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J Bourguignon

Publications and source records attributed to J Bourguignon.

At least 55 records · Page 3Linked to original sources
PubMed1994

Detection by the polymerase chain reaction of two polymorphisms in exon 14 of the human inter-alpha-trypsin inhibitor heavy chain H1 gene.

Two polymorphisms were detected within exon 14 of the inter-alpha-trypsin inhibitor heavy chain H1 (ITIH1) gene. The polymorphisms are detected by digesting the same 202-bp polymerase reaction product with the PstI and Hph1 restriction endonucleases. These gene polymorphisms lead to the change of two amino acids in the mature protein. The polymorphisms can be used for the analysis of 3p21 deletion in human carcinomas as well as to develop a better understanding of the protein polymorphism already described.

Alpha-Globulins↗
PubMed1994

The polymorphism of the plasma inter-alpha-trypsin inhibitor (ITI) and its relationship to the heavy chain H1 subunit gene (ITIH1) at 3p211-212.

We investigated the ITI protein polymorphism in linkage analysis, using DraI and SstI as restriction fragment length polymorphism (RFLP) markers for the ITIH1 gene. Isoelectric focusing (IEF) classification from 76 individual plasma samples and RFLP analysis from the corresponding DNA preparations disclosed linkage disequilibrium between the phenotypic IEF patterns of the two common ITI alleles, ITI*1 and ITI*2, and the diallelic DNA polymorphisms of two ITIH1 RFLPs, represented by DraI 4.0 kb and DraI 2.4 + 1.6 kb, and by SstI 6.7 kb and SstI 6.0 + 0.7 kb, for the ITI 1 and ITI 2 IEF phenotypes, respectively, and by DraI 4.0/2.4 + 1.6 kb and SstI 6.7/6.0 + 0.7 kb for the heterozygous ITI 1-2 IEF phenotype. Linked segregation between either of the RFLPs and the polymorphic ITI plasma protein locus has been established in nine informative family pedigrees. The less frequent allele in Europeans, ITI*3, is not represented by a further allelic restriction fragment in either RFLP. The significant linkage disequilibrium observed in this genetic study indicates that the ITI locus, with the alleles ITI*1 and ITI*2, must be close to, or reside within, the ITIH1 gene. The diallelic ITI protein polymorphism therefore provides an informative phenotypic marker system for chromosome 3p211-212.

Alleles↗
PubMed1994

Light microscopical detection of inter-alpha-trypsin inhibitor and its different mRNAs in cultured hepatoma Hep G2 cells using immunocytochemical and in situ hybridization techniques.

The Hep G2 hepatoma cell line synthesizes the inter-alpha-trypsin inhibitor (ITI). This protease inhibitor and the other proteins of this family include four polypeptides chains: three heavy chains (HC1, HC2, HC3) and one light chain (bikunin). In the present study, we have demonstrated by immunofluorescence that ITI is detected mainly in perinuclear cytoplasmic zones comparable to those of albumin or alpha-1-antitrypsin. The presence of the mRNAs of the four polypeptide chains in all Hep G2 cells of a non-synchronized culture have been demonstrated by in situ hybridization. An evaluation of the transcription of the four ITI genes through an analysis of markings brings to the fore a clearly much higher rate of mRNAs from the light chain than from the heavy chains. The mRNAs corresponding to the HC2 chains are more heavily represented than are those corresponding to the HC1 and HC3 chains. In Hep G2 cells in culture, a quantification of mRNAs based on the in situ hybridization technique shows that their relative quantities, in decreasing order, are those of L, HC2, HC3 and HC1.

Alpha-Globulins↗
PubMed1993

Isolation and characterization of the human inter-alpha-trypsin inhibitor heavy-chain H1 gene.

