title: Persistent signaling by thyrotropin-releasing hormone receptors correlates with G-protein and receptor levels.
abstract: G-protein-coupled receptors with dissociable agonists for thyrotropin, parathyroid hormone, and sphingosine-1-phosphate were found to signal persistently hours after agonist withdrawal. Here we show that mouse thyrotropin-releasing hormone (TRH) receptors, subtypes 2 and 1(TRH-R2 and TRH-R1), can signal persistently in HEK-EM293 cells under appropriate conditions, but TRH-R2 exhibits higher persistent signaling activity. Both receptors couple primarily to Gα(q/11). To gain insight into the mechanism of persistent signaling, we compared proximal steps of inositolmonophosphate (IP1) signaling by TRH-Rs. Persistent signaling was not caused by slower dissociation of TRH from TRH-R2 (t(1/2)=77 ± 8.1 min) compared with TRH-R1 (t(1/2)=82 ± 12 min) and was independent of internalization, as inhibition of internalization did not affect persistent signaling (115% of control), but required continuously activated receptors, as an inverse agonist decreased persistent signaling by 60%. Gα(q/11) knockdown decreased persistent signaling by TRH-R2 by 82%, and overexpression of Gα(q/11) induced persistent signaling in cells expressing TRH-R1. Lastly, persistent signaling was induced in cells expressing high levels of TRH-R1. We suggest that persistent signaling by TRHRs is exhibited when sufficient levels of agonist/receptor/G-protein complexes are established and maintained and that TRH-R2 forms and maintains these complexes more efficiently than TRH-R1.
authors.affiliation: Laboratory of Endocrinology and Receptor Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892-8029, USA.
authors.forename: Alisa
authors.initials: A
authors.lastname: Boutin
authors.forename: Michael D
authors.initials: MD
authors.lastname: Allen
authors.forename: Susanne
authors.initials: S
authors.lastname: Neumann
authors.forename: Marvin C
authors.initials: MC
authors.lastname: Gershengorn
mesh_terms: D000818:Animals; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000502:physiology*; D007295:Inositol Phosphates / Q000096:biosynthesis; D051379:Mice; D043562:Receptors, G-Protein-Coupled; D018025:Receptors, Thyrotropin-Releasing Hormone / Q000819:agonists / Q000235:genetics / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D007295:Inositol Phosphates; D043562:Receptors, G-Protein-Coupled; D018025:Receptors, Thyrotropin-Releasing Hormone; C528487:TRH-R1 protein, mouse; C002647:inositol 1-phosphate; D043802:GTP-Binding Protein alpha Subunits, Gq-G11