Thyroglobulin Synthesis Research Articles

A variant of adenomatous goiter with characteristic histology and possible hereditary thyroglobulin abnormality.

A variant type of adenomatous goiter was identified in 24 of 2160 patients with adenomatous goiter who underwent thyroidectomy. The characteristics of the thyroid gland in these 24 patients included large goiter, small follicles, scant colloid, and columnar follicular cells containing yellow-green granules on hematoxylin-eosin staining. The thyroid gland was slightly orange-red, and electron microscopic examination showed abundant lysosomes with colloid droplets. When comparing the features of this group with those of 24 patients with common adenomatous goiter, the incidence of familial predisposition to thyroid diseases in the former group was higher. The age at the time of detection of goiter was lower, i.e. 17 +/- 15 vs. 44 +/- 17 yr (P < 0.001, variant type vs. common type), the serum total T4 concentrations were lower (84 +/- 21 vs. 103 +/- 18 nmol/L; P < 0.01), and the serum TSH concentrations were higher (2.4 +/- 2.1 vs. 1.0 +/- 0.9 mU/L; P < 0.01). Thyroid radioiodine uptake was remarkably increased (49 +/- 22 vs. 16 +/- 9%; P < 0.001), and lower levels of serum thyroglobulin were noted (33 +/- 51 vs. 484 +/- 603 micrograms/L; P < 0.01). The thyroglobulin content was low in the thyroid gland studied. The data suggest that the etiology of this variant type of goiter is a hereditary abnormality in thyroglobulin synthesis, and this type of goiter may be distinguished from common adenomatous goiter by the characteristic morphology of the thyroid gland in addition to clinical findings.

Co-expression of vimentin and 19S-thyroglobulin in follicular cells located in the C-cell complex of dog thyroid gland.

The localization of vimentin, an intermediate filament protein, in the dog thyroid cells was examined by immunohistochemistry with a monoclonal antibody against vimentin and polyclonal anti-19S-thyroglobulin and anti-calcitonin antibodies. In the dog thyroid gland, C-cells are usually distributed as large cell groups in an interfollicular location. In addition, C-cell complexes that are special C-cell clusters associated with other epithelial elements and cysts are frequently located around the parathyroid gland IV. The C-cell complexes are remnants of embryonic ultimobranchial bodies and retain fetal characteristics for long periods. Follicular cells in various stages of differentiation are observed in the C-cell complexes and large C-cell groups of postnatal dogs: (a) cell masses not yet organized into follicles, (b) primitive and small follicles storing colloid, and (c) small follicles without colloid storage. The immature follicular cells in the C-cell complexes and large C-cell groups express immunoreactivity for vimentin in addition to 19S-thyroglobulin. When the follicular cells stored considerable amounts of colloid in the follicular lumina, vimentin immunoreactivity disappeared from them. Typical thyroid follicles showed no immunoreactivity for vimentin. In dog fetuses in which follicular cells were arranged in cell clusters or were vigorously forming follicles, no immunoreactivity for vimentin was observed in the thyroid parenchyma. These results indicate that vimentin is specifically expressed in the immature follicular cells derived from the ultimobranchial anlage. The vimentin filaments may participate in increased cellular activities in the cells, i.e., thyroglobulin synthesis and folliculogenesis.

A 138-nucleotide deletion in the thyroglobulin ribonucleic acid messenger in a congenital goiter with defective thyroglobulin synthesis.

