TSH Receptor Gene Research Articles

Mutation of a Gene for Thyroid Transcription Factor-1 (TITF1) in a Patient with Clinical Features of Resistance to Thyrotropin

Resistance to TSH (RTSH [MIM 275200]) is a heterogeneous condition defined by variable degree of insensitivity to biologically active TSH. While this condition is classically caused by loss-of-function mutations of the TSH receptor gene (TSHR), several patients have exhibited RTSH-like phenotype in the apparent absence of TSHR mutations, and some of them have mutations of PAX8 or GNAS1. We identified a Japanese boy with congenital hypothyroidism who suffered from recurrent lower respiratory infection during infancy and choreoathetosis at a later age. At 14 years of age, he was diagnosed as having RTSH, on the basis of compensated hypothyroidism (TSH, 30.2 mU/L; FT4, 1.2 ng/dl), disproportionate increments of thyroid hormones and TSH during a TRH test (DeltaFT3, 0.4 pg/ml; DeltaT3, 13 ng/dl; and DeltaTSH, 88.3 mU/L), and normal ultrasound thyroid image and radioactive iodine uptakes. Molecular analysis for TITF1 revealed a novel de novo heterozygous deletion/insertion mutation (c.470_479delinsGCG,) that is predicted to lose the entire homeodomain and the NK2-specific domain. We suggest that a heterozygous loss-of-function TITF1 mutation can also cause RTSH-compatible phenotype.

The Genetic Basis of Thyroid Autoimmunity

Autoimmune thyroid diseases (AITDs), including Graves' disease (GD) and Hashimoto's thyroiditis (HT), are prevalent autoimmune diseases, affecting up to 5% of the general population. AITDs arise due to interplay between environmental and genetic factors. In the past decade, significant progress has been made in our understanding of the genetic contribution to the etiology of AITDs. Excitingly, several AITD susceptibility genes have been identified and characterized. Some of these susceptibility genes are specific to either GD or HT, while others confer susceptibility to both conditions. The first AITD susceptibility gene locus identified was the Human-Leukocyte-Antigen DR (HLA-DR) gene locus. Subsequently, a quintet of non-HLA genes, including the cytotoxic T lymphocyte antigen (CTLA-4), CD40, protein tyrosine phosphatase-22 (PTPN22), thyroglobulin, and thyroid-stimulating hormone receptor (TSHR) gene, has been shown to contribute to the susceptibility to AITDs. Recently, the mechanisms by which these new AITD genes predispose to AITDs have been dissected. In this review, we overview and highlight the recent data on the genes predisposing to AITDs and the putative mechanisms by which they confer susceptibility to disease.

Multifocal hyperfunctioning thyroid carcinoma without metastases

Hyperthyroidism due to thyroid carcinoma is rare, and most cases are caused by hyperfunctioning metastatic thyroid carcinoma rather than primary carcinoma. Among primary hyperfunctioning thyroid carcinoma, multifocal thyroid carcinoma is exceedingly rare, with the only one case being reported in the literature. Here, we describe the case of a 62-year-old woman with multifocal functioning thyroid carcinoma. Technetium-99 m ( 99mTc) scintigraphic imaging showed four hot areas in the thyroid gland. Histopathological examination of all four nodules revealed papillary carcinoma, corresponding to hot areas in the 99mTc scintigram. DNA sequencing of the thyrotropin receptor ( TSH-R) gene from all nodules revealed no mutation, indicating that activation of TSH-R was unlikely in the pathophysiogenesis of hyperfunctioning thyroid carcinoma in the present case.

