scholarly journals A Novel Homozygous Mutation of Thyroid Peroxidase Gene Abolishes a Disulfide Bond Leading to Congenital Hypothyroidism

2020 ◽  
Vol 2020 ◽  
pp. 1-8
Author(s):  
Fumiyoshi Yakou ◽  
Hirotsugu Suwanai ◽  
Takuya Ishikawa ◽  
Mariko Itou ◽  
Jumpei Shikuma ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Congenital Hypothyroidism ◽  
Disulfide Bond ◽  
Hormone Replacement ◽  
Hek293 Cells ◽  
Dimensional Structure ◽  
Thyroid Peroxidase ◽  
Homozygous Mutation ◽  
Wild Type ◽  
Cousin Marriage

Congenital hypothyroidism (CH) is the most prevalent congenital endocrine disorder and causes mental retardation. A male Japanese patient with first cousin marriage parents was diagnosed as CH at 10 months. He was born before introduction of mass screening for CH. With continuous thyroid hormone replacement therapy, normal thyroid hormone status was maintained until adulthood. Genetic screening of next-generation sequencing was performed at the age of 52 years, and we identified a new homozygous thyroid peroxidase (TPO) gene mutation (GRCh38.p13, chromosome 2 at position 1493997, c.1964 G>T, p.Cys655Phe). TPO is an important enzyme to produce thyroid hormone. As demonstrated by a homology analysis of TPO proteins among different species, cysteine 655 residue is highly conserved, suggesting an important role in maintaining TPO function and structure. An in silico study with three-dimensional structure of the novel mutation was performed and suggested that the mutation abolished disulfide bond between cysteines at positions 598 and 655. An in vitro functional analysis using HEK293 cells revealed that TPO activity of the mutant was significantly impaired compared with that of the wild type. Furthermore, study of immunohistochemistry showed that localization of TPO in cells did not differ between the wild type and the mutant. In conclusion, this single disulfide bond loss mutation of a new TPO homozygous mutation, p.Cys655Phe, reduced TPO activity and caused congenital hypothyroidism without affecting subcellular localization of TPO proteins.

The Disulfide Bond between Cys22 and Cys27 in the Protease Domain Modulate Clotting Activity of Coagulation Factor X

2019 ◽  
Vol 119 (06) ◽  
pp. 871-881 ◽  
Author(s):  
Fang Li ◽  
Changming Chen ◽  
Si-Ying Qu ◽  
Ming-Zhu Zhao ◽  
Xiaoling Xie ◽  
...  
Keyword(s):  
Disulfide Bond ◽  
Catalytic Domain ◽  
Coagulation Factor ◽  
Major Change ◽  
Coagulation Factors ◽  
Hek293 Cells ◽  
Factor X ◽  
Wild Type ◽  
Protease Domain ◽  
Dynamic Simulations

AbstractThe Cys22-Cys27 disulfide bond of factor X (FX) protease domain is not conserved among coagulation factors and its contribution to the physiological haemostasis and implication in the pathogenesis of haemostatic and thrombotic disorders remain to be elucidated. Mutation p.Cys27Ser was identified in a pedigree of congenital FX deficiency and fluorescence labelling study of transiently transfected HEK293 cells showed accumulation of FX p.Cys27Ser within cell, indicating incompetent secretion partially responsible for the FX deficiency. The clotting activity of FX p.Cys27Ser was decreased to about 90% of wild-type, while amidolytic and pro-thrombinase activities (kcat/Km) determined with recombinant FXa mutant were 1.33- and 4.77-fold lower. Molecular dynamic simulations revealed no major change in global structure between FXa p.Cys27Ser and wild-type FXa; however, without the Cys22-Cys27 disulfide bond, the insertion of newly formed N terminal of catalytic domain after the activation cleavage is hindered, perturbing the conformation transition from zymogen to enzyme. The crystal structure of FXa shows that this disulfide bond is solvent accessible, indicating that its stability might be subject to the oxidation/reduction balance. As demonstrated with FX p.Cys27Ser here, Cys22-Cys27 disulfide bond may modulate FX clotting activity, with reduced FX pertaining less pro-coagulant activity.

scholarly journals TSH Compensates Thyroid-Specific IGF-I Receptor Knockout and Causes Papillary Thyroid Hyperplasia

2011 ◽  
Vol 25 (11) ◽  
pp. 1867-1879 ◽  
Author(s):  
Kathrin Müller ◽  
Dagmar Führer ◽  
Jens Mittag ◽  
Nora Klöting ◽  
Matthias Blüher ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Thyroid Tissue ◽  
Strong Increase ◽  
Thyroid Peroxidase ◽  
Receptor Signaling ◽  
Papillary Thyroid ◽  
Wild Type ◽  
Igf I ◽  
Thyroid Hyperplasia

