Truncation of the thyrotropin-releasing hormone receptor carboxyl tail causes constitutive activity and leads to impaired responsiveness in Xenopus oocytes and AtT20 cells.
We studied the activity of a truncated thyrotropin-releasing hormone receptor (TRH-R), which lacks the last 59 amino acids of the carboxyl tail, where Cys-335 was mutated to a stop codon (C335Stop) (Nussenzveig, D. R., Heinflink, M., and Gershengorn, M. C. (1993) J. Biol. Chem. 268, 2389-2392). In Xenopus laevis oocytes expressing C335Stop TRH-Rs, TRH binding was higher, whereas chloride current, 45Ca2+ efflux, and [Ca2+]i responses evoked by TRH were 23, 39, and 21%, respectively, of those in oocytes expressing wild type mouse pituitary TRH-Rs (WT TRH-Rs). In oocytes expressing C335Stop TRH-Rs, basal 45Ca2+ efflux and [Ca2+]i were twice those in oocytes expressing WT TRH-Rs; chelation of Ca2+ caused a rapid increase in holding current, which is consistent with basal activation; and coexpression with other receptors caused inhibition of the responses to the other cognate agonists. In AtT20 pituitary cells stably expressing C335Stop TRH-Rs, thyrotropin-releasing hormone (TRH)-independent inositol phosphate formation was 1.32 +/- 0.11-fold higher, basal [Ca2+]i was 1.8 +/- 0.2-fold higher, and the [Ca2+]i response to TRH was much lower than in cells expressing WT TRH-Rs. We conclude that a TRH-R mutant truncated at Cys-335 exhibits constitutive activity that results in desensitization of the response to TRH.
Matus-Leibovitch
N
N
Department of Physiology and Pharmacology, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv, Israel.
Nussenzveig
D R
DR
Gershengorn
M C
MC
Oron
Y
Y
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D007700:Kinetics; D051379:Mice; D009154:Mutation; D009865:Oocytes / Q000378:metabolism*; D018025:Receptors, Thyrotropin-Releasing Hormone / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014982:Xenopus laevis
D018025:Receptors, Thyrotropin-Releasing Hormone; D002118:Calcium