The human inter-alpha-trypsin inhibitor heavy chain H1 (ITI heavy chain H1) gene was isolated from two overlapping clones. It spans 14 kbp and is composed of 22 exons from 15 bp to 281 bp in size and has consensus splice sites. Intron sizes range from 80 bp to 2000 bp. It codes for the precursor of HC1 that is part of the serum ITI form of 220 kDa. Two major transcriptional initiation sites were identified in the 5'-flanking region, which contained putative promoter elements, but no typical TATA and CAAT boxes. mRNA for the ITI heavy chain H1 was found only in liver. The tissue-specific transcription of the gene might be due to the presence of binding sites for the hepatocyte nuclear factor HNF-5 and to the octameric motifs. A previous overlapping of cDNA clones indicated the absence of 29 bp in one of these clones. The present study shows that the 29 bp is located within the gene at the end of exon 21. A reverse-transcriptase polymerase chain reaction mapping analysis of liver mRNA identified the two types of the mRNA for ITI heavy chain H1. Accordingly, the data demonstrate that there is alternative splicing of at least one exon of the ITI heavy chain H1 gene.

Alpha-Globulins↗
PubMed1993

Glycine decarboxylase complex from higher plants. Molecular cloning, tissue distribution and mass spectrometry analyses of the T protein.

cDNA clones encoding the precursor of the T protein of the glycine decarboxylase complex have been isolated from a pea leaf cDNA library in lambda gt11. The longest cDNA insert of 1430 bp encodes a polypeptide of 408 amino acid residues of which 30 residues constitute an N-terminal cleavable presequence and 378 residues make up the mature protein. Several results confirmed the identity of the cDNA and the exactness of the predicted primary structure. Firstly, we purified the T protein to homogeneity and its mass was measured by mass spectrometry. The mass obtained (40966 +/- 5 Da) was the value predicted from the cDNA (40961 Da). Secondly, the purified T protein was chemically cleaved with cyanogen bromide and the peptide fragments were analysed by high-performance liquid chromatography/electrospray ionization mass spectrometry and/or fast-atom-bombardment mass spectrometry. The mass values of all the peptides generated by chemical cleavage and measured by these techniques were very close to the values calculated from the predicted primary structure. Thirdly, microsequencing of some of these peptides, which represent 35% of the total protein, fits perfectly with the primary structure deduced from the cDNA. In the present HPLC/electrospray ionization MS studies we never detected the presence of covalently bound tetrahydropteroylpolyglutamate (H4PteGlun), either in the native T protein or in the different peptide fragments generated by the chemical cleavage. The absence of H4PteGlun bound to the T protein in our experimental conditions demonstrates that H4PteGlun is not covalently linked to the T protein. Northern blot analysis showed that the steady-state level of the mRNA corresponding to the T protein was high in green leaves compared to the level in etiolated leaves (approximately 8-10-fold higher). Surprisingly, a non-negligible amount of mRNA corresponding to the T protein was present in roots whereas the mRNA encoding the H protein was not detectable. Western blot analysis showed that the P, L and T proteins of the glycine decarboxylase complex were present in roots whereas the H protein was not detectable. Southern hybridization to pea genomic DNA indicated the presence of a single gene encoding the T protein of the glycine decarboxylase complex in the haploid genome.

Amino Acid Sequence↗
PubMed1993

Human pre-alpha-trypsin inhibitor-precursor heavy chain. cDNA and deduced amino-acid sequence.

Pre-alpha-trypsin inhibitor (P alpha I) is a serine-proteinase inhibitor of M(r) 130,000 found in human serum. This protein belongs to the family of proteins called inter-alpha-trypsin inhibitor (ITI). P alpha I is composed of a heavy chain (HC3) and of a light chain (bikunin), synthesized by two separate mRNA. Bikunin is identical to the ITI light chain, the structure of which has already been established. The HC3 is obtained from a precursor called H3. The bikunin is covalently linked to HC3 by a chondroitin-4-sulfate glycosaminoglycan. We report here the H3 full-length cDNA sequence and the deduced amino-acid sequence of the heavy-chain H3 precursor. The high degree of similarity between the nucleotide and amino-acid sequences of ITI heavy-chain families H1, H2, H3 is examined with respect to their probable structure and assembly with bikunin in the final proteins, P alpha I and ITI.

Amino Acid Sequence↗
PubMed1992

Cloning of the gene (gdcH) encoding H-protein, a component of the glycine decarboxylase complex of pea (Pisum sativum L.).