Abstract Although TSH stimulates all aspects of thyroid physiology IGF-I signaling through a tyrosine kinase-containing transmembrane receptor exhibits a permissive impact on TSH action. To better understand the importance of the IGF-I receptor in the thyroid in vivo, we inactivated the Igf1r with a Tg promoter-driven Cre-lox system in mice. We studied male and female mice with thyroidal wild-type, Igf1r+/−, and Igf1r−/− genotypes. Targeted Igf1r inactivation did transiently reduce thyroid hormone levels and significantly increased TSH levels in both heterozygous and homozygous mice without affecting thyroid weight. Histological analysis of thyroid tissue with Igf1r inactivation revealed hyperplasia and heterogeneous follicle structure. From 4 months of age, we detected papillary thyroid architecture in heterozygous and homozygous mice. We also noted increased body weight of male mice with a homozygous thyroidal null mutation in the Igf1r locus, compared with wild-type mice, respectively. A decrease of mRNA and protein for thyroid peroxidase and increased mRNA and protein for IGF-II receptor but no significant mRNA changes for the insulin receptor, the TSH receptor, and the sodium-iodide-symporter in both Igf1r+/− and Igf1r−/− mice were detected. Our results suggest that the strong increase of TSH benefits papillary thyroid hyperplasia and completely compensates the loss of IGF-I receptor signaling at the level of thyroid hormones without significant increase in thyroid weight. This could indicate that the IGF-I receptor signaling is less essential for thyroid hormone synthesis but maintains homeostasis and normal thyroid morphogenesis.

scholarly journals Screening and Functional Analysis of TPO Gene Mutations in a Cohort of Chinese Patients With Congenital Hypothyroidism

2021 ◽  
Vol 12 ◽  
Author(s):  
Huijjuan Wang ◽  
Wenxia Wang ◽  
Xi Chen ◽  
Hailong Shi ◽  
Yinmin Shi ◽  
...  
Keyword(s):  
Congenital Hypothyroidism ◽  
High Throughput Sequencing ◽  
Biological Function ◽  
Northwest China ◽  
Chinese Patients ◽  
Thyroid Peroxidase ◽  
Missense Mutations ◽  
Wild Type ◽  
Hormone Synthesis

BackgroundsAs a crucial enzyme in thyroid hormone synthesis, the genetic defective thyroid peroxidase (TPO) was one of the main genetic factors leading to congenital hypothyroidism (CH).MethodsMutations in the TPO gene were screened and identified in 219 patients with CH from northwest China by using high-throughput sequencing and bioinformatics analysis. The biological function of detected variants was studied by in vitro experiments and homology modeling.ResultsNineteen rare variants, including seven novel ones, were detected in 17 of 219 patients (7.8%). Most cases were detected with one single heterozygous variant, and only two patients were detected with multiple variants, i.e., compounds for (1) IVS7-1G>A, p.Ala443Val, and p.Arg769Trp and (2) p.Asn592Ser and p.Asn798Lys. The biological function of the four missense mutations (i.e., p.Ala443Val, p.Arg769Trp, p.Asn592Ser, and p.Asn798Lys) they carried were further studied. Experimental data showed that these four mutations did not affect the protein expression level of the TPO gene but remarkably reduced the peroxidase activity toward guaiacol oxidation, retaining 8–32% of activity of the wild-type protein. The comparison of the predicted 3-D structures of wild-type and mutant TPO proteins showed that these four amino acid substitutions changed the non-covalent interactions of studied residues that might alter the structure and function of the TPO protein.ConclusionThis study was the first to analyze the TPO mutation spectrum of patients with CH in northwest China. Our data indicated that the TPO mutation was not a common reason to cause CH in China. The functional data may help to clarify the structure-function relationship of the TPO protein and provide further evidence for the elucidation of the genetic etiology of CH.

scholarly journals Male congenital hypothyroid Pax8−/− mice are infertile despite adequate treatment with thyroid hormone

2007 ◽  
Vol 192 (1) ◽  
pp. 99-109 ◽  
Author(s):  
Joachim Wistuba ◽  
Jens Mittag ◽  
C Marc Luetjens ◽  
Trevor G Cooper ◽  
Ching-Hei Yeung ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Congenital Hypothyroidism ◽  
Clinical Symptoms ◽  
Hormone Replacement ◽  
Serum Testosterone ◽  
Fluid Secretion ◽  
Pituitary Hormones ◽  
Developmental Defect ◽  
Adequate Treatment ◽  
Efferent Ducts

Severe forms of congenital hypothyroidism lead to serious clinical symptoms if thyroid hormone replacement therapy is not instituted immediately after birth. In this study, Pax8−/− mice that are born without a thyroid gland were used as an animal model to study the consequences of congenital hypothyroidism. As expected, adequate treatment of these animals with thyroxine restored the general deficits of congenital hypothyroidism; however, Pax8-deficient male mice were infertile. We report here that in these mice, the efferent ducts and epididymides are either absent or the efferent ducts exhibit a reduced lumen and extensive connective tissue, which appears to impair testicular drainage and subsequently leads to complete absence of spermatozoa from the epididymis. The results suggest that, starting with the onset of pubertal testicular fluid secretion, a backpressure is created in the testis by the absence of efferent ducts or constriction of their tubule lumen when present. This subsequently leads to secondary disorganization of the seminiferous epithelium that increases with age, resulting in mixed atrophy of the testis in the adult. Serum testosterone levels as well as mRNA expression of anterior pituitary hormones are in the normal range, indicating that the observed infertility is not due to hormonal imbalance, but rather to a developmental defect of the efferent ducts. The demonstration of Pax8 expression in the epithelia of the epididymis and the efferent ducts suggests a direct morphogenic role of Pax8 in the development of these organs. It remains to be elucidated whether congenital hypothyroid male patients with mutations in the Pax8 gene are similarly affected.

