Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:4.6.1.1 (adenylate cyclase)
19,190 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We studied the effect of bromolevamisole (BL) and other imidazo [2,1-b] thiazole derivatives--bromodexamisole (BD) and levamisole (LV)--on adenylate cyclase (AC) activity. BL and BD both inhibited forskolin-activated human thyroid AC, while LV had no effect. This inhibition was non-stereospecific and the IC50 values, as measured with 1 mM ATP and 40 microM forskolin, were 0.95 and 0.80 mM for BL and BD, respectively. In contrast, human thyroid alkaline phosphatase (ALP) inhibition was stereospecific, with IC50 values of 0.0012 mM for BL and 0.9 mM for BD. LV was a 10-fold weaker inhibitor of ALP than BL. These results show that ALP inhibition is not correlated with forskolin-activated AC inhibition. Furthermore, in the presence of a competitive inhibitor of GTP (0.1 mM guanosine 5'-O-(2-thiodiphosphate), BL retained its antagonizing effect on forskolin-activated AC which suggests a direct action on the catalytic subunit. The inhibition was of the mixed type, indicating a complex interaction between BL and AC. Glucagon-activated AC activity in rat liver membranes was also inhibited by BL, although to a slightly lesser degree than thyroid stimulating hormone (TSH)-activated AC from human thyroid for a given BL concentration. In cultured human thyroid cells, BL (0.25 mM) induced a potent decrease in cAMP accumulation after 2 hr of stimulation by TSH. Taken together, these results show that BL inhibits AC and that this inhibition is not organ-specific.
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PMID:Effect of bromolevamisole and other imidazo [2,1-b] thiazole derivatives on adenylate cyclase activity. 131 3

The ligand specificity and biochemical properties of the human (h) FSH receptor are poorly characterized due to the low abundance of these receptors and the limited availability of human tissues. Using a fragment of rat FSH receptor cDNA, we screened a human testicular cDNA library and obtained a FSH receptor cDNA covering the entire amino acid-coding region. After transfection of a human fetal kidney cell line (293) with the hFSH receptor cDNA, radioligand receptor analysis revealed the presence of high affinity (Kd, 1.7 x 10(-9) M) FSH-binding sites on the plasma membrane. Both recombinant and wild-type hFSH displaced [125I]hFSH binding, with ED50 values of 25 and 70 ng/ml, respectively, whereas hLH, hCG, and hTSH were ineffective. Although human, rat(r), and ovine FSH as well as equine CG competed for rat testicular FSH receptor binding, only hFSH and rFSH interacted effectively with the recombinant hFSH receptor, suggesting that species-specific ligand recognition exists between human and rodent receptors. After incubation of transfected cells with hFSH, but not recombinant hLH or hCG, a dose-dependent increase (ED50, 10 ng/ml) in extracellular cAMP accumulation was observed, indicating a functional coupling of the expressed human receptor with the endogenous adenyl cyclase. In cells cotransfected with the FSH receptor expression plasmid and a luciferase reporter gene driven by the promoter of a cAMP-responsive gene, treatment with hFSH, but not hCG, resulted in a dose-dependent increase in luciferase activity. Northern blot analysis using a cRNA probe derived from the human receptor cDNA indicated the presence of multiple FSH receptor mRNA transcripts (7.0, 4.2, and 2.5 kilobases) in RNA prepared from human follicular phase ovary, but not from human corpus luteum or placenta. Additionally, two FSH-binding sites of 76 and 112 kilodaltons were detected in transfected 293 cells after ligand/receptor cross-linking and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. These results demonstrate the expression of functional hFSH receptor with unique ligand specificity and provide new data on the biochemical properties of the human receptor at the mRNA and protein levels.
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PMID:Expression of recombinant human follicle-stimulating hormone receptor: species-specific ligand binding, signal transduction, and identification of multiple ovarian messenger ribonucleic acid transcripts. 132 83