Effects of serum TSH and FT4 levels and the TSHR‐Asp727Glu polymorphism on bone: the Rotterdam Study

TSH and thyroid hormone may have independent effects on bone. In this study we investigated the association of TSH and free T4 (FT4) with different bone parameters in human subjects. TSH and FT4 are known to be associated with body mass index (BMI) and a higher BMI gives a higher bone mineral density (BMD). Thus, we aimed to determine whether the effects of TSH and FT4 on bone are mediated by BMI. As TSH exerts its biological effect through the TSH receptor (TSHR), the TSHR gene might be a candidate gene affecting bone mass. The TSHR-Asp727Glu polymorphism is associated with lower TSH levels. We therefore examined the association of this polymorphism with bone parameters. Genotypes were determined by Taqman assay in 4934 elderly Caucasian men and women of the Rotterdam Study, of whom BMD and bone geometry data were available. Serum thyroid parameters were available in a random set of 1327 subjects. Femoral neck BMD as well as narrow neck BMD and cortical thickness increased with serum TSH. However, FT4 was more strongly and negatively associated with bone parameters. Regression models showed BMI-dependent and -independent effects of both TSH and FT4 on bone. Carriers of the TSHR-Glu(727) allele had a 2.3% higher femoral neck BMD. In line with the effect of TSH on bone in mice, serum TSH shows a positive trend with BMD in human subjects, a finding that is strengthened by the association between the TSHR-Asp727Glu polymorphism and femoral neck BMD. However, serum FT4 has a much greater influence on bone than TSH.

Identification of TSH receptor mutations in three families with resistance to TSH

Genetic analysis of the TSH receptor gene in seven subjects with subclinical hypothyroidism (SH), in whom the diagnosis of autoimmune thyroid disease had been excluded by laboratory and instrumental techniques currently available. Three families where different members (2 children and 5 adults) affected by SH were studied. GENETIC ANALYSIS: Genomic DNA was extracted from peripheral lymphocytes and the entire coding sequence of the TSHr gene was sequenced. pSVL-TSHr construct harbouring a Q8fsX62 insertion was obtained by site-directed mutagenesis. COS-7 cells transfected with wild-type and mutant receptor were used for binding studies, flow cytometry, and cyclic AMP (cAMP) determination. A four base pair (bp) duplication in position 41 (41TGCAins), leading to a premature stop of translation at codon 62 (Q8fsX62), was found to be heterozygous in the proband, the father and the sister in Family 1. In Family 2 the proband and the sister were heterozygous for the mutation D410N. In Family 3 the proband and the father were heterozygous for the mutation P162A. After transfection in COS-7 cells, the mutant receptor Q8fsX62 displayed a low expression at the cell surface, and a reduced response to bovine TSH (bTSH) in terms of cAMP production. We identified TSH receptor mutations in seven members of three families with subclinical hypothyroidism.

The Comparative Effects of Gene Modulators on Thyroid-Specific Genes and Radioiodine Uptake

The aim of this study was to comparatively investigate the effects of 5-azacytidine-C (5-Aza), trichostatin-A (TSA), and all-trans retinoic acid (ATRA) on the mRNA expressions of the sodium and iodine (Na/I) symporter (NIS), thyroglobulin (Tg), thyroid peroxidase (TPO), and the thyroid-stimulating hormone receptor (TSH-R), as well as radioiodine (RAI) uptake in cancer (B-CPAP) and normal (Nthy-ori 3-1) thyroid cell lines. Cell lines were treated with 10 ng/mL of TSA, 5 microM of 5-AZA, and 1 microM of ATRA, according to the MTT (methyl-thiazol-tetrazolium) test results. Additionally, recombinant thyroid-stimulating hormone (rTSH) was also applied, with a selected dose of 100 ng/mL. Following the treatment, NIS, Tg, TPO, and TSH-R mRNA levels were detected by real-time-polymerase chain reaction (RT-PCR) and RAI uptakes were measured by using a well counter as counts/cell number. 5-Aza increased TSH-R mRNA expression in both of the cell lines and decreased TPO, NIS, and Tg mRNA levels in the cancer cell line. In the normal thyroid cell line, 5-AZA increased TPO mRNA levels by 2-fold and made no differences in NIS and Tg mRNA levels. TSA treatment repressed NIS and Tg mRNA levels and made no change on other thyroid-specific genes that were investigated in the cancer cell line. In the normal thyroid cell line, TSA increased TSH-R mRNA levels in 72 hours and created no important difference in the other genes. ATRA repressed the TSH-R mRNA levels in the normal thyroid cell line and increased the TPO and Tg mRNA levels slightly in both the cell lines. Furthermore, in short-term treatment, ATRA repressed the NIS gene expression slightly, but in the long term, this repression turned to basal levels. 5-Aza, TSA, and ATRA did not make any changes in RAI uptake in the cancer cell line, but rTSH increased RAI uptake significantly. In the normal thyroid cell line, TSA and ATRA decreased RAI uptake (to 1/10 and 1/2, respectively), but 5-Aza and rTSH increased RAI uptake significantly (2- and 4-fold, respectively). In our study, we showed an increase in TSH-R gene expression and radioiodine uptake with 5-Aza. Further in vitro and in vivo studies are needed to support our findings and the potential clinical use of this agent.

