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G Vassart

Publications and source records attributed to G Vassart.

At least 253 records · Page 14Linked to original sources
PubMed1989

The human genes for calbindin 27 and 29 kDa proteins are located on chromosomes 8 and 16, respectively.

Genomic clones coding for the brain calcium-binding protein, calbindin 29 kDa, were isolated from a human library. A fragment containing exon 2 was used as a probe to investigate the presence of the gene in human x rodent somatic cell hybrids. The gene was unambiguously assigned to chromosome 16. The closely-related calbindin 27 kDa gene was previously assigned to chromosome 8. These two genes, deriving from a common ancestor, thus appear to have been separated during vertebrate evolution.

Animals↗
PubMed1989

Antibodies to human thyroid peroxidase in autoimmune thyroid disease: studies with a cloned recombinant complementary deoxyribonucleic acid epitope.

Previous studies carried out by screening a lambda gt11 human thyroid cDNA library with serum samples from selected patients with Hashimoto's thyroiditis and a polyclonal antibody to porcine thyroid peroxidase (TPO) confirmed, at the molecular level, that TPO is a major component of the thyroid microsomal antigen (M). That investigation led to the isolation of a clone (C2) which encodes an 85-amino acid segment of TPO and harbors a major epitope recognized by serum from several patients with autoimmune thyroid disease that contained anti-M autoantibodies (MAb). In this study, C2 antigen that was produced as a beta-galactosidase fusion protein was used to establish an enzyme-linked immunoabsorbent assay for the detection of anti-C2 autoantibodies (C2Ab). C2Ab then were assayed in 191 patients with different autoimmune and nonautoimmune thyroid disorders, and 50 patients with nonthyroidal autoimmune diseases. The results were compared with the titers of anti-TPO antibodies (TPOAb; as detected by monoclonal antibody-assisted RIA) and MAb (as detected by passive hemagglutination). Positive C2Ab was found in the serum of 85 of 136 (63%) patients whose serum contained TPOAb and/or MAb. A significant positive correlation was found between the levels of C2Ab and those of TPOAb (r = 0.76; P less than 0.001) or MAb (r = 0.69; P less than 0.001), which was independent of the type of underlying autoimmune thyroid disorder. Low levels of C2Ab also were found in 10 of 105 (9%) serum samples that did not contain TPOAb. Western blot analysis carried out on the latter samples showed that in 2 samples the apparent C2Ab reactivity was due to the presence of antibodies reacting with beta-galactosidase. In conclusion, we confirmed the validity of screening lambda gt11 cDNA human thyroid libraries to better characterize thyroid autoantigens and demonstrated the feasibility of using recombinant proteins to establish diagnostic assays for autoantibodies.

Antibodies↗
PubMed1989

Control of thyroperoxidase and thyroglobulin transcription by cAMP: evidence for distinct regulatory mechanisms.

The expression of the genes coding for thyroglobulin (TG), and thyroperoxidase (TPO), are regulated by TSH. These effects are mediated by cAMP as they are reproduced by forskolin. In vitro run-on transcription assays performed on nuclei isolated from dog thyrocytes in culture or from dog thyroid slices, indicate that the forskolin-induced transcriptional stimulation of TG and TPO genes are very different. For the TG gene, the kinetics of transcriptional activation vary according to the experimental model: it is rapid (1 h) in thyroid slices and slow (8 h) in primary cultures. In contrast, TPO induction is rapid in both cases. In primary cultures, insulin is responsible for the basal level and for a part of forskolin-induced TG transcription, whereas TPO transcription is not affected by insulin. The forskolin-induced increase of TG transcription requires ongoing protein synthesis, as it is blocked by cycloheximide. TPO gene transcription is unaffected by cycloheximide. Taken together with previous data on the two genes, our results suggest that while TPO regulation corresponds to the classical model of genes in which the promoter is regulated directly via cAMP regulatory elements, TG gene regulation involves the synthesis of an intermediary, rapid turnover trans-acting protein.

Animals↗
PubMed1989

Towards an antigenic map of human thyroglobulin: identification of ten epitope-bearing sequences within the primary structure of thyroglobulin.

The aim of this study was to define, at the molecular level, epitopes of thyroglobulin recognized by heterologous antibodies and autoantisera. One hundred thousand clones from a lambda gt 11 human thyroid cDNA library were screened using a rabbit antibody to human thyroglobulin at a 1:2000 dilution. Twenty clones were plaque-purified to homogeneity, and characterization and sequencing of their cDNA inserts showed that they represented four distinct regions of the thyroglobulin molecule, one of them being the 22 carboxyl-terminal amino acids. Rescreening of the library with the same rabbit antibody to human thyroglobulin absorbed with peptides encoded by the carboxyl-terminal clone, led to the definition of six further epitope-bearing fragments of thyroglobulin. The ten regions that we have identified were recognized by ten further rabbit antibodies to human thyroglobulin, showing that they are representative of the repertoire of heterologous epitopes. In contrast, none of the ten heteroepitope-bearing fragments was recognized by sera from ten patients with autoimmune thyroid disease with various titres of thyroglobulin antibodies. Screening of 2 x 10(6) clones from the library using a pool of ten autoantisera (individual sera diluted to 1:1000), and of 1 x 10(6) clones using a single autoantiserum of very high antithyroglobulin titre (diluted 1:400) resulted in no thyroglobulin clones being isolated. The significance of these results to the immune process is discussed.