The syndrome of Albright hereditary osteodystrophy (AHO), pseudohypoparathyroidism (PHP) and pseudopseudohypoparathyroidism (PPHP) is clinically and genetically heterogeneous. Classically, patients with PHP have the skeletal features of AHO, resistance to multiple hormones that work via cAMP such as parathyroid hormone and thyroid stimulating hormone, and deficient activity of Gs protein, the guanine nucleotide-binding protein that stimulates adenylate cyclase. However, patients without hormone resistance but with AHO and Gs deficiency were described (PPHP), as well as patients with multiple hormone resistance but without AHO or Gs deficiency. In a few patients with deficient Gs activity, hypothyroidism rather than hypocalcemia was the initial presentation of the disorder. We describe here a new variant of the syndrome, affecting 5 individuals in a 3 generation family with AHO, normal Gs activity and hypothyroidism. In the first 2 generations, mild features of AHO were present. The 2 sibs in the third generation had severe manifestations of AHO, including mild mental retardation as well as hypothyroidism. Diagnosis of congenital osteoma cutis at birth of the proband led to the diagnosis of the family. Elucidation of the molecular defect will shed light on the relationship between hormone resistance and AHO, as well as on the physiological mechanism of hormonal signal transduction.
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PMID:Albright hereditary osteodystrophy with hypothyroidism, normocalcemia, and normal Gs protein activity: a family presenting with congenital osteoma cutis. 162 72

To ascertain the thyrotropic activity of human chorionic gonadotropin in sera of normal pregnant women, we examined the adenylate cyclase activation in the cultured FRTL-5 cells by extracted hCG from 7 normal pregnant women. hCG was extracted from the sera using anti-hCG-beta subunit monoclonal antibody-coated microwells, eluted with 2 mol/l guanidine-HCl, and reconstituted with hypotonic Hanks' solution. FRTL-5 cells were precultured in 5H medium, incubated for 2 h with the serum extracts, and the cAMP released into the medium was measured. hCG levels in serum extracts ranged from 1100 to 6800 IU/l; values corresponded to 1.4-19.8% compared with those in the original serum samples. Addition of the extracts to FRTL-5 cells resulted in significant increases in the cAMP accumulation, ranging from 9.8 to 59.0 nmol/l. cAMP levels were also increased in a dose-dependent manner by adding purified hCG as well as crude hCG and hTSH to FRTL-5 cells. These findings suggest that the thyroid gland of normal pregnant women may actually be stimulated by hCG itself.
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PMID:Thyroid-stimulating activity of human chorionic gonadotropin in sera of normal pregnant women. 170 May 69

A chemical relatedness has been described between the beta-subunit of cholera toxin and that of the four dimeric glycoprotein hormones (hCG, hLH, hFSH and hTSH). However, antibodies induced by cholera toxin did not crossreact, when tested by labeled hCG binding and immunocytochemistry, with the beta-subunit of hCG. It appears that differences in the tertiary structures, as shown in this study, account for distinct epitopes. Similarities in biological activity between these two compounds, such as induction of adenyl cyclase or a protective effect against some tumors, are not based on immunological mechanisms.
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PMID:Lack of immunological analogy between the beta-subunits of cholera toxin and human choriogonadotropin. 242 25