Susceptibility genes in Graves’ ophthalmopathy: searching for a needle in a haystack?

The variety of clinical presentations of eye changes in patients with Graves' disease suggests that complex interactions between genetic, environmental, endogenous and local factors influence the development/severity of Graves' ophthalmopathy (GO). At present, the role of genetic factors in the development of GO remains unknown. Based on small case-control association studies with candidate genes, several susceptibility loci in GO have been proposed. These are human leucocyte antigen (HLA, 6p21.3), cytotoxic T-lymphocyte antigen-4 (CTLA-4, 2q33), tumour necrosis factor (TNF, 6p21.3), interferon-gamma (IFN-gamma, 12q14), intercellular adhesion molecule-1 (ICAM-1, 19p13), and thyroid stimulating hormone receptor gene (TSH-R, 14q31). Unfortunately, these results were either not confirmed or require replication in larger studies. There are many reasons for the lack of reproducibility of association studies in GO, including poor characterization of the studied groups and small sample sizes, which may result in both false positive and negative results. Thus, the genetic background of GO remains to be elucidated in future research. However, the possibility that GO may be a genetically heterogeneous disorder, or that the development of GO may be predominantly influenced by environmental factors such as cigarette smoking, can not be disregarded.

The impact of a TSH receptor gene polymorphism on thyroid‐related phenotypes in a healthy Danish twin population

The Asp727Glu polymorphism in the TSH receptor (TSHR) gene is associated with serum TSH levels. However, the proportion of genetic variation accounted for by this polymorphism is unknown. In this study, we (1) examined the association of the Asp727Glu polymorphism with thyroid size, serum levels of TSH, thyroid hormones, and thyroid antibodies in 1241 healthy Danish twin individuals and (2) assessed the contribution of the polymorphism to the trait variation and the genetic variance. The effect of the genotype on the traits (mean +/- SD) was established; associations between the TSHR-Asp727Glu polymorphism and measures of thyroid homeostasis were assessed and the effect of the polymorphism on the trait's phenotypic variability was quantified by incorporating the genotype information in structural equation modelling. The genotype distribution was Asp/Asp 84.9%; Asp/Glu 14.5% and Glu/Glu 0.6%. Carriers of the TSHR-Glu727 allele had lower TSH levels (noncarriers vs. carriers: 1.78 +/- 0.93 vs. 1.60 +/- 0.84 mU/l, P = 0.04). Regression analysis showed an association between the TSHR-Asp727Glu polymorphism and serum TSH (P = 0.007). The polymorphism accounted for 0.91% of the total phenotypic variance in serum TSH levels. Including the genotype in quantitative genetic modelling improved the model fit (P = 0.001); however, the genetic influence on serum TSH not attributable to this specific genetic variant was only reduced from 68.2% to 67.8%. The polymorphism was not significantly associated with thyroid size, thyroid hormones or thyroid antibody levels. The TSHR-727Glu allele was associated with decreasing TSH levels; however, the contribution to the genetic variance was very small. No association was found with other thyroid-related measures.

