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Biomedical subjects

F Libert

Publications and source records attributed to F Libert.

At least 55 records · Page 3Linked to original sources
PubMed1990

Molecular cloning of a dog thyrotropin (TSH) receptor variant.

A clone coding for a variant form of thyrotropin receptor was isolated from a dog thyroid cDNA library. It was characterized by a 75 bp deletion in the coding region, additionally to minor modifications in the 3' untranslated region. The corresponding 25 amino acids deletion is located in the long NH2 terminal extracellular domain which is characteristic of the glycoprotein hormone receptors. This region of the protein is composed of imperfect repeats and the deletion corresponds exactly to one of the repeat units. This suggests that the repeats correspond to individual exons in the thyrotropin receptor chromosomal gene. It is not known whether the deletion of the repeat and the concomitant suppression of one of the N-glycosylation sites of the molecule do alter the receptor function.

Amino Acid Sequence↗
PubMed1990

An efficient screening morphological test for the identification and characterization of cyclic AMP-coupled hormone receptors.

We describe here a new method for the identification of adenylate cyclase-regulating receptors using the morphological response of Y1 cells to cAMP. The efficiency of this method was demonstrated with a cloned beta 2-adrenergic receptor: the coding region of a genomic beta 2-adrenergic receptor clone was amplified using the polymerase chain reaction, so that in vitro RNA transcripts of this DNA could be obtained. Y1 cells were microinjected with this RNA and assayed for sensitivity to isoproterenol. In the presence of this agonist, the microinjected cells rapidly showed morphological changes identical to those observed in the presence of ACTH, forskolin, or cholera toxin, agents that enhance cAMP accumulation in the control uninjected cells. This suggests the integration, within the injected cell membrane, of a functional beta 2-adrenergic receptor, whose activation by isoproterenol resulted in the intracellular formation of cAMP. This new qualitative screening method for the identification of adenylate cyclase-regulating receptors can be extended to any unknown receptor coupled to adenylate cyclase and absent in these cells. It has been applied to the identification of the dog thyrotropin receptor.

Adrenal Gland Neoplasms↗
PubMed1990

[A cloned protein belonging to the G protein-coupled receptor family has an essentially striatal distribution copying that of the major component of the D1 receptor].

RDC8 (correction of RCD8), a recently cloned new putative member of the G protein-coupled receptor family protein, is exclusively present in the medium-sized neurons of the striatum. This restricted localisation mimics the major striatal dopamine D1 receptor localisation and is of major importance for the understanding of basal ganglia physiology and degenerative diseases pathogeny such as Huntington's and Parkinson's disease. RDC7, another putative G protein-coupled receptor chemically closely related to RDC8 (correction of RCD8), is mainly distributed in pyramidal neurons of the cerebral cortex, the hippocampus and the claustrum, and in the amygdala, and may represent the minor extra-striatal variant of the D1 receptor.

Animals↗
PubMed1990

Distinct transcriptional effects of cAMP on 2 thyroid specific genes: thyroperoxidase and thyroglobulin.

The production of the thyroid hormones by the thyroid tissue is regulated by thyrotropin (TSH). TSH, through cAMP, enhances all steps of T3 and T4 synthesis, among which transcription of the genes encoding the precursor protein, thyroglobulin (TG) and the enzyme responsible for the iodination and coupling mechanisms, thyroperoxidase (TPO). Run-on transcription assays show that the kinetics of TG gene transcriptional activation by cAMP is slow (8 to 16 hours) in dog thyrocytes in primary culture, while it is rapid (1 hour) in dog thyroid slices. Activation is sensitive to cycloheximide, reflecting the need for ongoing protein synthesis. In contrast, stimulation of TPO gene transcription is rapid in both experimental systems and is not inhibited in the presence of cycloheximide. It is concluded that different regulatory mechanisms are implicated in the control of Tg and TPO gene transcription by cAMP. However, the stimulation of TG and TPO gene transcription are equally suppressed by inhibition of cAMP-dependent protein kinase, which suggests that both regulatory mechanisms involve protein phosphorylation.

