[The effect of thyroid hormones and 5-azacytidine on the gene transcription of malic enzyme and 6-phosphogluconate dehydrogenase]. / Vliianie tireoidnykh gormonov i 5-azatsitidina na transkriptsiiu genov malik-fermenta i 6-fosfogliukonatdegidrogenazy.

The effect of thyroid hormones and the methylation inhibitor 5-azacytidine on the level of expression of thyroid hormone-responsive genes (malic enzyme and 6-phosphogluconate dehydrogenase genes) has been studied. DNA-RNA hybridization has shown there is an inverse correlation between the level of DNA methylation and gene expression of malic enzyme and 6-phosphogluconate dehydrogenase. Thus it suggests that the regulation of thyroid hormone gene expression can take place via DNA methylation blocking and that DNA demethylation is part of structural changes essential to the binding of thyroid hormones with DNA elements recognized by thyroid hormone receptors and to further induction of synthesis of specific mRNA.

[Human thyroid gland DNA methylase in nodular and diffuse goiter]. / DNK-metilazy shchitovidnoi zhelezy cheloveka pri uzlovom i diffuznom zobe.

The heterogeneity and some properties of DNA-methylases isolated from nonmalignant human thyroid formations--nodular and diffuse goiters--have been studied. Isoelectrofocusing of methylase preparations produced 6-7 distinct activity peaks distinguished by pI, activity towards Ca2+ and Mg2+, sensitivity towards dithiothreitol and capacity to methylase cytosine into mono-, di- and tripyrimidine blocks in vitro. The degree of DNA methylation in vivo depended on the origin of of the goiter. Analysis of several methylase fractions by two-dimensional electrophoresis performed according to O'Farrell revealed the presence of major polypeptides having similar molecular masses (approximately 36 kDa). Besides the major component, all the preparations under study contained 6-10 characteristic minor components. The data obtained are suggestive of a structural and functional heterogeneity of human methylases.

[The regulation of the nuclear RNA polymerase activity of the liver and brain in rats of different ages by a cytoplasmic modulator of thyroxine action]. / Reguliatsiia aktivnosti RNK-polimerazy iader pecheni i golovnogo mozga krys razlichnogo vozrasta tsitoplazmaticheskim moduliatorom deistviia tiroksina.

The effect of physiological, concentration of thyroxine and cytoplasm thyroxine-binding protein (thyroxine action modulator, TAM4) on the RNA-polymerase activity in rat liver and bain cells in ontogenesis was studied. The physiological doses of the hormone were shown not to effect the RNA-polymerase of isolated liver and brain nuclei, while TAM4 caused the stimulation of enzymatic activity in these organs. ++TAM4 stimulating effect in liver correlated with increasing animal age, while the maximum RNA-polymerase activation by tAM was discovered in young rats under one month-At later stage the RNA-polymerase activity was same as in the controls. In is suggested that the cytoplasmic hormone-binding proteins take part in the formation of liver and brain sensitivity to the action of the thyroid gland hormones.

[Liver nuclear phospholipids of rats with varying thyroid status]. / Fosfolipidy iader pecheni krys s razlichnym tireoidnym statusom.

Thin-layer chromatography was used for fractionation of rat liver nuclear phospholipids after in vivo administration of 14C-sodium acetate. The administration of T3 to thyroidectomized rats caused a sharp increase in the incorporation of the label in all phospholipids of the nuclear fraction. The action of sphingomyelin and sphingomyelase on RNA-polymerase of nuclei isolated from the liver of thyroidectomized rats was tested. Sphingomyelin was shown to cause stimulation of RNA nuclear synthesis; parallel incubation with sphingomyelinase eliminated a stimulating effect of this phospholipid.

[The effect of thyroid hormones on DNA methylation in rat liver in vivo and in vitro]. / Vliianie tireoidnykh gormonov na metilirovanie DNK pecheni krys in vivo i in vitro.

Methylation of rat liver DNA was studied in vivo and in vitro in presence of various content of thyroid hormones. Both administration of triiodothyronine into intact rats and thyroidectomy led to considerable alterations in activity of endogenous DNA-methylases and in content of m5C in DNA although distinct correlation between these two factors was not detected. Alterations in the acceptor activity of endogenous DNA towards bacterial DNA-methylases of thy Mbu type demonstrated the processes occurring in vivo. The methylase probe 5'...GGA...3' proved to be the universal means for testing of all the types of thyroid status involving either free DNA or whole nuclei.

[Multiple forms of DNA methylases in hepatic cell nuclei of animals in the normal state and in pathology]. / Mnozhestvennye formy DNK-metilaziz iader pechenizhivotnykh v norme i pri patologii.

Using hydrophobic chromatography multiple nature of DNA-methylating enzymes have been revealed. It was established that the most active enzymatic fraction of rats' liver and that of chicken are completely identical.