The Comparative Effects of Gene Modulators on Thyroid-Specific Genes and Radioiodine Uptake

The aim of this study was to comparatively investigate the effects of 5-azacytidine-C (5-Aza), trichostatin-A (TSA), and all-trans retinoic acid (ATRA) on mRNA expressions of Na/I symporter (NIS), thyroglobulin (Tg), thyroid peroxidase (TPO), and thyroid stimulating hormone receptor (TSH-R), and radioiodine (RAI) uptake in cancer (B-CPAP) and normal (Nthy-ori 3-1) thyroid cell lines. Cell lines were treated with 10 ng/mL of TSA, 5 microM of 5-Aza, and 1 microM of ATRA, according to the MTT (methyl-thiazol-tetrazolium) test results. Additionally, recombinant thyroid stimulating hormone (rTSH) was also applied, with a selected dose of 100 ng/mL. Following the treatment, NIS, Tg, TPO, and TSH-R mRNA levels were detected by real-time-polymerase chain reaction (RT-PCR) and RAI uptakes were measured by using a well counter as the counts/cell number. 5-Aza increased TSH-R mRNA expression in both of the cell lines and decreased TPO, NIS, and Tg mRNA levels in the cancer cell line. In the normal thyroid cell line, 5-Aza increased TPO mRNA levels 2-fold and made no differences in NIS and Tg mRNA levels. TSA treatment repressed NIS and Tg mRNA levels, and made no differences on other thyroid specific genes investigated in the cancer cell line. In the normal thyroid cell line, TSA increased TSH-R mRNA levels in 72 hours and created no important differences in other genes. ATRA repressed the TSH-R mRNA levels in the normal thyroid cell line and increased the TPO and Tg mRNA levels slightly in both cell lines. Furthermore, in short-term treatment, ATRA repressed NIS gene expression slightly, but in the long term, this repression turned to basal levels. 5-Aza, TSA, and ATRA did not make any differences in RAI uptake in the cancer cell line, but rTSH increased RAI uptake significantly. In the normal thyroid cell line, TSA and ATRA decreased RAI uptake (to 1/10 and 1/2, respectively), but 5-Aza and rTSH increased RAI uptake significantly (2- and 4-fold, respectively). We have shown an increase in TSH-R gene expression and radioiodine uptake with 5-Aza. Further in vitro and in vivo studies are needed to support our findings and the potential clinical use of this agent.

Suppression of BRAF/MEK/MAP Kinase Pathway Restores Expression of Iodide-Metabolizing Genes in Thyroid Cells Expressing the V600E BRAF Mutant

The V600E BRAF mutant plays an important role in the pathogenesis of papillary thyroid cancer (PTC) and is associated with loss of expression of thyroid iodide-metabolizing genes. This study was done to investigate the restorability of expression of these genes by suppressing the BRAF/extracellular signal-regulated kinase kinase (MEK)/mitogen-activated protein (MAP) kinase pathway in V600E BRAF-harboring thyroid cells and to explore the mechanisms involved. We used inducible expression of V600E BRAF, small interfering RNA transfection, and MEK-specific inhibitor to alter the MAP kinase pathway activities and subsequently examined the changes in expression, promoter activities, and methylation status of thyroid genes. MEK inhibitor U0126 or cessation of V600E BRAF expression in PCCL3 cells restored expression of thyroid genes silenced by induced expression of V600E BRAF. U0126 also restored the expression of these genes in V600E BRAF-harboring PTC-derived NPA cells. Knockdown of BRAF by specific small interfering RNA restored expression of some of these genes in NPA cells. Luciferase reporter assay using thyroid-stimulating hormone receptor gene as a model showed that the promoter activity was modulated by the MAP kinase pathway. Promoter methylation in association with DNA methyltransferase expression played a role in gene silencing by MAP kinase pathway in NPA cells. We showed the restorability of expression of thyroid iodide-metabolizing genes silenced by V600E BRAF, and linked this process to gene methylation in PTC cells. The results provide clinical implications that therapeutic targeting at the BRAF/MEK/MAP kinase pathway may be a good approach in restoring thyroid gene expression for effective radioiodine therapy for BRAF mutation-harboring PTC.

