Article: article from journal or magazin.
The adaptor complex 2 directly interacts with the alpha 1b-adrenergic receptor and plays a role in receptor endocytosis.
Journal of Biological Chemistry
Using the yeast two-hybrid system, we identified the mu 2 subunit of the clathrin adaptor complex 2 as a protein interacting with the C-tail of the alpha 1b-adrenergic receptor (AR). Direct association between the alpha 1b-AR and mu 2 was demonstrated using a solid phase overlay assay. The alpha 1b-AR/mu 2 interaction occurred inside the cells, as shown by the finding that the transfected alpha 1b-AR and the endogenous mu 2 could be coimmunoprecipitated from HEK-293 cell extracts. Mutational analysis of the alpha 1b-AR revealed that the binding site for mu 2 does not involve canonical YXX Phi or dileucine motifs but a stretch of eight arginines on the receptor C-tail. The binding domain of mu 2 for the receptor C-tail involves both its N terminus and the subdomain B of its C-terminal portion. The alpha 1b-AR specifically interacted with mu 2, but not with the mu 1, mu 3, or mu 4 subunits belonging to other AP complexes. The deletion of the mu 2 binding site in the C-tail markedly decreased agonist-induced receptor internalization as demonstrated by confocal microscopy as well as by the results of a surface receptor biotinylation assay. The direct association of the adaptor complex 2 with a G protein-coupled receptor has not been reported so far and might represent a common mechanism underlying clathrin-mediated receptor endocytosis.
Adaptor Protein Complex 2/chemistry, Adaptor Protein Complex 2/genetics, Adaptor Protein Complex mu Subunits/chemistry, Adaptor Protein Complex mu Subunits/genetics, Amino Acid Sequence, Animals, Binding Sites/genetics, Biotinylation, Blotting, Western, Cell Line, Clathrin/physiology, Cricetinae, Electrophoresis, Polyacrylamide Gel, Endocytosis/drug effects, Epinephrine/pharmacology, Escherichia coli/genetics, Gene Deletion, Gene Expression, Glutathione Transferase/genetics, Green Fluorescent Proteins, Humans, Immunosorbent Techniques, Luminescent Proteins/genetics, Microscopy, Confocal, Molecular Sequence Data, Mutagenesis, Polymerase Chain Reaction, Receptors, Adrenergic, alpha-1/chemistry, Receptors, Adrenergic, alpha-1/genetics, Recombinant Fusion Proteins, Transfection, Two-Hybrid System Techniques
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