Amino Acid Sequence↗
PubMed1989

Definition, at the molecular level, of a thyroglobulin-acetylcholinesterase shared epitope: study of its pathophysiological significance in patients with Graves' ophthalmopathy.

The nature of the putative autoantigen in Graves' ophthalmopathy (Go) remains an enigma but the sequence similarity between thyroglobulin (Tg) and acetylcholinesterase (ACHE) provides a rationale for epitopes which are common to the thyroid gland and the eye orbit. In an attempt to define the shared epitope, we have screened a lambda gt 11 human thyroid cDNA library using a polyclonal antibody to Torpedo ACHE and isolated two clones, which upon sequencing, were shown to contain Tg segments, corresponding to portions of the C terminal part of the molecule which has a high similarity with ACHE. Having demonstrated the existence of an epitope common to Tg and ACHE, the clones have been further tested and found to be positive in lysis plaque assays with 1/10 sera from patients with Hashimoto's thyroiditis (HT), 8/8 from patients with Graves' ophthalmopathy and 0/8 normal sera. We have investigated the physiological significance of this common epitope by in situ immunolocalization studies in which the polyclonal antibody to Torpedo ACHE (which was used for screening the library) and immunoglobulins (Igs) from 6 Go patients tested were shown to bind to end plate regions of human foetal muscle fibres which were concurrently shown to be rich in cholinesterase activity: Igs from 3 normal individuals and 2 patients with Hashimoto's thyroiditis did not bind. The results demonstrate and characterize an epitope which is common to Tg and ACHE and show that Go patients Igs contain antibodies which bind to muscle end plates rich in cholinesterase. The significance of these findings to the pathogenesis of Go is discussed.

Acetylcholinesterase↗
PubMed1988

Thyroid-specific and cAMP-dependent hypersensitive regions in thyroglobulin gene chromatin.

Two regions hypersensitive to DNase I digestion were found in a 7-kb segment of thyroglobulin gene 5'-flanking sequences in the chromatin from bovine thyroid. The most upstream site (-2000 to -1600 bp relative to the transcriptional start) was found in thyroid chromatin only, but independently of actual expression of the gene. It therefore represents a tissue-specific characteristic which may be associated with the commitment of the thyroglobulin gene to transcriptional activity. The very 5' end of the gene and the proximal promoter sequences (-100 to +60 bp relative to transcriptional start), constitute the second site, the hypersensitive character of which could be directly correlated with transcriptional activity. The structural changes occurring in this region of the chromatin were dependent on cAMP stimulation of the thyroid cells.

Animals↗
PubMed1988

Use of DNA bar codes to resolve a canine paternity dispute.

The DNA fingerprinting method was used to resolve a canine paternity dispute. During the same estrus, a Shih Tzu bitch was inseminated by 2 dogs--a Shih Tzu and a Coton de Tulear. Because both breeds are alike phenotypically, it was difficult to decide whether the pups were purebred or of mixed breeding. The DNA bar codes indicated unambiguously that the 2 sires had fathered one pup each, thus documenting superfecundation.

Alleles↗
PubMed1988

The human pregnancy-specific beta 1-glycoprotein (PS beta G) and the carcinoembryonic antigen (CEA)-related proteins are members of the same multigene family.

Pregnancy-specific beta 1-glycoprotein (PS beta G), a major product of the placenta with unknown function, consists of a set of glycoproteins synthesized by the syncytiotrophoblast. We report here the molecular cloning of 3 cDNA encoding different members of the PS beta G family. Two clones (C, D) correspond to a single transcript undergoing differential splicing. The third one (E) originates from a different gene. All three clones have identical (C, D) or similar (E) coding sequences except for the last residues at their carboxyl end. They contain 93 residue motifs related to the ancestral Ig-like domain which makes them new members of this gene superfamily. A striking sequence similarity (50 to 60%) is observed between PS beta G and carcinoembryonic antigen (CEA)-related proteins. The evolutionary relationship between CEA and PS beta G points to a possible common function in the control of cell invasion and/or metastasis.

Amino Acid Sequence↗
PubMed1988

Carrier prediction of cystic fibrosis in 36 families by means of restriction fragment length polymorphism.