Lack of association between thyrotropin receptor gene polymorphisms and subclinical hypothyroidism in children

Subclinical hypothyroidism is defined as a serum TSH level above the statistically set reference range, associated to normal free thyroid hormone concentrations. Genetic and environmental factors contribute to the inter- and intra-individual biological variations of TSH levels, sometimes leading to uncertainty of treatment in the clinical practice, especially when moderate elevations above the upper limit of the reference range are considered (5< TSH <10 mIU/l). In this view, the study of association between subclinical hypothyroidism and possible molecular effectors, such as polymorphisms in the TSH receptor (TSHR) gene, could be interesting. In this paper, we analyzed the TSHR gene polymorphisms in 103 hyperthyrotropinemic infants. A control group of 120 newborns of the same ethnic background was used to evaluate the frequencies of each polymorphism in the population. We found a statistically significant difference in the allelic frequency of the P52T polymorphism, being that the T variant was more represented in the control group (p=0.03). However, no significant results have been obtained in the analysis of the association between genotypes and serum TSH levels. In conclusion, we analyzed 7 polymorphic variants of TSHR gene in subclinical hypothyroidism. The only significant result refers to the allelic frequency of A in the P52T polymorphism, which is statistically reduced when compared with that of a control group.

A Novel Thyrotropin Receptor Germline Mutation (Asp617Tyr) Causing Hereditary Hyperthyroidism

Constitutively activating germline mutations of the thyrotropin receptor (TSHR) gene have been identified as a molecular cause of hereditary nonautoimmune hyperthyroidism. We describe here a Japanese kindred with two affected individuals who showed overt hyperthyroidism and mild goiter in the absence of TSHR antibodies. A novel heterozygous germline point mutation, identified in both individuals, resulted in an amino acid substitution of aspartic acid for tyrosine at codon 617 (Asp617Tyr) in the third intracellular loop of the TSHR. Screening of 7 additional family members led to the identification of the same mutation in 4 relatives: 1 had undergone thyroidectomy due to hyperthyroidism but 3 were asymptomatic with subclinical hyperthyroidism. In vitro functional studies of the Asp617Tyr TSHR demonstrated a constitutive activation of the cyclic adenosine monophosphate pathway, but not of the inositol phosphate cascade, with data similar to those of Asp619Gly, the first constitutively activating mutant TSHR identified. Treatment with inorganic iodine for 7 months successfully relieved all symptoms of hyperthyroidism in both patients.

Subclinical hyperthyroidism due to a thyrotropin receptor (TSHR) gene mutation (S505R)

To identify the molecular defect by which non-autoimmune subclinical hyperthyroidism was caused in a 6-mo-old infant who presented with weight loss. Congenital non-autoimmune hyperthyroidism is caused by activating germline mutations in the thyrotropin receptor (TSHR) gene. Therefore, the TSHR gene was sequenced directly from the patient's genomic DNA. Molecular analysis revealed a heterozygous point mutation (S505R) in the TSHR gene as the underlying defect. A constitutively activating mutation in the TSHR gene has to be considered not only in patients with severe congenital non-autoimmune hyperthyroidism, but also in children with subclinical non-autoimmune hyperthyroidism.