Two siblings (HSN and AcSN) with congenital goitrous hypothyroidism were investigated in terms of clinical, biochemical, and molecular biology. Diagnosis of defective thyroglobulin (Tg) was based on findings of low serum T4, low normal or normal serum T3, a negative percholate discharge test, and the virtual absence of the serum Tg response to challenge by bovine TSH. Only minute amounts of Tg-related antigens were detected by RIA in the goitrous tissue (HSN, 0.82 mg/g, compared to 70-90 mg/g in normal thyroid tissue), as confirmed by sodium dodecyl sulfate-agarose gel electrophoresis that indicated the virtual absence of Tg. The Tg messenger ribonucleic acids (mRNAs) from controls and HSN thyroid tissue were first reverse transcribed and then divided into several portions from positions 57-8448; the resulting complementary DNAs were, in turn, amplified by reverse polymerase chain reaction. The amplification of nucleotides 5165-6048 from control thyroid tissue Tg mRNA showed a fragment of 884 base pairs (bp). In contrast, the fragment present in the HSN was +/- 750 bp and lacked the normal fragment. The sequencing of the smaller fragment revealed that 138 bp were missing between positions 5590-5727 of the HSN Tg mRNA. This deletion does not affect the reading frame of the resulting mRNA and is potentially fully translatable into a Tg polypeptide chain that is shorter by 46 residues. A cysteine residue is maintained by the junction between the proximal T from leucine 1831 and the distal GT from cysteine 1877. DNA genomic polymerase chain reaction amplification excludes a deletion in the Tg gene and indicates that the deleted 138-nucleotide sequences lie in the same exon. The functional consequences of the deletion are not entirely clear, but it is conceivable that the excision of this segment of the Tg molecule could affect the protein structure, resulting in its premature degradation, very low colloid storage, and diminished thyroid hormone production rate.

A 138-nucleotide deletion in the thyroglobulin ribonucleic acid messenger in a congenital goiter with defective thyroglobulin synthesis

A 138-nucleotide deletion in the thyroglobulin ribonucleic acid messenger in a congenital goiter with defective thyroglobulin synthesis

Clinical and Morphologic Findings of Familial Goiter in Bongo Antelope (Tragelaphus eurycerus)

Inherited defects of thyroglobulin synthesis resulting in congenital goiter are well described in certain breeds of domestic ungulates and in human beings. Goiter associated with synthesis of an abnormal thyroglobulin and the presence of thyroidal albumin was identified in five closely related bongo antelopes (Tragelaphus eurycerus). The goiter had an adult onset, and the affected bongos appeared to remain euthyroid with normal serum T3 and T4 values, normal serum cholesterol concentrations, and nonelevated concentrations of circulating thyroid stimulating hormone (TSH). Goitrous bongos had significant reproductive difficulties, including reduced cyclic activity and prolonged gestations, but were otherwise normal. Over the course of the disease, the thyroid glands greatly enlarged (up to 10 x 20 cm) and became polycystic. Microscopically, there was an admixture of giant colloid-filled follicles and follicles of normal size lined with variable follicular epithelium ranging from squamoid to mildly to moderately hyperplastic. The pathogenesis of goiter in the bongo may reflect a mixture of genetic predisposition coupled with environmental factors, including a period of exposure to a goitrogen.

Hormonal regulation of some steps of thyroglobulin synthesis and secretion in bicameral cell culture

Porcine thyroid cells were cultured for 15 days on porous bottom chambers with or without different mixtures of hormones added to serum-free basal medium. Assays with 10% serum were also performed for comparison with previously published results. The effects of the hormones, particularly insulin, TSH and hydrocortisone, were studied on total RNA content, thyroglobulin mRNA level, the amount of thyroglobulin secreted into the apical medium and on glycosylation. Insulin and TSH similarly increased the total RNA content, and their effects were additive. Thyroglobulin mRNA content was increased twofold by insulin and threefold by TSH. When they were added simultaneously, the maximal level of thyroglobulin mRNA was reached, showing that TSH and insulin effects on thyroglobulin gene expression were additive. Hydrocortisone alone did not modify total RNA or thyroglobulin mRNA content but the hormone amplified total RNA when insulin and TSH were present together. The basal level of thyroglobulin secreted into the apical medium was increased threefold by insulin and fourfold by TSH. The effects of these two hormones added together appeared to be additive. Hydrocortisone had no effect alone or even when combined with insulin or TSH. However, when the three hormones were added together, the hormonal response was amplified. TSH effect and insulin effect on the incorporation of 3H-mannose into thyroglobulin as well as on the anionic residue content of the molecule were additive.