Transmission of cystic fibrosis (CF) was studied in 36 families with at least one affected and one unaffected child. DNA was prepared from peripheral leukocytes and submitted to restriction fragment length polymorphism (RFLP) analysis with two CF probes (pj3.11 and met). Twenty families were shown to be informative so that accurate predictions could be made of the status of the offspring. Sixteen were only partially informative. The allele frequency was similar to that originally reported except for one Msp I site detected with the pj3.11 probe, for which we found a significantly higher heterozygote frequency, making it more informative than expected in our population sample. Pedigree analysis demonstrated no obligate recombinant between CF and the polymorphic markers.

Cystic Fibrosis↗
PubMed1988

Transcriptional regulation of the thyroperoxydase gene by thyrotropin and forskolin.

The expression of the gene coding for thyroperoxydase, the main enzyme involved in the synthesis of the thyroid hormones, is controlled by thyroid stimulating hormone. In vitro transcription assays performed on nuclei isolated from dog thyroid cells in primary culture showed that this control is at the transcription level, takes place rapidly (1 h) and is cyclic AMP-dependent as it is mimicked by forskolin. Insulin does not seem to be an important modulator of the thyroperoxydase gene expression.

Animals↗
PubMed1988

DNA fingerprinting in domestic animals using four different minisatellite probes.

Four probes known to allow DNA fingerprinting in the human (M13, Jeffreys' core sequence, the human alpha globin hypervariable region [HVR], and a mouse probe related to the Drosophila Per gene) were checked for their ability to reveal "genetic bar codes" in cattle, horses, pigs, dogs, chickens, and a European cyprinid fish, the barbel (Barbus barbus L.). Individual-specific patterns were obtained in cattle using M13, Jeffreys' core sequence, and the alpha globin HVR, in horses, dogs, and pigs using M13, Jeffreys' core sequence, and the Per probe, and in chicken and fish using the four different probes. Although we observed a considerable heterogeneity in the extent of interindividual variation, depending on the particular probe-species combination, the fingerprints are polymorphic enough to be used efficiently in animal identification, paternity testing, and as a source of genetic markers for linkage analysis. These markers should substantially accelerate the mapping of genes affecting economically important traits.

Animals↗
PubMed1988

Regional localization of the gene for thyroid peroxidase to human chromosome 2pter----p12.

A 2.0-kb thyroid peroxidase cDNA of human origin was used as probe for Southern blot hybridization of genomic DNA from human somatic cells and human-rodent somatic cell hybrids. The results showed that the gene coding for human thyroid peroxidase is located on chromosome. 2. Further analysis of hybrids derived from Burkitt lymphoma cells carrying a (2;8)(p12;q24) translocation revealed that the gene maps to the region 2pter----p12.

Animals↗
PubMed1987

Human 27-kDa calbindin complementary DNA sequence. Evolutionary and functional implications.

Human 27-kDa calbindin cDNA clones were selected by antibody screening from lambda gt11 brain libraries. The sequence revealed an open reading frame coding for a protein of 261 amino acids, containing four active calcium-binding domains, and two modified domains that had presumably lost their calcium-binding capability. Comparison with chick and bovine calbindins showed that the protein was highly conserved in evolution (evolutionary rate: 0.3 x 10(-9) amino acid-1 year-1) and that active and inactive domains were equally conserved. From the data we postulate that calbindin has an important physiological function involving protein--protein interactions. Comparison of calcium-binding domains from various proteins suggested that all members of the troponin C superfamily derive from a common two-domained ancestor, but that duplications leading to calbindin and to the four-domained calcium-binding proteins took place independently on different branches of the evolutionary tree. Preliminary data showed that another calcium-binding protein, homologous to calbindin, is present in the brain and encoded by a different gene.

Amino Acid Sequence↗
PubMed1987

Thyroperoxidase, an auto-antigen with a mosaic structure made of nuclear and mitochondrial gene modules.

A lambda gt11 cDNA library was constructed from a normal human thyroid and screened with a rabbit anti-porcine thyroperoxidase antibody. A series of thyroperoxidase (TPO) clones were obtained which allowed determination of the complete primary structure of the protein. The library was also screened with serum from a patient with Hashimoto's thyroiditis, an autoimmune disease characterized by the presence in the serum of high titers of autoantibodies directed against the 'microsomal antigen' (McAg). Comparison of the cDNA sequences from TPO clones and McAg clones provides definite proof that the McAg is TPO. A short segment of TPO was characterized as bearing a major epitope involved in autoimmunity. The primary structure of TPO was 42% homologous to myeloperoxidase (MPO). It contains, in addition, a C-terminal extension with a membrane anchor region contiguous to two domains encoded by modules belonging to the EGF and C4b gene families. The existence in TPO of still another domain presenting a significant homology with a putative heme-binding region of cytochrome C oxidase polypeptide I raises the possibility that a mitochondrial gene module has contributed a piece to the evolution of a typical nuclear mosaic gene.

Amino Acid Sequence↗