Clinical significance of heterozygous carriers associated with compensated hypothyroidism in R450H, a common inactivating mutation of the thyrotropin receptor gene in Japanese

Loss-of-function mutations in the thyrotropin receptor (TSHR) gene were described as a syndrome characterized by thyroid hyposensivity to biologically active TSH, ranging from euthyroid to severe hypothyroidism. In Japanese, a common mutation in the TSHR gene is R450H, which demonstrated moderately impaired receptor function. We studied six subjects of Japanese origin whose major abnormality was persistent hyperthyrotropinemia by genetic sequence analysis of the TSHR gene. Three subjects were homozygous for the R450H mutation, whereas the three remaining subjects were single heterozygous. Homozygous subjects displayed mild hypothyroidism confirmed by moderately elevated basal TSH levels and excessive TSH response to TRH administration. Heterozygous subjects also demonstrated fully or partially compensated hypothyroidism, but less severe than that of homozygous subjects. More frequent involvement of the R450H mutation in the TSHR gene in Japanese was identified. In addition, a good correlation between phenotype and genotype was demonstrated in respect to biochemical analysis and drug dosage. Our observations showed clinical significance of heterozygosity associated with compensated hypothyroidism in spite of only mildly impaired receptor function.

A Novel Mutation in the Thyrotropin (Thyroid-Stimulating Hormone) Receptor Gene in a Case of Congenital Hypothyroidism

Congenital hypothyroidism (CH) occurs approximately with a frequency of 1 in 3000-4000 births, being a disease caused by defects in thyroid hormone synthesis associated either with goiter presence or with agenesis or ectopy of the thyroid gland. A study of some familial cases has allowed identification of mutations in several known genes, including that encode the thyroid-stimulating hormone receptor (TSHR). We report a familial case of CH that transmitted as a recessive trait and caused by a novel homozygous nonsense mutation in TSHR with an initial diagnosis of thyroid agenesis hypoplasia. Genomic DNA was obtained from two siblings and their parents; TSHR was amplified using pairs of overlapping exonic primers; and polymerase chain reaction products were automatically sequenced. The propositus was homozygous (genotype: M/M) for a novel C to G transversion (1431C>G), producing a nonsense mutation, Y444X, in the first intracellular loop of TSHR, rendering a truncated receptor. Thus, the observed unresponsiveness to TSHR may be due to absent insertion of the truncated receptor into the cell membrane (if it gets translated at all) or the truncation may lead to nonsense-mediated mRNA degradation (its unresponsive to TSH). Both parents were heterozygous (wWt/M) and unrelated, as known from family history. The other daughter was homozygous for both wild-type alleles (wWt/wWt).

Hypothyroïdie congénitale

Congenital hypothyroidism is the principle cause of preventable mental retardation, with a prevalence of 1 in 3,500 neonates. The disorder may be permanent or transitory. Permanent congenital hypothyroidism is caused principally by thyroid dysgenesis. In industrialized countries, mass screening allows the disorder to be diagnosed at birth. The severity is variable but is generally more pronounced in females. The majority of studies point to a genetic origin for the disease and no consistent evidence has been found to suggest a major role for environmental factors. The genetic factors have already been identified and involve several elements (mutations in the TTF-1, TTF-2, PAX8 and TSH receptor genes). The etiological diagnosis is based on scintigraphy, ultrasound and the level of circulating thyroglobulin. At present, treatment is administered at an adapted dose during the first two weeks of life and should allow the child to reach its full intellectual potential. However, minor anomalies have been reported in some treated children, suggesting that this treatment cannot compensate for a certain degree of foetal hypothyroidism.

A Family of 3 Generations With Mild Nonautoimmune Hyperthyroidism Resulting From a Novel Activating Mutation of the Thyrotropin Receptor Gene (M626I)

A Family of 3 Generations With Mild Nonautoimmune Hyperthyroidism Resulting From a Novel Activating Mutation of the Thyrotropin Receptor Gene (M626I)

Identification and Functional Analysis of Novel Inactivating Thyrotropin Receptor Mutations in Patients with Thyrotropin Resistance