Monoclonal antibodies that bind to the TSH receptor were obtained by an autoantiidiotypic approach in which immunization of BALB/c mice was performed with mixtures of bovine (b) and human (h) TSH. Two of 28 positive wells were selected for cloning and characterization: D2 and 4G11. Their antiidiotypic character was evidenced by TSH-inhibitable binding to affinity-purified polyclonal anti-TSH. The specificity of D2 and 4G11 for the hormone-binding region of the TSH receptor was demonstrated by several findings: 1) they inhibited the binding of [125I]iodo-bTSH to receptor in a dose-dependent manner; 2) their binding to partially purified thyroid plasma membranes could be completely inhibited by bTSH and hTSH; and 3) they inhibited the TSH-dependent growth and adenylate cyclase stimulation in FRTL-5 cells in a dose-dependent manner. By Western blot analysis of bovine thyroid membranes, D2 bound to a polypeptide of 188,000-195,000 mol wt under nonreducing conditions and 54,000-59,000 mol wt after treatment of membranes with beta-mercaptoethanol; the 4G11 epitope was undetectable. Scatchard analysis of the binding of 125I-labeled antibodies to receptor showed that 4G11 bound to a single site with a Kd of 5.7 X 10(-9) M, whereas D2 showed complex binding characterized by high affinity (Kd = 1.74 X 10(-11) M) and low affinity (Kd = 1.3 X 10(-8) M) sites. Binding studies in which D2 and 4G11 competed with each other for the TSH receptor showed mutual but unequal inhibition. The data suggest that portions of the D2 and 4G11 epitopes overlap, but that there is a high affinity binding site(s) for D2 for which 4G11 competes less effectively. The binding of D2 and 4G11 to TSH receptor was inhibited by monoclonal antibodies secreted by Graves' heterohybridomas, showing that D2 and 4G11 share characteristics with autoantibodies of Graves' disease and lending support to the hypothesis that idiotypic network interactions may play a role in the pathogenesis of Graves' disease.
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PMID:Monoclonal antibodies to the thyrotropin receptor raised by an autoantiidiotypic protocol and their relationship to monoclonal autoantibodies from Graves' patients. 245 50

In cultured porcine thyroid cells, during 60 min incubation phorbol 12-myristate 13-acetate (PMA) had no effect on basal cyclic AMP accumulation and slightly stimulated cyclic AMP accumulation evoked by thyroid stimulating hormone (TSH) or forskolin. Cholera toxin-induced cyclic AMP accumulation was significantly stimulated by PMA. On the other hand, cyclic AMP accumulation evoked by prostaglandin E1 or E2 (PGE1 or PGE2) was markedly depressed by simultaneous addition of PMA. These opposing effects of PMA on cyclic AMP accumulation evoked by PGE and cholera toxin were observed in a dose-related fashion, with half-maximal effect of around 10(-9) M in either case. The almost same effects of PMA on cyclic AMP accumulation in basal and stimulated conditions were also observed in freshly prepared thyroid cells. The present study was performed in the presence of phosphodiesterase inhibitor, 3-iso-butyl-1-methylxanthine (IBMX), indicating that PMA affected adenylate cyclase activity. Therefore, it is suggested that PMA may modulate the production of cyclic AMP in response to different stimuli, possibly by affecting several sites in the adenylate cyclase complex in thyroid cells.
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PMID:Phorbol esters modulate cyclic AMP accumulation in porcine thyroid cells. 245 71

Humoral and cellular immune responses are both involved in autoimmune disorders of the thyroid gland. In the last five years, new substantial data have been obtained on the nature and the expression of thyroid cell surface autoantigens and on the demonstration of the functional heterogeneity of autoantibodies to the thyroid stimulating hormone (TSH) receptor. In the present report, attention will be mainly focused on recent studies carried out in our laboratory. The main autoantigens so far identified include the 'microsomal' antigen, thyroglobulin and the TSH receptor. For many years the 'microsomal' antigen (M) was considered a poorly characterized constituent of the cytoplasm of the thyroid cell. In the last five years, several lines of evidence were provided indicating that M is also well represented on the surface of the follicular cell and is identical to thyroid peroxidase (TPO). The use of anti-TPO monoclonal antibodies, presently available, have confirmed this antigenic identity. Microsomal (anti-TPO) antibodies are very useful markers of autoimmune thyroid disorders and are generally present in Hashimoto's thyroiditis, idiopathic myxedema and Graves' disease. TSH receptor antibodies (TRAb) are present in the sera of patients with Graves' disease. TRAb are able to stimulate thyroid adenylate cyclase and also to mimic TSH in its thyroid growth stimulation. Thus, these antibodies may have a pathogenetic role in goiter formation and in thyroid hyperfunction in Graves' disease. TRAb were also shown to inhibit both TSH binding to its receptor and TSH-stimulated adenylate cyclase activity. Recently TRAb, which inhibited TSH-stimulated adenylate cyclase activity, were found in idiopathic myxedema patients and may be responsible for impairment of thyroid function.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Thyroid autoantigens and their relevance in the pathogenesis of thyroid autoimmunity. 249 24