Congenital euthyroid goitre with impaired thyroglobulin transport

A case of congenital goitre with defective thyroglobulin (Tg) synthesis was studied from the clinical, biochemical and morphological perspectives. The patient, 5.5-year-old boy, who was clinically euthyroid, showed a positive perchlorate discharge test (37.2%). However, the iodination system seemed to be normal since radioiodine uptake into the thyroid was very high, and inspection of the H2O2-generating system using thyroid slices and an assay for peroxidase activity in microsomes showed no abnormalities. On the other hand, virtually no Tg was detected in the serum, and the amount of Tg in thyroid tissue, estimated with gel electrophoresis, was below 10% of the normal value, the quality of Tg being unchanged. Morphological observations demonstrated the presence of Tg in the markedly distended rough endoplasmic reticulum of the cytoplasm of follicular cells and a lack of Tg in the colloid of the follicular lumen. These results suggest that the thyroid is defective in Tg synthesis, probably associated with impaired transport of Tg from the cells to the lumen.

Hypothyroidism

Thyroid Development THYROID HORMONE SYNTHESIS AND CONTROL Thyroid hormones (TH) are critical for normal growth, development, and metabolism during infancy and childhood. They are synthesized by the thyroid gland follicular cells, the function of which is to concentrate iodide from the blood and return it to peripheral tissues in a hormonally active form. The thyroid cell iodide-concentrating mechanism, often referred to as the iodide pump, confers on the gland its ability to concentrate iodide to many times its level in plasma. The steps involved in synthesis and release of TH include: 1) iodide trapping by the thyroid gland; 2) synthesis of thyroglobulin; 3) organification of trapped iodide as iodotyrosines (monoiodotyrosine, MIT, and diiodotyrosine, DIT); 4) coupling of the iodotyrosines to form the iodothyronines thyroxine (T4) and triiodothyronine (T3) and storage in follicular colloid; 5) endocytosis of colloid droplets and hydrolysis of thyroglobulin to release MIT, DIT, T4, and T3; and 6) deiodination of MIT and DIT with intrathyroidal recycling of the iodine. The concentration of iodide in plasma under normal circumstances is less than 0.2 µg/dL. The iodide is removed from plasma almost entirely by the kidney and the thyroid gland. Transport of iodide across the thyroid cell membrane is the first and ratelimiting step in TH biosynthesis.

Hypothyroidism

Thyroid DevelopmentTHYROID HORMONE SYNTHESIS AND CONTROLThyroid hormones (TH) are critical for normal growth, development, and metabolism during infancy and childhood. They are synthesized by the thyroid gland follicular cells, the function of which is to concentrate iodide from the blood and return it to peripheral tissues in a hormonally active form. The thyroid cell iodide-concentrating mechanism, often referred to as the iodide pump, confers on the gland its ability to concentrate iodide to many times its level in plasma. The steps involved in synthesis and release of TH include: 1) iodide trapping by the thyroid gland; 2) synthesis of thyroglobulin; 3) organification of trapped iodide as iodotyrosines (monoiodotyrosine, MIT, and diiodotyrosine, DIT); 4) coupling of the iodotyrosines to form the iodothyronines thyroxine (T4) and triiodothyronine (T3) and storage in follicular colloid; 5) endocytosis of colloid droplets and hydrolysis of thyroglobulin to release MIT, DIT, T4, and T3; and 6) deiodination of MIT and DIT with intrathyroidal recycling of the iodine.The concentration of iodide in plasma under normal circumstances is less than 0.2 µg/dL. The iodide is removed from plasma almost entirely by the kidney and the thyroid gland. Transport of iodide across the thyroid cell membrane is the first and ratelimiting step in TH biosynthesis.

Dyshormonogenetic goiter: A morphological and immunohistochemical study.