H-protein is the lipoyl-protein component of the glycine decarboxylase complex, which catalyses, with serine hydroxymethyltransferase, the mitochondrial step of photorespiration in plants. We have isolated and characterized the gene (gdcH) encoding the H-protein in pea (Pisum sativum L.). The H-protein gene is distributed in a stretch of about 1.55 kbp and contains three introns (75, 64 and 185 bp) located in the coding region. No intervening sequences were detected in the 5' and 3' non-coding regions. This intron-exon structure contrasts with the preliminary H-protein gene structures reported for human and chicken, where these genes (dispersed on 13 and 8 kbp genomic fragments respectively) are composed of five highly conserved exons and are interrupted by four long introns. Two main transcription sites were detected by primer extension of RNA. The first transcriptional initiation site was assigned the +1 position and correlated with a putative TATA box located at position -26. The second transcriptional start site was not correlated with a putative TATA box, but may be regulated by an 'initiator' element described by Smale & Baltimore [(1989) Cell (Cambridge, Mass.) 57, 103-113] which contains, within itself, the transcription start site. The presence of two potential promoters may be related to the specialized overexpression pattern of H-protein in leaves, in order to support photorespiration.

Amino Acid Oxidoreductases↗
PubMed1992

Human inter-alpha-trypsin inhibitor: full-length cDNA sequence of the heavy chain H1.

Inter-alpha-trypsin inhibitor (ITI), called inter-alpha-inhibitor, is a 220 kDa serine proteinase inhibitor found in human serum. It is composed of at least three distinct polypeptide chains. These chains, named H1, H2 and L, are an independently synthesized and proteolytically processed precursor protein. Only the complete structure of H2 and L has been established so far. We used a PCR-based cloning approach and a cDNA screening library to isolate the full-length cDNA H1. The amino acid sequences of the two heavy chains deduced from the cDNA are highly similar (40% identity). Nevertheless, the structure of the signal peptide and propeptide in the N-terminal region is different in these two chains. A complex posttranslational cleavage at both ends of H1 and H2 may be proposed prior to assembly of the ITI chains.

Alpha-Globulins↗
PubMed1992

Isolation, characterization, and sequence analysis of a cDNA clone encoding L-protein, the dihydrolipoamide dehydrogenase component of the glycine cleavage system from pea-leaf mitochondria.

L-protein is the dihydrolipoamide dehydrogenase component of the glycine decarboxylase complex which catalyses, with serine hydroxymethyltransferase, the mitochondrial step of photorespiration. We have isolated and characterized a cDNA from a lambda gt11 pea library encoding the complete L-protein precursor. The derived amino acid sequence indicates that the protein precursor consists of 501 amino acid residues, including a presequence peptide of 31 amino acid residues. The N-terminal sequence of the first 18 amino acid residues of the purified L-protein confirms the identity of the cDNA. Alignment of the deduced amino acid sequence of L-protein with human, porcine and yeast dihydrolipoamide dehydrogenase sequences reveals high similarity (70% in each case), indicating that this enzyme is highly conserved. Most of the residues located in or near the active sites remain unchanged. The results described in the present paper strongly suggest that, in higher plants, a unique dihydrolipoamide dehydrogenase is a component of different mitochondrial enzyme complexes. Confidence in this conclusion comes from the following considerations. First, after fractionation of a matrix extract of pea-leaf mitochondria by gel-permeation chromatography followed by gel electrophoresis and Western-blot analysis, it was shown that polyclonal antibodies raised against the L-protein of the glycine-cleavage system recognized proteins with an Mr of about 60000 in different elution peaks where dihydrolipoamide dehydrogenase activity has been detected. Second, Northern-blot analysis of RNA from different tissues such as leaf, stem, root and seed, using L-protein cDNA as a probe, indicates that the mRNA of the dihydrolipoamide dehydrogenase accumulates to high levels in all tissues. In contrast, the H-protein (a specific protein component of the glycine-cleavage system) is known to be expressed primarily in leaves. Third, Southern-blot analysis indicated that the gene coding for L-protein in pea is most likely to be present in a single copy/haploid genome.

Amino Acid Sequence↗
PubMed1992

Stimulation by GH of IGF1 proforms synthesized by rabbit chondrocytes cultured with bFGF in serum-free medium.