Congenital Hypothyroidism, Dwarfism, and Hearing Impairment Caused by a Missense Mutation in the Mouse Dual Oxidase 2 Gene, Duox2

2007 ◽  
Vol 21 (7) ◽  
pp. 1593-1602 ◽  
Author(s):  
Kenneth R. Johnson ◽  
Coleen C. Marden ◽  
Patricia Ward-Bailey ◽  
Leona H. Gagnon ◽  
Roderick T. Bronson ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Congenital Hypothyroidism ◽  
Genetic Model ◽  
Mutant Mice ◽  
Tectorial Membrane ◽  
Thyroid Peroxidase ◽  
Hormone Synthesis ◽  
Thyroid Hormone Synthesis ◽  
Organ Systems ◽  
Dual Oxidase

Abstract Dual oxidases generate the hydrogen peroxide needed by thyroid peroxidase for the incorporation of iodine into thyroglobulin, an essential step in thyroid hormone synthesis. Mutations in the human dual oxidase 2 gene, DUOX2, have been shown to underlie several cases of congenital hypothyroidism. We report here the first mouse Duox2 mutation, which provides a new genetic model for studying the specific function of DUOX2 in the thyroid gland and in other organ systems where it is hypothesized to play a role. We mapped the new spontaneous mouse mutation to chromosome 2 and identified it as a T>G base pair change in exon 16 of Duox2. The mutation changes a highly conserved valine to glycine at amino acid position 674 (V674G) and was named “thyroid dyshormonogenesis” (symbol thyd) to signify a defect in thyroid hormone synthesis. Thyroid glands of mutant mice are goitrous and contain few normal follicles, and anterior pituitaries are dysplastic. Serum T4 in homozygotes is about one-tenth the level of controls and is accompanied by a more than 100-fold increase in TSH. The weight of adult mutant mice is approximately half that of littermate controls, and serum IGF-I is reduced. The cochleae of mutant mice exhibit abnormalities characteristic of hypothyroidism, including a delayed formation of the inner sulcus and tunnel of Corti and an abnormally thickened tectorial membrane. Hearing thresholds of adult mutant mice are on average 50–60 decibels (dB) above those of controls.

scholarly journals Molecular insights into the role of genetic determinants of congenital hypothyroidism

2021 ◽  
Vol 19 (3) ◽  
pp. e29
Author(s):  
Yedukondalu Kollati ◽  
Radha Rama Devi Akella ◽  
Shaik Mohammad Naushad ◽  
Rajesh K. Patel ◽  
G. Bhanuprakash Reddy ◽  
...  
Keyword(s):  
Thyroid Hormones ◽  
Congenital Hypothyroidism ◽  
In Silico ◽  
In Silico Analysis ◽  
Thyroid Stimulating Hormone ◽  
Binding Energies ◽  
Thyroid Peroxidase ◽  
Transcriptional Level ◽  
Missense Mutations ◽  
Wild Type

In our previous studies, we have demonstrated the association of certain variants of the thyroid-stimulating hormone receptor (TSHR), thyroid peroxidase (TPO), and thyroglobulin (TG) genes with congenital hypothyroidism. Herein, we explored the mechanistic basis for this association using different in silico tools. The mRNA 3'-untranslated region (3'-UTR) plays key roles in gene expression at the post-transcriptional level. In TSHR variants (rs2268477, rs7144481, and rs17630128), the binding affinity of microRNAs (miRs) (hsa-miR-154-5p, hsa-miR-376a-2-5p, hsa-miR-3935, hsa-miR-4280, and hsa-miR-6858-3p) to the 3'-UTR is disrupted, affecting post-transcriptional gene regulation. TPO and TG are the two key proteins necessary for the biosynthesis of thyroid hormones in the presence of iodide and H2O2. Reduced stability of these proteins leads to aberrant biosynthesis of thyroid hormones. Compared to the wild-type TPO protein, the p.S398T variant was found to exhibit less stability and significant rearrangements of intra-atomic bonds affecting the stoichiometry and substrate binding (binding energies, ΔG of wild-type vs. mutant: ‒15 vs. ‒13.8 kcal/mol; and dissociation constant, Kd of wild-type vs. mutant: 7.2E-12 vs. 7.0E-11 M). The missense mutations p.G653D and p.R1999W on the TG protein showed altered ΔG (0.24 kcal/mol and 0.79 kcal/mol, respectively). In conclusion, an in silico analysis of TSHR genetic variants in the 3'-UTR showed that they alter the binding affinities of different miRs. The TPO protein structure and mutant protein complex (p.S398T) are less stable, with potentially deleterious effects. A structural and energy analysis showed that TG mutations (p.G653D and p.R1999W) reduce the stability of the TG protein and affect its structure-functional relationship.

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