We identified and analyzed novel thyrotropin (TSH) receptor mutations in three Japanese families with resistance to TSH. The TSH receptor gene was sequenced and the mutations were determined. The mutant TSH receptors were transfected into COS-7 cells, and their functions were analyzed. The patients were compound-heterozygotes for the R450H mutation and novel mutations in the TSH receptor gene. The first patient was a compound-heterozygote for R450H and V473I. The second sibling possessed R450H and R519C. The third sibling had R450H and R519G. The R450H mutant exhibited moderately impaired receptor functions and a moderately decreased cell surface expression in agreement with previous results. The V473I mutant exhibited an almost normal TSH binding, a slightly decreased cyclic adenosine monophosphate (cAMP) response, a moderately decreased inositolphosphate (IP) response, and an almost normal cell surface expression. TSH binding and TSH stimulation of cAMP and IPs were markedly decreased in the R519C and R519G mutants. Cell surface expression was decreased in the R519C mutant and negligible in the R519G mutant. All of these mutants showed normal intracellular synthesis of TSH receptors. These novel inactivating mutations contribute to understanding of the structure-function relationship of the TSH receptor. To date, all of the patients with TSH resistance resulting from TSH receptor mutations identified in Japan possessed the R450H mutation at least in one allele. These observations suggest that the R450H mutation is a commonly observed TSH receptor mutation in patients with TSH resistance in Japan.

Case History: A Novel Activating Mutation in Transmembrane Helix 6 of the Thyrotropin Receptor as Cause of Hereditary Nonautoimmune Hyperthyroidism

Constitutively-activating germline mutations of the thyrotropin receptor (TSHR) gene are very rare and are considered the cause of hereditary nonautoimmune hyperthyroidism. We describe four affected individuals from a Caucasian family: a mother and her three children, and an unaffected father. The mother and her first two children presented in a similar manner: lifelong histories of heat intolerance, hyperactivity, fast heart rate, reduced energy, increased appetite, and scrawny build. They all developed goiter in childhood and showed a suppressed TSH and elevated thyroxine (T(4)). The last child, a 12-year-old female, presented with no clinical symptoms or palpable neck mass, but with a suppressed TSH, elevated T(4) and thyromegaly detected by ultrasound. Mutation analysis of the TSHR gene in all family members revealed a novel heterozygous germline mutation resulting in the substitution of phenylalanine (TTC) by serine (TCC) at codon 631 in transmembrane helix 6 in the mother and all three children. Functional characterization of this germline mutation showed constitutive activation of the G(s)-mediated cyclic adenosine monophosphate (cAMP) pathway, which controls thyroid hormone production and thyroid growth. Molecular characterization of F631S demonstrates that this activating mutation plays a key role in the development of hereditary hyperthyroidism in this family although the timing of onset of clinical manifestations in the subjects may depend on other, as yet unidentified, factors.

Case Reports: Follicular Carcinoma Presenting as Autonomous Functioning Thyroid Nodule and Containing an Activating Mutation of theTSH Receptor(T620I) and a Mutation of TheKi-RAS(G12C) Genes

Most autonomous functioning thyroid nodules (AFTN) are benign thyroid follicular neoplasms. There are rare reports of malignant hot nodules, in which activating mutations of the TSH receptor (TSHR) were found. We report a case of follicular carcinoma presenting as an AFTN causing subclinical hyperthyroidism in a 64-year-old woman who had a 6-cm hot nodule in the left thyroid lobe. Genomic DNA was extracted from paraffin-embedded tissues from the tumor and extratumoral thyroid tissue. Sequence analyses revealed point mutations in two different genes: the normal ACC sequence at codon 620 of the TSHR gene was replaced by ATC, changing the threonine by isoleucine (T620I); and the wild-type GGT at codon 12 of Ki-RAS mutated to TGT, replacing glycine by cysteine (G12C). In transfection experiments the T620I mutant showed constitutive activity in terms of cyclic adenosine monophosphate (cAMP) production when permanently transfected in 3T3 cells. Here, we describe for the first time an activating mutation in 3codon 620 of the TSHR. In addition, the cancerous AFTN also contained a G12C Ki-RAS mutation. We hypothesize that the combination of these two mutations might have played an important role in both the hyperfunction of the tumor and the carcinogenetic process.