Functional expression and molecular characterization of the thyrotrophin-releasing hormone receptor from the rat anterior pituitary gland

Abstract

We have isolated the TRH receptor (TRH-R) from a rat anterior pituitary cDNA library, determined its sequence and examined its functional characteristics. Expression studies were carried out in Xenopus oocytes and in COS-7 cells. Microinjection of sense RNA transcripts into Xenopus oocytes showed electrophysiological responses of between 800 and 1000 nA under voltage-clamp conditions. COS-7 cells were transiently transfected with the cDNA clone under the control of a cytomegalovirus promoter and inositol phosphate (IP) measurements carried out. In TRH-R transfected cells, TRH (100 nM) produced an approximately twofold increase in total IP production. In-situ hybridization in the rat anterior pituitary revealed a heterogeneous distribution of label, a characteristic pattern of TRH-R expression. The rat 3.3 kb insert coded for a protein of 411 amino acids compared with 393 for the mouse protein. Over its length, the rat TRH-R protein showed considerable homology with that of the mouse, except for a deletion of 232 bp in the 3'-coding region. This deletion did not appear to affect the functional characteristics of the receptor, as shown by electrophysiological studies with Xenopus oocytes and by transfection of the cDNA into COS-7 cells. The sequence given for the 3'-untranslated region is 1.5 kb longer than that reported for the mouse receptor, and extends to the poly(A) tail.

Real Time Visualization of Agonist-mediated Redistribution and Internalization of a Green Fluorescent Protein-tagged Form of the Thyrotropin-releasing Hormone Receptor

“…TRH functions via binding to a seven-transmembrane element-G protein-coupled receptor (3), which, by interacting selectivity with G q and G 11 , causes activation of phospholipase C␤1 and the hydrolysis of phosphatidylinositol 4,5-bisphosphate (4 -7). As with other G protein-coupled receptors there has been great interest in the mechanisms of regulation of the TRH receptor (TRHR) (6 -11).…”

Homo- and Hetero-oligomerization of Thyrotropin-releasing Hormone (TRH) Receptor Subtypes

“…Our findings suggest that TRHR oligomerization results in the creation of novel receptor units that can be differentially regulated by ␤-arrestins. Eukaryotic Expression Constructs-Rat TRHR1 coding region was PCR-amplified from a previously described clone (38) and inserted into pcDNA3 vector. The TRHR1/Rluc and TRHR1/EYFP constructs have been previously described (30).…”

The human neuroendocrine thyrotropin-releasing hormone receptor promoter is activated by the haematopoietic transcription factor c-Myb

“…TRH and TRH receptor (TRHR) are found distributed throughout the central and peripheral nervous systems as well as in other tissues [3][4][5][6] including thymus [6] and small intestine epithelial cells [7], consistent with its proposed role in the immune system. cDNAs encoding the highly conserved TRHR have been cloned from mouse, rat and hamster cell lines [8][9][10], from human brain [11] and from avian and bovine tissues [12,13]. This receptor is now also designated TRHR1, to be distinguished from TRHR2, a recently discovered subtype of the rat TRH receptor, 51 % identical to TRHR1 but with a distinct mRNA tissue distribution [14,15].…”