Polarity of TRH receptors in transfected MDCK cells is independent of endocytosis signals and G protein coupling.

TitlePolarity of TRH receptors in transfected MDCK cells is independent of endocytosis signals and G protein coupling.
Publication TypeJournal Article
Year of Publication1996
AuthorsYeaman C, Heinflink M, Falck-Pedersen E, Rodriguez-Boulan E, Gershengorn MC
JournalAm J Physiol
Issue3 Pt 1
Date Published1996 Mar
KeywordsAmino Acid Sequence, Animals, Cell Line, Cell Membrane, Dogs, Endocytosis, GTP-Binding Proteins, Kidney, Kinetics, Molecular Sequence Data, Protein Structure, Secondary, Radioligand Assay, Receptors, Thyrotropin-Releasing Hormone, Recombinant Proteins, Sequence Deletion, Signal Transduction, Thyrotropin-Releasing Hormone, Transfection, Tritium

Information concerning the molecular sorting of G protein-coupled receptors in polarized epithelial cells is limited. Therefore, we have expressed the receptor for thyrotropin-releasing hormone (TRH) in Madin-Darby canine kidney (MDCK) cells by adenovirus-mediated gene transfer to determine its distribution in a model cell system and to begin analyzing the molecular information responsible for its distribution. Equilibrium binding of [methyl-3H]TRH to apical and basolateral surfaces of polarized MDCK cells reveals that TRH receptors are expressed predominantly (>80%) on the basolateral cell surface. Receptors undergo rapid endocytosis following agonist binding; up to 80% are internalized in 15 min. A mutant receptor missing the last 59 residues, C335Stop, is poorly internalized (<10%) but is nevertheless basolaterally expressed (>85%). A second mutant TRH receptor, delta218-263, lacks essentially all of the third intracellular loop and is not coupled to G proteins on binding agonist. This receptor internalizes TRH approximately half as efficiently as wild-type TRH receptors but is nevertheless strongly polarized to the basolateral surface (>90%). These results indicate that molecular sequences responsible for basolateral accumulation of TRH receptors can be segregated from signals for ligand-induced receptor endocytosis and coupling to heterotrimeric G proteins.

Alternate JournalAm J Physiol
PubMed ID8638654
Grant ListR01 GM034107 / GM / NIGMS NIH HHS / United States
DK-07313 / DK / NIDDK NIH HHS / United States
DK-43046 / DK / NIDDK NIH HHS / United States
GM-34107-11 / GM / NIGMS NIH HHS / United States

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