A retrospective morphological and immunohistochemical study of 21 cases of dyshormonogenetic goiter was carried out correlating patterns of hyperplasia and the atypias of the glandular tissues with specific defects in hormonal synthesis, including (1) thyroglobulin synthesis defect (Group I, n = 8); (2) defective organification of iodide (Group II, n = 11); and (3) iodide transport defect (Group III, n = 2). Microfollicular, trabecular, papillary, and oxyphilic cell patterns were more frequent in Group II compared with Group I (Group III was excluded because of the small number of cases). The combined microfollicular and trabecular patterns were more frequently seen in patients in Group II. Two cases of thyroglobulin synthesis defect demonstrated certain morphological specificity characterized by an alveolar pattern. Atypias were more frequent and severe in patients in Group II relative to patients in Group I, but features of malignancy were not found in any patients. Immunohistochemical study using thyroglobulin antiserum demonstrated correlation between morphology and positivity of follicular cells. Scarce C cells were verified in these cases by immunohistochemistry. Using two-paired samples, respectively, of 21 endemic and 21 dyshormonogenetic goiters, we distinguished 85.7% of the cases examined, presented in a double-blind fashion. Scarcity of colloid and prominent cellular atypia were highly suggestive of dyshormonogenetic goiter. Considering the relative rarity of dyshormonogenetic goiter, our studies point out the most common patterns of hyperplasia and atypias in this pathology to avoid misdiagnosis, principally when considering the possibility of malignancy.

A nonsense mutation causes human hereditary congenital goiter with preferential production of a 171-nucleotide-deleted thyroglobulin ribonucleic acid messenger.

Defective or impaired thyroglobulin (Tg) synthesis usually results in congenital goitrous hypothyroidism, virtual absence of Tg in thyroid tissue, and the presence of an elevated concentration of iodoalbumin. The final result of these abnormalities is a decreased rate of T3 and T4 synthesis. We have previously reported two siblings with this syndrome that was attributable to decreased levels of thyroid tissue Tg mRNA, resulting in decreased translation of a fully mature Tg. Further molecular studies in this family are the subject of this report. The Tg mRNA from normal and goitrous thyroid tissue was first reverse transcribed and divided into five overlapping portions from positions 57-8448, and the resulting cDNAs were amplified by polymerase chain reaction and analyzed by agarose gel electrophoresis. The amplification of nucleotides (nt) 4502-5184 from control thyroid tissue Tg mRNA showed a predominant fragment of 683 basepairs (bp) and a minor fragment of 512 bp. This latter fragment contained a 171-nt deletion that mapped between positions 4567 and 4737 of the Tg mRNA. In contrast, the fragment predominantly present in the congenital goiter was 512 bp. The sequencing of the 683-bp fragment revealed that the responsible mutation is a cytosine to thymine transition, creating a stop codon at position 1510. This results in loss of a TaqI restriction site. The point mutation is, thus, removed from a portion of the transcripts by the preferential accumulation in the goiter of a 171-nt-deleted Tg mRNA. The reading frame is maintained and is potentially fully translatable into a Tg polypeptide chain shorter by 57 residues. The presence of the deleted Tg mRNA in normal thyroid tissue, albeit at a low level, strongly suggests that the deleted mRNA sequence corresponds to a complete exon. Our studies suggest that the shorter, alternatively spliced Tg mRNA predominates in the goitrous tissue and probably has a shorter half-life. This would explain the tissue's low Tg mRNA levels, previously reported. Moreover, translation of the mutated transcript would generate a severely truncated Tg polypeptide with limited ability to generate thyroid hormone, resulting in congenital goitrous hypothyroidism.

Defective thyroglobulin synthesis and secretion causing goiter and hypothyroidism [published erratum appears in Endocr Rev 1994 Aug;15(4):438

Defective thyroglobulin synthesis and secretion causing goiter and hypothyroidism [published erratum appears in Endocr Rev 1994 Aug;15(4):438

Defective Thyroglobulin Synthesis and Secretion Causing Goiter and Hypothyroidism *

