Homo- and hetero-oligomerization of thyrotropin-releasing hormone (TRH) receptor subtypes. Differential regulation of beta-arrestins 1 and 2.
G-protein-coupled receptors (GPCRs) are regulated by a complex network of mechanisms such as oligomerization and internalization. Using the GPCR subtypes for thyrotropin-releasing hormone (TRHR1 and TRHR2), the aim of this study was to determine if subtype-specific differences exist in the trafficking process. If so, we wished to determine the impact ... of homo- and hetero-oligomerization on TRHR subtype trafficking as a potential mechanism for the differential cellular responses induced by TRH. Expression of either beta-arrestin 1 or 2 promoted TRHR1 internalization. In contrast, only beta-arrestin 2 could enhance TRHR2 internalization. The preference for beta-arrestin 2 by TRHR2 was supported by the impairment of TRHR2 trafficking in mouse embryonic fibroblasts (MEFs) from either a beta-arrestin 2 knockout or a beta-arrestin 1/2 knockout, while TRHR1 trafficking was only abolished in MEFs lacking both beta-arrestins. The differential beta-arrestin-dependence of TRHR2 was directly measured in live cells using bioluminescence resonance energy transfer (BRET). Both BRET and confocal microscopy were also used to demonstrate that TRHR subtypes form hetero-oligomers. In addition, these hetero-oligomers have altered internalization kinetics compared with the homo-oligomer. The formation of TRHR1/2 heteromeric complexes increased the interaction between TRHR2 and beta-arrestin 1. This may be due to conformational differences between TRHR1/2 hetero-oligomers versus TRHR2 homo-oligomers as a mutant TRHR1 (TRHR1 C335Stop) that does not interact with beta-arrestins, could also enhance TRHR2/beta-arrestin 1 interaction. This study demonstrates that TRHR subtypes are differentially regulated by the beta-arrestins and also provides the first evidence that the interactions of TRHRs with beta-arrestin may be altered by hetero-oligomer formation.
Mesh Terms:
Animals, Arrestins, COS Cells, Cell Line, Cercopithecus aethiops, Cloning, Molecular, Fibroblasts, Genetic Vectors, Green Fluorescent Proteins, Kidney, Kinetics, Luminescent Proteins, Macromolecular Substances, Mice, Mice, Knockout, Polymerase Chain Reaction, Protein Isoforms, Rats, Receptors, Thyrotropin-Releasing Hormone, Recombinant Fusion Proteins, Transfection
Animals, Arrestins, COS Cells, Cell Line, Cercopithecus aethiops, Cloning, Molecular, Fibroblasts, Genetic Vectors, Green Fluorescent Proteins, Kidney, Kinetics, Luminescent Proteins, Macromolecular Substances, Mice, Mice, Knockout, Polymerase Chain Reaction, Protein Isoforms, Rats, Receptors, Thyrotropin-Releasing Hormone, Recombinant Fusion Proteins, Transfection
J. Biol. Chem.
Date: Dec. 27, 2002
PubMed ID: 12393857
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