Animals↗
PubMed1989

Cloning, sequencing and expression of the human thyrotropin (TSH) receptor: evidence for binding of autoantibodies.

A human thyroid cDNA library was screened by hybridization with a dog thyrotropin receptor (TSHr) cDNA. Sequencing of the resulting clones identified a 2292 residue open reading frame encoding a 744 amino acid mature polypeptide presenting 90.3% similarity with the dog TSHr. Two major transcripts (4.6 and 4.4 kilobases) were identified in the human thyroid which suggests that alternative splicing could generate multiple forms of human TSHr. Transfection of the coding sequence in COS-7 cells conferred to a membrane preparation of these cells the ability to bind specifically TSH. TSH binding was completely displaced by immunoglobulin preparations from patients with idiopathic myxoedema.

Amino Acid Sequence↗
PubMed1989

Molecular cloning of the thyrotropin receptor.

The pituitary hormone thyrotropin, or thyroid-stimulating hormone (TSH), is the main physiological agent that regulates the thyroid gland. The thyrotropin receptor (TSHR) was cloned by selective amplification with the polymerase chain reaction of DNA segments presenting sequence similarity with genes for G protein-coupled receptors. Out of 11 new putative receptor clones obtained from genomic DNA, one had sequence characteristics different from all the others. Although this clone did not hybridize to thyroid transcripts, screening of a dog thyroid complementary DNA (cDNA) library at moderate stringency identified a cDNA encoding a 4.9-kilobase thyroid-specific transcript. The polypeptide encoded by this thyroid-specific transcript consisted of a 398-amino acid residue amino-terminal segment, constituting a putative extracellular domain, connected to a 346-residue carboxyl-terminal domain that contained seven putative transmembrane segments. Expression of the cDNA conferred TSH responsiveness to Xenopus oocytes and Y1 cells and a TSH binding phenotype to COS cells. The TSHR and the receptor for luteinizing hormone-choriogonadotropin constitute a subfamily of G protein-coupled receptors with distinct sequence characteristics.

Amino Acid Sequence↗
PubMed1989

Selective amplification and cloning of four new members of the G protein-coupled receptor family.

An approach based on the polymerase chain reaction has been devised to clone new members of the family of genes encoding guanosine triphosphate-binding protein (G protein)-coupled receptors. Degenerate primers corresponding to consensus sequences of the third and sixth transmembrane segments of available receptors were used to selectively amplify and clone members of this gene family from thyroid complementary DNA. Clones encoding three known receptors and four new putative receptors were obtained. Sequence comparisons established that the new genes belong to the G protein-coupled receptor family. Close structural similarity was observed between one of the putative receptors and the 5HT1a receptor. Two other molecules displayed common sequence characteristics, suggesting that they are members of a new subfamily of receptors with a very short nonglycosylated (extracellular) amino-terminal extension.

Amino Acid Sequence↗
PubMed1989

Cloning and sequencing of a calcium-binding protein regulated by cyclic AMP in the thyroid.

p24 is a thyroid protein (Mr 24,000) identified by two-dimensional gel electrophoresis on the basis that its synthesis and phosphorylation are up-regulated by thyrotropin and cyclic AMP agonists. p24 cDNA was cloned from a lambda gt11 cDNA library using a polyclonal antibody raised against the protein recovered from a Western blot spot. The encoded polypeptide (189 residues) displays a putative target-site for phosphorylation by cyclic AMP-dependent protein kinase and belongs to the superfamily of proteins binding Ca2+ through 'EF hand' domains. It presents four such domains of which two agree closely with the consensus. The ability of p24 to bind Ca2+ has been directly confirmed on Western blots. p24 was detected in many tissues including the salivary glands, the lung and the brain. The ubiquitous nature of p24, together with its regulatory and sequence characteristics suggest that it constitutes an important target common to the cyclic AMP and Ca2+-phosphatidylinositol cascades.

Animals↗