Department of Medicine, Unidade de Enfermedades Tiroideas e Metabolicas, University of Santiago de Compostela, Santiago de Compostela, Spain.
Thyroid. 2011 Feb;21(2):103-9. doi: 10.1089/thy.2010.0187. Epub 2010 Dec 27.
Resistance to thyrotropin (TSH) causes congenital hypothyroidism (CH). TSH receptor (TSHR) and adenylate cyclase-stimulating G alpha protein subunit (GNAS) loss-of-function mutations cause TSH resistance. We describe a family with TSH resistance and CH bearing a combination of inactivating mutations in TSHR and GNAS genes. We describe studies to determine the molecular mechanisms involved in TSH resistance in this family.
DNA sequencing to identify TSHR and GNAS gene mutations was performed. In vitro effects of the mutations on cAMP production and TSH binding were investigated in COS7 cells. cAMP production was evaluated by transfecting a cAMP response element (CRE)-luciferase reporter with pSVL-TSHR and pSVK3-GNAS vectors. For binding studies, cells transfected with pSVL-TSHR vectors were incubated with iodine-125 bovine TSH ((125)IbTSH).
Family members with and without CH were heterozygous for the TSHR mutant p.E34K or the GNAS mutant c.750_751insA (=GNASMut). The propositus had CH and he was heterozygous for TSHR p.E34K; his mother, also heterozygous for TSHR p.E34K, did not have CH. The euthyroid propositus' wife was heterozygous for GNASMut. The propositus' two daughters had CH, one was heterozygous for GNASMut and the other a compound heterezygous for TSHR p.E34K and GNASMut. Albright's hereditary osteodystrophy phenotype was present in those with GNASMut mutation but only the daughters had pseudohypoparathyroidism type 1a. Cells transfected with TSHRE34K had lower TSH affinity and less CRE-luciferase response than cells transfected with TSHR wild-type (WT). Cells transfected with GNASMut did not stimulate CRE-luciferase activity, but when cells were transfected with GNASMut plus GNASWT, a similar response to GNASWT alone was observed. The combination of TSHRWT and GNASWT showed higher CRE-luciferase response than TSHRWT and TSHRE34K with either GNASWT or GNASWT plus GNASMut.
CH was caused by loss-of-function mutations in TSHR and/or GNAS. The absence of CH in the propositus' mother argues against a role for TSHR p.E34K being the only cause of CH. The minimal thyroidal phenotypic differences between the sisters with pseudohypoparathyroidism type 1a and TSH resistance, both heterozygous for GNAS c.750_751insA but only one bearing the TSHR p.E34K mutant, suggest that the main cause for CH was preferential expression of the mutated maternal GNAS allele in the thyroid gland.
促甲状腺激素（TSH）抵抗会导致先天性甲状腺功能减退症（CH）。促甲状腺激素受体（TSHR）和腺苷酸环化酶刺激 G 蛋白亚单位（GNAS）的失活功能突变会导致 TSH 抵抗。我们描述了一个患有 TSH 抵抗和 CH 的家族，该家族同时存在 TSHR 和 GNAS 基因的失活突变。我们描述了研究该家族 TSH 抵抗相关分子机制的实验。
对 TSHR 和 GNAS 基因突变进行 DNA 测序。在 COS7 细胞中研究突变对 cAMP 产生和 TSH 结合的体外影响。通过转染 cAMP 反应元件（CRE）-荧光素酶报告载体 pSVL-TSHR 和 pSVK3-GNAS 来评估 cAMP 产生。对于结合研究，用碘-125 牛 TSH（（125）IbTSH）孵育转染了 pSVL-TSHR 载体的细胞。
患有 CH 和不患有 CH 的家族成员均为 TSHR 突变 p.E34K 或 GNAS 突变 c.750_751insA（=GNASMut）的杂合子。先证者患有 CH，且为 TSHR p.E34K 的杂合子；他的母亲也是 TSHR p.E34K 的杂合子，没有 CH。甲状腺功能正常的先证者的妻子为 GNASMut 的杂合子。先证者的两个女儿患有 CH，一个为 GNASMut 的杂合子，另一个为 TSHR p.E34K 和 GNASMut 的复合杂合子。有 GNASMut 突变的人存在 Albright 遗传性骨营养不良表型，但只有女儿患有 1a 型假性甲状旁腺功能减退症。转染 TSHRE34K 的细胞与转染 TSHR 野生型（WT）的细胞相比，TSH 亲和力更低，CRE-荧光素酶反应更低。转染 GNASMut 的细胞不能刺激 CRE-荧光素酶活性，但当用 GNASMut 加上 GNASWT 转染细胞时，观察到与单独用 GNASWT 相似的反应。TSHRWT 和 GNASWT 的组合比 TSHRWT 和 TSHRE34K 与 GNASWT 或 GNASWT 加上 GNASMut 的组合具有更高的 CRE-荧光素酶反应。
CH 是由 TSHR 和/或 GNAS 的失活功能突变引起的。先证者的母亲没有 CH，这表明 TSHR p.E34K 不是导致 CH 的唯一原因。1a 型假性甲状旁腺功能减退症和 TSH 抵抗的姐妹之间甲状腺表型差异极小，她们均为 GNAS c.750_751insA 的杂合子，但只有一人携带 TSHR p.E34K 突变，这表明 CH 的主要原因是突变的母系 GNAS 等位基因在甲状腺中的优先表达。
