title: TSH receptor mutation V509A causes familial hyperthyroidism by release of interhelical constraints between transmembrane helices TMH3 and TMH5.
abstract: Mutations of the human thyrotrophin receptor (TSH-R) are a cause of thyroid adenomas and hyperthyroidism. Here we study mechanisms of receptor activation in a genomic TSH-R variant V509A located in transmembrane helix (TMH) 3, which we identify in a family with congenital hyperthyroidism, multiple adenomas and follicular thyroid cancer. Using molecular modelling and dynamic simulation, we predicted the release of amino acid residue A593 (located opposite in domain TMH5) from a tight 'knob-and-hole' interaction with TMH3, physiologically constrained in the native receptor state by the bulky side chain of V509. To experimentally validate this concept, we generated mutant TSH-R expression constructs for functional in vitro studies. TSH-R mutant V509A showed a 2.8-fold increase in basal cAMP production, confirming constitutive TSH-R activation. The addition of a second site suppressor mutant A593V to TSH-R V509A resulted in the normalization of basal cAMP release, and the dose-responsiveness to TSH ligand was maintained. These data thus demonstrate that TSH-R V509A activation is caused by the release of TMH3-TMH5 interhelical constraints, while the native TSH-R conformation is re-stabilized by the introduction of a spacious valine residue at position 593. In conclusion, we delineate a novel mechanism of constitutive TSH-R activation, leading to thyroid hyperfunction and neoplasia.
authors.affiliation: Pediatric Endocrinology, University Children's Hospital, University of Ulm, Prittwitzstrasse 43, D-89075 Ulm, Germany. beate.karges@medizin.uni-ulm.de
authors.forename: Beate
authors.initials: B
authors.lastname: Karges
authors.forename: Gerd
authors.initials: G
authors.lastname: Krause
authors.forename: Janos
authors.initials: J
authors.lastname: Homoki
authors.forename: Klaus-Michael
authors.initials: KM
authors.lastname: Debatin
authors.forename: Nicolas
authors.initials: N
authors.lastname: de Roux
authors.forename: Wolfram
authors.initials: W
authors.lastname: Karges
mesh_terms: D000236:Adenoma / Q000235:genetics; D000293:Adolescent; D001483:Base Sequence; D018265:Carcinoma, Papillary, Follicular / Q000235:genetics; D002675:Child, Preschool; D000242:Cyclic AMP / Q000378:metabolism; D004247:DNA; D005260:Female; D020022:Genetic Predisposition to Disease; D018095:Germ-Line Mutation; D006042:Goiter / Q000235:genetics; D018257:Helix-Loop-Helix Motifs; D006801:Humans; D006980:Hyperthyroidism / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D008958:Models, Molecular; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D010375:Pedigree; D011989:Receptors, Thyrotropin / Q000235:genetics* / Q000378:metabolism; D013964:Thyroid Neoplasms / Q000235:genetics
chemical_list: D011989:Receptors, Thyrotropin; D004247:DNA; D000242:Cyclic AMP