The integrity of the Tg structure as a protein is essential for adequate synthesis of thyroid hormone. Also a large supply of iodine and of thyroid hormone is stored into the Tg molecule and available for secretion on demand. Mutations in Tg gene or hyposialylated Tg due to a defective sialyltransferase activity would cause a structurally defective protein and severely impair the functional ability of Tg. In this review we attempt to cover the abnormalities in the synthesis of Tg described in both animals and man. Hereditary congenital goiter with or without hypothyroidism is the phenotypic major clinical finding in these species. Affected animals include sheep, bovine cattle, bongo antelope, goats, and mice. As in man the inheritance mode is autosomal recessive. In most animal studies structurally abnormal Tg is present. The molecular basis for the defective Tg synthesis was attributable to nonsense mutation in exon 9 (Afrikander cattle) and in exon 8 (Dutch goats). In man the Tg defective synthesis has been reported in 89 subjects and frequently more than one sibling is affected in a given generation. Characteristically these patients exhibit hereditary congenital goiter with relatively low Tg levels that do not increase after stimulation with bovine TSH. High PBI concentrations with low serum T4 values indicate the serum presence of iodinated proteins (mainly iodoalbumin). Also iodinated peptides are frequently excreted into the urine. Tissue studies confirm that there is an absent Tg peak at gel filtration, and virtually no immunoassayable Tg is present in the tissue extracts. The molecular basis of these defects have been recently reported in a patient and includes low tissue Tg mRNA probably due to premature degradation of a defective Tg mRNA. The responsible mutation is a cytosine to thymine transition creating a stop codon at position 1510. The point mutation is removed by the preferential accumulation of a 171 nt deleted Tg mRNA. In another subject molecular studies revealed that exon 4 was missing from the major Tg transcript due to a cytosine to guanine transversion at position minus 3 in the acceptor splice site of intron 3. It is anticipated that other mutations responsible for these defects will be identifiable in the near future.

Molecular basis of the thyroglobulin synthesis defect in Dutch goats

A thyroglobulin (Tg) synthesis defect in Dutch goats causes congenital goiter and hypothyroidism. The disease is inherited in an autosomal recessive way and is linked to restriction fragment length polymorphisms (RFLPs) in the Tg gene. Previous studies showed that Tg mRNA isolated from the goiters was of normal size (8.4 kilobases). Translation of high mol wt polysomal Tg mRNA isolated from goiter in a cell-free rabbit reticulocyte lysate resulted in a single 35,000 mol wt Tg polypeptide. Tg antigens analyzed in T4-arrested goiters were glycosylated and had mol wt of 40,000 and 32,000. The aim of this study was to identify the molecular lesion responsible for this disease. Polysomal Tg mRNA, therefore, was isolated, and cDNA was made using oligonucleotides as primers. This cDNA was multiplied by the polymerase chain reaction and cloned. In comparing the normal and abnormal sequences, we found a C—>G point mutation in exon 8 causing a change from TAC (Tyr)—>TAG (termination signal) at amino acid position 296. This mutation resulted in the appearance of a KpnI restriction site in the goiter DNA. The sequence of Tg mRNA preceding the stop codon was equal for normal and goitrous goats, except for one C—>T mutation in exon 5 which gave a Ser—>Leu transition. The KpnI site introduced by the C—>G point mutation was present in chromosomal DNA of the goitrous goats, making it possible to distinguish goats heterozygous for the defect from normal and goitrous animals. We calculated that the stop codon in exon 8 would result in a Tg polypeptide chain with a mol wt of 39,000, in good agreement with the mol wt of the in vitro and in vivo translation products. In conclusion, the C—>G mutation causing a stop codon in exon 8 is responsible for the Tg synthesis defect in Dutch goats.

Molecular basis of the thyroglobulin synthesis defect in Dutch goats.

