Congenital neonatal hyperthyroidism caused by germline mutations in the TSH receptor gene.

Neonatal hyperthyroidism, a rare and serious disorder, occurs in two forms. An autoimmune form associated with maternal Graves' disease, resulting from transplacental passage of maternal thyroid-stimulating antibodies, and a non-autoimmune form, resulting from mutations in the stimulatory G protein or the thyrotropin receptor (TSHR) causing constitutive activation of intracellular signaling cascades. To date, 29 separate cases of thyrotoxicosis caused by germline mutations of the TSHR have been documented. These cases have expressed themselves in a range of clinical consequences. This report describes a new case of a newborn with non-autoimmune hyperthyroidism secondary to a constitutively active TSHR mutation (S281N) whose clinical course was complicated by severe respiratory compromise. Typical clinical findings in this disorder are discussed by a review of all previously published cases.

Repulsive separation of the cytoplasmic ends of transmembrane helices 3 and 6 is linked to receptor activation in a novel thyrotropin receptor mutant (M626I).

Ligand-dependent activation of G protein-coupled receptors (GPCRs) involves repositioning of the juxtacytoplasmic ends of transmembrane helices TM3 and TM6. This concept, inferred from site-directed spin labeling studies, is supported by chemical cross-linking of the cytoplasmic ends of TM3 and TM6 blocking GPCR activation. Here we report a novel constitutive active mutation (M626I) in TM6 of the TSH receptor (TSHR), identified in affected members of a family with nonautoimmune hyperthyroidism. The specific constitutive activity of M626I, measured by its basal cAMP generation corrected for cell surface expression, was 13-fold higher than that of wild-type TSHR. Homology modeling of the TSHR serpentine domain based on the rhodopsin crystal structure suggests that M626 faces the side chain of I515 of TM3 near the membrane-cytoplasmic junction. Steric hindrance of the introduced isoleucine by I515 is consistent with the fact that shorter or more flexible side chains at position 626 did not increase constitutivity. Furthermore, a reciprocal mutation at position 515 (I515M), when introduced into the M626I background, acts as revertant mutation by allowing accommodation of the isoleucine sidechain at position 626 and fully restoring the constitutive activity to the level of wild-type TSHR. Thus, repulsive separation of the juxtacytoplasmic TM6 and TM3 in the M626I model conclusively demonstrates a direct link between the opening of this cytoplasmic face of the receptor structure and G protein coupling.

Thyroid transcription factor 1 rescues PAX8/p300 synergism impaired by a natural PAX8 paired domain mutation with dominant negative activity.

Mutations in the paired domain transcription factor PAX8 are a rare cause of congenital hypothyroidism due to thyroid dysgenesis. We identified a novel and unique PAX8 mutation segregating in seven affected members of a three-generations family. The mutation replaces an invariant serine residue within helix 2 of the paired DNA-binding domain for phenylalanine. The mutant protein (PAX8-S48F) does not induce the thyroglobulin promoter in nonthyroid cells, but displays almost half of wild-type PAX8 activity in thyroid cells. PAX8-S48F shows no defect in expression, nuclear targeting, or DNA binding and retains the ability to synergize with thyroid transcription factor 1 (TTF-1, NKX2.1). However, we found that in nonthyroid cells, the acetylation-independent synergism with the general transcriptional adaptor p300 is completely abrogated, suggesting that PAX8-S48F may be unable to efficiently recruit p300. Reconstitution experiments in nonthyroid cells reveal that TTF-1 can partially rescue PAX8-S48F/p300 synergism and thus reproduce the situation in thyroid cells. These functional characteristics result in a dominant negative effect of PAX8-S48F on coexpressed wild-type PAX8 activity, which is not observed in paired domain mutations with DNA binding defect. Our results describe the first dominant negative missense mutation in a paired domain and provide evidence for a crucial role of the p300 coactivator in mediating the functional synergism between PAX8 and TTF-1 in thyroid-specific gene expression.

Resistance to thyroid hormone in a patient with thyroid dysgenesis.

We report a patient with the unusual coincidence of two rare congenital disorders, lingual ectopy of the thyroid gland and resistance to thyroid hormone (RTH), resulting in impaired thyroid hormone production and action, respectively. The proposita had a positive thyrotropin (TSH) newborn screening test (350 mU/L, confirmed) with normal thyroxine (T4) and no clinical signs of hypothyroidism. A scintiscan revealed lingual but no orthotopic thyroid tissue. Levothyroxine (LT4) replacement failed to reduce TSH and was discontinued after four months owing to significantly elevated free T4. Her physical and mental development was unremarkable, and she was considered to be clinically euthyroid throughout childhood, even though she received either no T4 or a dose insufficient to lessen hyperthyrotropinemia. At the age of 15 years, T4 was gradually increased to a supraphysiological dose of 300 microg/d, resulting in the normalization of the serum TSH level, and subjective improvements in her ability to concentrate. The proposita's mother was clinically euthyroid, had a palpable diffuse goiter, and thyroid function tests consistent with RTH. This diagnosis was confirmed by detection of a heterozygous mutation (R320H) in the thyroid hormone receptor-beta (TR-beta) gene found in both the proposita and her mother. Under the high-dose T4 regimen, the patient's TSH and free T4 values resembled those of untreated patients with TRbeta R320H mutation, suggesting that a compensated state could be achieved, at least at the pituitary level. In the proposita, treatment of hyperthyrotropinemia is clearly mandatory because of potential complications inflicted by TSH-stimulated growth of the lingual tissue. To our knowledge, this represents the first report of congenital hypothyroidism secondary to thyroid dysgenesis complicated by coincidental RTH.

Loss-of-function mutations in the thyrotropin receptor gene as a major determinant of hyperthyrotropinemia in a consanguineous community.

CONTEXT: Resistance to TSH (RTSH) is a condition of impaired responsiveness of the thyroid gland to TSH, characterized by elevated serum TSH, low or normal thyroid hormone levels, and hypoplastic or normal-sized thyroid gland. OBJECTIVES: The aim of the study was to evaluate the clinical course and the genotype-phenotype relationship of RTSH caused by two different TSH receptor (TSHR) gene mutations in a consanguineous population. PATIENTS AND METHODS: We conducted a clinical and genetic investigation of 46 members of an extended family and 163 individuals living in the same town. In vitro functional studies of the mutant TSHRs were also performed. RESULTS: Two TSHR gene mutations (P68S and L653V) were identified in 33 subjects occurring as homozygous L653V (five subjects), heterozygous L653V (20 subjects), heterozygous P68S (four subjects), and compound heterozygous L653V/P68S (four subjects). With the exception of one individual with concomitant autoimmune thyroid disease, all homozygotes and compound heterozygotes presented with compensated RTSH (high TSH with free T(4) and T(3) in the normal range). Only nine of 24 heterozygotes had mild hyperthyrotropinemia. The L653V mutation resulted in a higher serum TSH concentration and showed a more severe in vitro abnormality than P68S. Haplotype analysis predicted a founder of the L653V six to seven generations earlier, whereas the P68S is older. Cross-sectional and prospective longitudinal studies indicate that TSH and T(4) concentrations remain stable over time. CONCLUSIONS: High frequency hyperthyrotropinemia in an Israeli Arab-Muslim consanguineous community is attributed to two inactivating TSHR gene mutations. Concordant genotype-phenotype was demonstrated clinically and by in vitro functional analysis. Retrospective and prospective studies indicate that in the absence of concomitant autoimmune thyroid disease, elevated TSH levels reflect stable compensated RTSH.