[Changes in the chromatin structure of the thyroid cells related to the expression of the thyroglobulin gene]. / Izmeneniia struktury khromatina tireoidnykh kletok, sviazannye s ékspressiei gena tireoglobulina.

A highly purified thyroglobulin mRNA was isolated from human nodal euthyroid goiter. A full-length cDNA was synthesized from 33S RNA by using reverse transcriptase in the presence of human placenta ribonuclease inhibitor. A DNA complementary to human Tg mRNA was used in liquid hybridization experiments to quantify Tg mRNA. The amount of Tg mRNA in euthyroid nodal and congenital goiter was reduced. In thyroid cancer Tg specific mRNA was absent. Direct correlation between Tg gene expression in thyroid cells and DNAase-I hypersensitivity of chromatin from the thyroid gland nucleus was revealed.

[Thyroglobulin gene expression in human thyroid cells in various types of thyroid pathology]. / Ekspressiia gena tireoglobulina v kletkakh shchitovidnoi zhelezy cheloveka pri nekotorykh vidakh tireoidnoi patologii.

Highly purified thyroglobulin mRNA was isolated from human nodal euthyroid goiter. Full length cDNA was synthesized from 33S RNA by using reverse transcriptase in the presence of human placenta ribonuclease inhibitor. DNA complementary to human Tg mRNA was used in liquid hybridization experiments to determine the quantity of Tg mRNA. The amount of Tg mRNA in euthyroid nodal and congenital goiter was reduced. Tg specific mRNA was absent in thyroid cancer cells.

[Use of bacterial DNA-methylases for structuro-functional analysis of the eukaryotic genome]. / Ispol'zovanie bakterial'nykh DNK-metilaz dlia strukturno- funktsional'nogo issledovaniia genoma éukariot.

Bacterial DNA-methylases with known recognition sites (RS) were used as probes for structural-and-functional analysis of eukaryotic genome. Adenine and cytosine DNA-methylases recognizing 4 to 6-member unique and degenerative nucleotide sequences having a symmetrical and asymmetrical structure were used for probing. The use of a set of methylases enabled the selection of a probe that was the most sensitive for the given pathology. Thus, severe hypothyrosis was found to be associated with changes in the acceptor capacity of liver DNA in the heterologous++ methylation reaction as could be evidenced from testing by two probes, CCCC and GAATGC. In the cells of chicken liver hepatoma MC29, the acceptor capacity of DNA during GGA methylation appeared to be altered in the greatest degree. DNA-methylases with degenerative SR are weakly specific probes for the study of structural changes (methylation) of the animal genome.

[Mapping of the thyroglobulin gene in high molecular weight DNA from the human thyroid in normal conditions and in thyroid pathology]. / Kartirovanie tireoglobulinovogo gena v sostave bysokomolekuliarnoi DNK shchitovidnoi zhelezy cheloveka v norme i pri tireoidnoi patologii.

A highly purified thyroglobulin (Tg) mRNA was isolated from human nodal euthyroid goiter. Ultracentrifugation in a 5-20% sucrose density gradient revealed that the protein has a sedimentation coefficient of 33S. cDNA was synthesized from the 33S RNA, using reverse transcriptase in the presence of a human placental ribonuclease inhibitor. In order to detect the polymorphism of restriction fragment length, screening of high molecular weight DNA from normal human thyroid gland and from nodal euthyroid and diffuse toxic goiters was carried out, using Tg cDNA probes and plasmid DNA containing a Tg cDNA fragment. After restriction with endonuclease Hind III, three Tg-containing fragments were identified both in the normal and nodal euthyroid goiters, whereas restriction with endonuclease Bam HI revealed two Tg-containing fragments in nodal euthyroid and diffuse toxic goiters.

[Stimulation with triiodothyronine of the synthesis of DNA, total proteins, nuclear proteins and nuclear matrix proteins in cultured cells of hepatoma Morris 7777]. / Stimuliatsiia triiodtironinom sinteza DNK, total'nykh belkov, belkov iadra i iadernogo matriksa v kul'tiviruemykh kletkakh gepatomy Morrosa 7777.

Intensity of DNA and protein biosynthesis was decreased in Morris hepatoma 7777 cultivated cells in absence of thyroid hormones. Physiological concentrations of triiodothyronine increased synthesis of DNA and total proteins after lag phase within 12-48 hrs, while synthesis of nuclear and nuclear matrix proteins was stimulated already within 2 hrs. This suggests that stimulation of nuclear proteins biosynthesis occurred prior to the hormone effect on proliferation and cell metabolism. Alterations in sensitivity of chromatin to DNAase I were not observed. Response of the Morris hepatoma cells to thyroid hormones may be used in studies of molecular mechanisms of thyroid hormones action.