A thyroglobulin (Tg) synthesis defect in Dutch goats causes congenital goiter and hypothyroidism. The disease is inherited in an autosomal recessive way and is linked to restriction fragment length polymorphisms (RFLPs) in the Tg gene. Previous studies showed that Tg mRNA isolated from the goiters was of normal size (8.4 kilobases). Translation of high mol wt polysomal Tg mRNA isolated from goiter in a cell-free rabbit reticulocyte lysate resulted in a single 35,000 mol wt Tg polypeptide. Tg antigens analyzed in T4-arrested goiters were glycosylated and had mol wt of 40,000 and 32,000. The aim of this study was to identify the molecular lesion responsible for this disease. Polysomal Tg mRNA, therefore, was isolated, and cDNA was made using oligonucleotides as primers. This cDNA was multiplied by the polymerase chain reaction and cloned. In comparing the normal and abnormal sequences, we found a C-->G point mutation in exon 8 causing a change from TAC (Tyr)-->TAG (termination signal) at amino acid position 296. This mutation resulted in the appearance of a KpnI restriction site in the goiter DNA. The sequence of Tg mRNA preceding the stop codon was equal for normal and goitrous goats, except for one C-->T mutation in exon 5 which gave a Ser-->Leu transition. The KpnI site introduced by the C-->G point mutation was present in chromosomal DNA of the goitrous goats, making it possible to distinguish goats heterozygous for the defect from normal and goitrous animals. We calculated that the stop codon in exon 8 would result in a Tg polypeptide chain with a mol wt of 39,000, in good agreement with the mol wt of the in vitro and in vivo translation products. In conclusion, the C-->G mutation causing a stop codon in exon 8 is responsible for the Tg synthesis defect in Dutch goats.

Histology and immunocytochemistry of differentiated thyroid carcinomas do not predict radioiodine uptake: a clinicomorphological study of 62 recurrent or metastatic tumours.

Sixty-two metastases or recurrences of differentiated thyroid carcinomas were investigated using conventional histology and immunocytochemistry for thyroglobulin (TG), thyroxine (T4) and triiodothyronine (T3). In each patient, 131I total body scans had been performed 4-10 weeks before surgery. Twenty-seven of the 62 tumours exhibited a predominance of follicles (A1), while 35 either exclusively or predominantly consisted of papillae or, in the case of follicular carcinomas, were predominantly trabecular or solid in structure (A2). TG and T4 immunoreactivity was observed in 60 cases, only 4 of these also expressing T3. Positive radioiodine uptake (RIU) was noted in 27 of 62 (44%) cases (A1:18/27 = 67%; A2:9/35 = 26%), 25 of which showed intraluminal TG and T4 positivity. Two follicular carcinomas showing RIU lacked follicular lumina, but exhibited strong diffuse cytoplasmic positivity for both TG and T4. In another 95 differentiated thyroid carcinomas, the structure of primary and secondary lesions was assessed. Of these, 27 (28%) showed a discordant pattern (A1/A2 or A2/A1) when comparing the structure of primary and secondary lesions. Our data suggest that differentiated thyroid carcinomas show a dissociation of TG/T4 expression and RIU, defects of iodine uptake and storage being found more frequently than a depression of TG and T4 synthesis. Intact synthesis of TG and T4, but not of T3 may be regarded as a prerequisite for RIU. Positive RIU is based on the presence of mature neoplastic follicles containing TG and T4 immunoreactive colloid and among follicular carcinomas, positive RIU may be encountered in neoplasms lacking follicular lumina but exhibiting strong cytoplasmic TG and T4 staining. Finally, the RIU of recurrent and metastatic PC and FC is not predictable from histological features of the primaries.

Thyroid albumin originates from blood.

Iodoalbumin has been found in the goiter of Dutch goats with a thyroglobulin synthesis defect. Immunohistochemical studies showed that in the goiter the percentage of follicles containing albumin was higher than that in normal thyroid glands. In the albumin-containing follicles of normal and goitrous glands, transferrin and immunoglobulin G could be found. Also, between the epithelial cells, serum proteins were detected. These results indicate intercellular passage of serum proteins. After in vivo labeling with 125I, goiter slices were incubated with [3H] leucine. Purified 125I-containing albumin did not contain [3H]leucine, while thyroglobulin antigens were double labeled. Cyanogen bromide treatment of albumin out of the goiter and of serum albumin gave comparable cleavage patterns, indicating that no great differences in amino acid composition, especially methionine, exist. In total RNA from goiter, no albumin mRNA could be detected after blotting and hybridization with goat albumin cDNA. Also, the polymerase chain reaction method with albumin DNA primers was unable to detect any albumin mRNA in normal and goitrous glands of goats. From these results we conclude that "thyralbumin" originates from blood and is not synthesized by the thyroid gland.

