title: WDR36 acts as a scaffold protein tethering a G-protein-coupled receptor, Gαq and phospholipase Cβ in a signalling complex.
abstract: We identified the WD-repeat-containing protein, WDR36, as an interacting partner of the β isoform of thromboxane A(2) 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.
authors.affiliation: 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.
authors.forename: Andréane
authors.initials: A
authors.lastname: Cartier
authors.forename: Audrey
authors.initials: A
authors.lastname: Parent
authors.forename: Pascale
authors.initials: P
authors.lastname: Labrecque
authors.forename: Geneviève
authors.initials: G
authors.lastname: Laroche
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Parent
mesh_terms: D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid / Q000494:pharmacology; D004789:Enzyme Activation; D005136:Eye Proteins / Q000378:metabolism*; D054769:G-Protein-Coupled Receptor Kinase 2 / Q000378:metabolism; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D047468:Immunoprecipitation; D007527:Isoenzymes / Q000378:metabolism; D008856:Microscopy, Fluorescence; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D046912:Multiprotein Complexes / Q000378:metabolism; D054799:Phospholipase C beta / Q000378:metabolism*; D011485:Protein Binding; D020033:Protein Isoforms / Q000378:metabolism; D021381:Protein Transport; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D044244:Receptors, Thromboxane A2, Prostaglandin H2 / Q000819:agonists / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D005136:Eye Proteins; D007527:Isoenzymes; D046912:Multiprotein Complexes; D020033:Protein Isoforms; D043562:Receptors, G-Protein-Coupled; D044244:Receptors, Thromboxane A2, Prostaglandin H2; C502341:WDR36 protein, human; D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; C496567:GRK2 protein, human; D054769:G-Protein-Coupled Receptor Kinase 2; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D054799:Phospholipase C beta; D043802:GTP-Binding Protein alpha Subunits, Gq-G11