WDR36 acts as a scaffold protein tethering a G-protein-coupled receptor, Gαq and phospholipase Cβ in a signalling complex

Author:

Cartier Andréane1,Parent Audrey1,Labrecque Pascale1,Laroche Geneviève1,Parent Jean-Luc1

Affiliation:

1. Service de Rhumatologie, Département de Médecine, Faculté de Médecine et des Sciences de la Santé, Université de Sherbrooke, Institut de Pharmacologie de Sherbrooke and Centre de Recherche Clinique Etienne-Lebel, Sherbrooke, QC J1H 5N4, Canada

Abstract

We identified the WD-repeat-containing protein, WDR36, as an interacting partner of the β isoform of thromboxane A2 receptor (TPβ) by yeast two-hybrid screening. We demonstrated that WDR36 directly interacts with the C-terminus and the first intracellular loop of TPβ by in vitro GST-pulldown assays. The interaction in a cellular context was observed by co-immunoprecipitation, which was positively affected by TPβ stimulation. TPβ–WDR36 colocalization was detected by confocal microscopy at the plasma membrane in non-stimulated HEK293 cells but the complex translocated to intracellular vesicles following receptor stimulation. Coexpression of WDR36 and its siRNA-mediated knockdown, respectively, increased and inhibited TPβ-induced Gαq signalling. Interestingly, WDR36 co-immunoprecipitated with Gαq, and promoted TPβ–Gαq interaction. WDR36 also associated with phospholipase Cβ (PLCβ) and increased the interaction between Gαq and PLCβ, but prevented sequestration of activated Gαq by GRK2. In addition, the presence of TPβ in PLCβ immunoprecipitates was augmented by expression of WDR36. Finally, disease-associated variants of WDR36 affected its ability to modulate Gαq-mediated signalling by TPβ. We report that WDR36 acts as a new scaffold protein tethering a G-protein-coupled receptor, Gαq and PLCβ in a signalling complex.

Publisher

The Company of Biologists

Subject

Cell Biology

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