[The effect of thyroid hormones on structural-functional relations in rat liver chromatin]. / Vliianie tireoidnykh gormonov na strukturno-funktsional'nye vzaimootnosheniia v khromatine pecheni krys.

The effects of thyroid hormones on chromatin structure at different levels of its functional activity were investigated. No differences in the sensitivity of hepatocyte nuclei to DNAase I were found, presumably due to the restriction of acceptor sites for thyroid hormones on DNA.

[Binding of triiodothyronine to the nuclear matrix of the rat liver. The effect of thyroid hormones on the phosphorylation of nuclear matrix proteins]. / Sviazyvanie triiodotironina s iadernym matriksom pecheni krys. Vliianie tireoidnykh gormonov na fosforilirovanie belkov iadernogo matriksa.

The interaction of thyroid hormones with rat liver nuclear matrix proteins was studied. It was shown that the nuclear matrix contains the sites which bind triiodothyronine with a high affinity (Ka = 1.07 X 10(9) M-1) and limited capacity (maximal binding capacity--28.5 fmol triiodothyronine/100 micrograms protein). Electrophoretic analysis of triiodothyronine-binding matrix proteins revealed that the molecular mass of the major triiodothyronine-binding fraction is 50 000-52 000 Da. Injections of triiodothyronine to thyroidectomized animals stimulated the phosphorylation of all protein fractions of the nuclear matrix.

[Determination of the ratio of membrane-bound and free polyribosomes and of the content of thyroglobulin in these fractions in normal thyroid gland cells and in several types of thyroid diseases]. / Opredelenie sootnosheniia membranosviazannykh i svobodnykh poliribosom i soderzhaniia v étikh fraktsiiakh tireoglobulinovykh poliribosom v kletkakh shchitovidnoi zhelezy v norme i pri nekotorykh vidakh tireoidnoi patologii.

The ratio of membrane-bound and free polyribosomes in thyroid gland cells was decreased two-fold in nodular euthyroid goiter and five-fold--in Hashimoto thyroiditis; a slight increase in thyroglobulin synthesis in free polyribosomes was noted in goiter. Impairments in transfer of the translation products across membranes of rough endoplasmic reticulum may be one of the causes of deteriorations in thyroglobulin synthesis in goiters.

[Thyroglobulin biosynthesis and the content of thyroglobulin polyribosomes of the thyroid gland cells in the normal state and in various types of thyroid pathology]. / Biosintez tireoglobulina i soderzhanie tireoglobulinovykh poliribosom v kletkakh shchitovidnoi zhelezy v norme i pri razlichnykh vidakh tireoidnoi patologii.

The content of polyribosomes in human thyroid cells was studied in health and different forms of thyroid pathology. In thyroid pathology, thyroglobulin (TG) polyribosome content in thyroid cells diminished in the following order: diffuse toxic goiter greater than mixed euthyroid goiter greater than nodular toxic goiter greater than Hashimoto's goiter and complete absence of TG synthesizing polyribosomes in thyroid cancer and congenital goiter cells. No correlation was revealed between polyribosome TG content and thyroid function. It is inferred that impairment of TG biosynthesis in thyroid diseases is related to the decreased content of polyribosome TG in thyroid cells.

[Protein biosynthesis and thyroglobulin polyribosome content of thyroid cells in congenital goiter]. / Biosintez belkov i soderzhanie tireoglobulinovykh poliribosom v kletkakh shchitovidnoi zhelezy pri vrozhdennom zobe.

The basic protein, synthetized in a cell-free (S 30) protein-synthetizing system from the thyroid cells in congenital goitre, is thyroalbumin, while 19S thyroglobulin synthesis is sharply reduced. Determination of the population extent of thyroglobulin polyribosomes has shown that they are practically absent in congenital goitre. It is suggested that thyroglobulin deficit in congenital goitre is provoked by deficiency in functioning thyroglobulin mRNA.

[Regulation of incorporation of amino acids into the thyroid gland proteins by thyroid hormones]. / Reguliatsiia vkliucheniia aminokislot v belki shchitovidnoi zhelezy tireoidnymi gormonami.

The effect of thyroid hormones and of iodide (1 x 10(-7)M, 5 x 10(-5)M) on the protein synthesis in the cell-free system made of the nodular goiter cells was investigated. As revealed, with the action of thyroid hormones the 14C-amino acids incorporation into the trichloracetic acid-precipitated proteins was considerably diminished. The greatest inhibition of the 14C-amino acids incorporation was observed in toxic (5 x 10(-5)M) hormone concentration. The inhibitory action of triiodthyronine was more expressed than that of thyroxin. Addition of potassium iodide into the incubation medium led to a slight inhibition of the labeled amino acids incorporation. The results obtained permit to regard thyroid hormones as highly specific inhibitors of protein synthesis in the thyroid gland.