We have studied the in vitro TSH secretion and the adenylate cyclase (AC) activity of a human pituitary adenoma surgically removed from a hyperthyroid patient showing high serum TSH levels. The tumor appeared almost homogeneously constituted by cells positive for an anti-TSH-beta antiserum and showing the ultrastructural characteristics of the adenomatous thyrotrophs. Adenoma fragments released in vitro a large amount of TSH (148.4 microU/mg prot/30 min), alpha-subunit (35.5 ng/mg prot/30 min) and TSH-beta (10.1 ng/mg prot/30 min). The effects of somatostatin (GHRIH) and dopamine (DA) on the hormone release have been tested in vitro. Both agents markedly inhibited the release of intact TSH and TSH-beta whereas the release of alpha-subunit was less affected. The two agents were effective at concentrations higher than 10(-8)M. The ability of GHRIH and DA in modulating the AC activity was investigated in membrane fraction preparations. GHRIH inhibited AC at concentrations higher than 10(-7)M. The maximal inhibition was 32% at 10(-5)M. Conversely, DA slightly stimulated AC activity. This effects was not mimicked by the dopaminergic ergot CH 29-717, which was completely ineffective on the enzyme. These results suggest that: 1) in this TSH-secreting pituitary adenoma a normal secretory response to the inhibiting agents (GHRIH and DA) is present; 2) different mechanisms of transduction of the GHRIH and DA signals (cAMP dependent and cAMP independent) could be operating in this tumor.
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PMID:In vitro studies on TSH secretion and adenylate cyclase activity in a human TSH-secreting pituitary adenoma. Effects of somatostatin and dopamine. 286 99

TSH is a heterodimeric glycoprotein hormone, whose dissociated subunits are without biological activity. This has precluded the assessment of the relative contribution of each subunit to hormone action. We have raised anti-idiotypes to monoclonal antibodies specific, respectively, for the alpha and beta hTSH subunits. The anti-beta anti-idiotype inhibited 125I-hTSH binding to the beta subunit-specific monoclonal quantitatively, whereas 125I-hTSH binding to the alpha subunit-specific monoclonal was not inhibited by anti-alpha anti-idiotypes, suggesting that only the former is an "internal image" anti-idiotype. Neither of the two anti-idiotypes nor equimolar mixtures thereof inhibited 125I-bTSH binding to thyroid membranes, even though radiolabelled anti-idiotypes showed saturable binding to thyroid plasma membrane which was inhibited 41-65% by bTSH. Each anti-idiotype alone caused 9% inhibition (compared to 50% by NRIgG) of thyroid plasma membrane adenylate cyclase. Equimolar mixtures (125 micrograms/ml IgG of each anti-idiotype) induced enzyme activity equivalent to 85% of that of 250 mU/ml of TSH. The TSH-like action of the two anti-idiotypes was also reflected in their capacity to increase (450% by 250 micrograms/ml IgG compared to normal rabbit IgG) the uptake of 131I into isolated thyrocytes and to promote the organization of such cells into follicular structures. At 250 micrograms/ml, anti-beta anti-idiotype promoted the organization of small follicles and only at a concentration of 500 micrograms/ml did it enhance 131I uptake.
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PMID:Use of anti-idiotypic antibodies as probes for the interaction of TSH subunits with its receptor. 349 59


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