The IR-IGF1 production by rabbit epiphyseal chondrocytes cultured in serum-free medium was analyzed. Cell proliferation was induced by the addition of 10 ng/ml basic fibroblast growth factor (bFGF) without or with 100 ng/ml recombinant human growth hormone (hGH). GH alone induced no cell multiplication. Chondrocytes treated with bFGF alone secreted an IR-IGF1 activity proportional to the mitotic activity of the cells. A specific positive IGF1 immunostaining was localized in the Golgi of control and hGH-treated cells. The IR-IGF1 activity recovered into culture medium was mainly composed of three fractions of apparent MW 6-8 kDa, 9-14 kDa, and 16-18 kDa. [35S]Methionine pulse-chase experiments indicated that the radiolabeled 16-18 kDa IR-IGF1 fraction was partly converted into the 9-14 kDa and 6-8 kDa fractions. At equilibrium, 70% of the chondrocyte IR-IGF1 activity was recovered as 9- to 18-kDa forms which contained high IR-proIGF1A activity. The 6-8 kDa fraction had biochemical characteristics similar to those of the mature IGF1 peptide. Similar results were observed when 4% fetal calf serum was added to the culture. The addition of 100 ng/ml of hGH significantly and specifically increased IGF1 precursor material, which thus represented 90% of total IR-IGF1 activity. On Day 16 of the culture, when cells stopped dividing, the amount of chondrocyte IR-IGF1 was significantly lower than during cell proliferation, and hGH had no effect on this production. These data indicate that cultured chondrocytes produce more IGF1 precursors than mature IGF1 and that GH specifically stimulates biosynthesis of IGF1 precursors but not IGF1 per se. A GH-dependent biological function of IGF1 proforms in chondrocytes remains to be demonstrated.

Animals↗
PubMed1990

Structural analysis of the human inter-alpha-trypsin inhibitor light-chain gene.

The human inter-alpha-trypsin inhibitor (ITI) light-chain gene, which codes for the two proteins alpha 1-microglobulin (protein HC) and ITI-derived human inhibitor of 30 kDa (HI-30), was isolated from a human genomic library. This gene, present as a single copy in the human genome, is composed of 10 exons and 9 introns distributed over 20 kbp. A single transcriptional initiation site was identified in the 5'-flanking region which contained promoter elements, but no typical TATA box. However a sequence equivalent to the TATA box is present on both sense and anti-sense strands in the 5'-flanking region of the first exon coding for HI-30. The exon-intron organization suggests that the regions coding for protein HC and other members of the lipocalin superfamily evolved from a common ancestral gene that is probably different from that coding for HI-30. These data suggest that two distinct ancestral genes could have existed and fused during evolution. Several direct and one inverted repeats are also found within this gene, as well as potential glucocorticoid-receptor binding sites.

Alpha-Globulins↗
PubMed1990

Interaction between the Component Enzymes of the Glycine Decarboxylase Multienzyme Complex.

The glycine decarboxylase multienzyme complex comprises about one-third of the soluble protein of the matrix of pea (Pisum sativum) leaf mitochondria where it exists at a concentration of approximately 130 milligrams protein/milliliter. Under these conditions the complex is stable with an approximate subunit ratio of 2 P-protein dimers:27 H-protein monomers:9 T-protein monomers:1 L-protein dimer. When the complex is diluted it tends to dissociate into its component enzymes. This prevents the purification of the intact complex by gel filtration or ultracentrifugation. In the dissociated state the H-protein acts as a mobile cosubstrate that commutes between the other three enzymes and shows typical substrate kinetics. When the complex is reformed, the H-protein no longer acts as a substrate but as an integrated part of the enzyme complex.

Journal Article↗
PubMed1990

In vitro insulin-like growth factor I interaction with cartilage cells derived from postnatal animals.

This paper reports data on the in vitro effects of insulin-like growth factor I (IGF-I) and basic fibroblast growth factor (bFGF) on the phenotypic expression of epiphyseal chondrocytes grown in serum-free (SF) culture medium. bFGF mostly stimulates chondrocyte DNA and inhibits sulfated proteoglycan synthesis and type II collagen mRNA. On the contrary, IGF-I is poorly mitogenic but strongly stimulates protein synthesis and type II collagen mRNA. In addition, IGF-I prevents the expression of type I collagen gene. Lastly, chondrocytes cultured in SF medium are able to locally produce IGF-I peptides. In conclusion, IGF-I and bFGF have opposite effects on the phenotypic expression of chondrocytes in vitro: bFGF is mostly mitogenic and IGF-I appears to be a differentiating factor.

Animals↗