MULTIPLE PHENOTYPES INDUCED BY MUTANT HA-RAS IN THYROID EPITHELIAL-CELLS - CORRELATION WITH LEVEL OF EXPRESSION

To investigate the role of ras mutations in thyroid epithelial tumorigenesis, we introduced wild-type or mutant v- or c-Ha-ras genes into a sub-cloned rat thyroid follicular epithelial cell line using retroviral vectors and a neutral selection method (G418 resistance). Mutant, but not wild-type, ras induced a spectrum of clonal phenotypes. Interestingly, many clones showed an unchanged phenotype despite ras expression at levels greater than that of the endogenous gene. Further increase in ras expression was associated with altered morphology, loss of thyroid-specific differentiation (thyroglobulin synthesis) and growth factor independence, but not. anchorage-independence or tumorigenicity. The mutation site (codon 12 or 61) did not have a significant influence. The results clearly emphasize the critical importance of expression level in determining the phenotypic effect of mutant ras on epithelial cells.

Hormonogenesis in thyroid cells cultured on porous bottom chambers

Different processes implied in thyroid hormonogenesis (thyroglobulin, thyroperoxidase and hydrogen peroxide generating system expressions) and their regulation by TSH and iodide have been studied using porcine thyroid cells cultured in porous bottomed chambers. This system allowed to reproduce the functional bipolarity. Cells form a tight and polarized monolayer. Both apical and basolateral poles of epithelial cells were independently accessible and the cell layer separated two compartments which can contain different media. A major polarized secretion of thyroglobulin into the apical compartment was observed; it was increased in the presence of TSH as well as the thyroglobulin synthesis and mRNA level. These TSH effects were the consequence of adenylcyclase stimulation. Active transport of iodide, iodination of thyroglobulin and hormonosynthesis took place only in the presence of TSH. These steps occurred at the apical pole of cells. In the culture chamber system, thyroglobulin was weakly iodinated (6 atoms of iodide per mole of thyroglobulin; in vivo up to 40 atoms per mole) but hormonogenesis efficiency was close to this one observed in vivo (40%). Iodide concentrations higher than 0.5 microM daily added to the basal medium inhibited iodination of thyroglobulin and hormonosynthesis. Some components contained in culture media were inhibitors for iodination when they were present in the apical medium such as vitamins, amino acids and phenol red. The culture system appears to be interesting for pharmacological and toxicological studies.

A 3' splice site mutation in the thyroglobulin gene responsible for congenital goiter with hypothyroidism.

A case of congenital goiter with defective thyroglobulin synthesis has been studied in molecular terms. The patient is the fifth of a kindred of six, three of which have a goiter. The parents are first cousins. Segregation of thyroglobulin alleles in the family was studied by Southern blotting with a probe revealing a diallelic restriction fragment length polymorphism (RFLP). The results demonstrated that the three affected siblings were homozygous for the RFLP. Northern blotting analysis of the goiter RNA with a thyroglobulin probe suggested that thyroglobulin mRNA size was slightly reduced. Polymerase chain reaction amplification of the 8.5-kb thyroglobulin mRNA as overlapping cDNA fragments demonstrated that a 200-bp segment was missing from the 5' region of the goiter mRNA. Subcloning and sequencing of the cDNA fragments, and of the patient genomic DNA amplified from this region, revealed that exon 4 is missing from the major thyroglobulin transcript in the goiter, and that this aberrant splicing is due to a C to G transversion at position minus 3 in the acceptor splice site of intron 3. The presence in exon 4 of a putative donor tyrosine residue (Tyrosine nr 130) involved in thyroid hormone formation provides a coherent explanation to the hypothyroid status of the patient.