FUNCTIONAL-CHARACTERISTICS OF 3 NEW GERMLINE MUTATIONS OF THE THYROTROPIN RECEPTOR GENE CAUSING AUTOSOMAL-DOMINANT TOXIC THYROID HYPERPLASIA

We report three unrelated families in which hyperthyroidism associated with thyroid hyperplasia was transmitted in an autosomal dominant fas hion, in the absence of signs of autoimmunity. Exon 10 of the TSH rece ptor gene was directly sequenced after PCR amplification from DNA of p eripheral leukocytes. In one family, a C to A transversion resulted in an S505R substitution in the third transmembrane segment; in the seco nd, an A to T transversion caused a N650Y substitution in the sixth tr ansmembrane segment; and in the third family, an A to G transition res ulted in an N670S substitution in the seventh transmembrane segment. W hen expressed by transfection in COS-7 cells, each mutated receptor di splayed an increase in constitutive stimulation of cAMP production; no effect on basal accumulation of inositol phosphates (IP) could be det ected. In binding studies, cells transfected with wild-type or mutated receptors showed similar levels of expression, with the mutated recep tors displaying similar or slightly increased affinity for bovine TSH (bTSH) binding. Cells transfected with S505R and N650Y mutants showed a similar cAMP maximal TSH-stimulated accumulation over the cells tran sfected with the wild type, whereas N670S transfectants showed a blunt ed response with an increase in EC(50). A higher IP response to 100 mU /mL bTSH over that obtained with the wild-type receptor was obtained i n cells transfected with N650Y; in contrast, cells transfected with S5 05R showed a blunted IP production (50% less), and the N670S mutant co mpletely lost the ability to stimulate IP accumulation in response to bTSH. The differential effects of individual mutations on stimulation by bTSH of cAMP or LP accumulation suggest that individual mutant rece ptors may achieve different active conformations with selective abilit ies to couple to G(s) alpha and to G(q) alpha.