in pull-down studies and FLAG-tagged CaR co-immunoprecipitated with EGFP (enhanced green fluorescent protein)-tagged 14-3-3θ when co-expressed in HEK (human embryonic kidney)-293 or COS-1 cells. The interaction between the CaR and 14-3-3θ did not require a putative binding site in the membrane-proximal region of the CaR tail and was independent of PKC (protein kinase C) phosphorylation. Confocal microscopy demonstrated co-localization of the CaR and EGFP-14-3-3θ in the ER (endoplasmic reticulum) of HEK-293 cells that stably expressed the CaR (HEK-293/CaR cells), but 14-3-3θ overexpression had no effect on membrane expression of the CaR. Overexpression of 14-3-3θ in HEK-293/CaR cells attenuated CaR-mediated Rho signalling, but had no effect on ERK (extracellular-signal-regulated kinase) 1/2 signalling. Another isoform identified from the library, 14-3-3ζ, exhibited similar behaviour to that of 14-3-3θ with respect to CaR tail binding, cellular co-localization and impact on receptor-mediated signalling. However, unlike 14-3-3θ, this isoform, when overexpressed, significantly reduced CaR plasma membrane expression. Results indicate that 14-3-3 proteins mediate CaR-dependent Rho signalling and may modulate the plasma membrane expression of the CaR.
Details
- Title
- The adaptor protein 14-3-3 binds to the calcium-sensing receptor and attenuates receptor-mediated Rho kinase signalling
- Authors/Creators
- A. Arulpragasam (Author/Creator) - Sir Charles Gairdner HospitalA.L. Magno (Author/Creator) - The University of Western AustraliaE. Ingley (Author/Creator) - The University of Western AustraliaS.J. Brown (Author/Creator) - Sir Charles Gairdner HospitalA. D. Conigrave (Author/Creator) - The University of SydneyT. Ratajczak (Author/Creator) - Sir Charles Gairdner HospitalB.K. Ward (Author/Creator) - Sir Charles Gairdner Hospital
- Publication Details
- Biochemical Journal, Vol.441(3), pp.995-1006
- Publisher
- Portland Press, Ltd.
- Identifiers
- 991005540194207891
- Copyright
- © The Authors
- Murdoch Affiliation
- Murdoch University
- Language
- English
- Resource Type
- Journal article
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