TRUNCATION OF THE THYROTROPIN-RELEASING-HORMONE RECEPTOR CARBOXYL TAIL CAUSES CONSTITUTIVE ACTIVITY AND LEADS TO IMPAIRED RESPONSIVENESS INXENOPUS OOCYTES AND ATT20 CELLS

We studied the activity of a truncated thyrotropin-releasing hormone r eceptor (TRH-R), which lacks the last 59 amino acids of the carboxyl t ail, where Cys-335 was mutated to a stop codon (C335Stop) (Nussenzveig , D. R., Heinflink, M., and Gershengorn, M. C. (1993) J. Biol. Chem. 2 68, 2389-2392). In Xenopus laevis oocytes expressing C335Stop TRH-Rs, TRH binding was higher, whereas chloride current, Ca-45(2+) efflux, an d [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 Ca-45(2+) efflux and [Ca2+](i) were twice those in oocytes expressing WT TRH-Rs; chela tion of Ca2+ caused a rapid increase in holding current, which is cons istent with basal activation; and coexpression with other receptors ca used inhibition of the responses to the other cognate agonists. In AtT 20 pituitary cells stably expressing C335Stop TRH-Rs, thyrotropin-rele asing 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 exhib its constitutive activity that results in desensitization of the respo nse to TRH.