11 results on '"Rique H"'
Search Results
2. Genomic organization and characterization of splice variants of the human histamine H3 receptor
- Author
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Cogé, F, Guénin, S P, Audinot, V, Renouard-Try, A, Beauverger, P, Macia, C, Ouvry, C, Nagel, N, Rique, H, Boutin, J A, and Galizzi, J P
- Subjects
DNA, Complementary ,Base Sequence ,Sequence Homology, Amino Acid ,Molecular Sequence Data ,CHO Cells ,Cell Biology ,Sulfur Radioisotopes ,Biochemistry ,Alternative Splicing ,Microscopy, Fluorescence ,Guanosine 5'-O-(3-Thiotriphosphate) ,Cricetinae ,Animals ,Humans ,Receptors, Histamine H3 ,Calcium ,Amino Acid Sequence ,Cloning, Molecular ,Molecular Biology ,Research Article ,Protein Binding - Abstract
In the present paper we report the genomic organization of the human histamine H3-receptor gene, which consists of four exons spanning 5.5 kb on chromosome 20. Using PCR, six alternative splice variants of the H3 receptor were cloned from human thalamus. These variants were found to be coexpressed in human brain, but their relative distribution varied in a region-specific manner. These isoforms displayed either a deletion in the putative second transmembrane domain (TM), H3(DeltaTM2, 431aa) or a variable deletion in the third intracellular loop (i3), H3(Deltai3, 415aa), H3(Deltai3, 365aa), H3(Deltai3, 329aa) and H3(DeltaTM5+Deltai3, 326aa). In order to determine the biological role of the H3 receptor variants compared with the 'original' H3(445aa) receptor, three isoforms, namely H3(445aa), H3(DeltaTM2, 431aa) and H3(Deltai3, 365aa), were expressed in CHO cells and their pharmacological properties were investigated. Binding studies showed that H3(DeltaTM2, 431aa) transiently expressed in CHO cells was unable to bind [125I]iodoproxyfan, whereas both the H3(445aa) and H3(Deltai3, 365aa) receptors displayed a high affinity for [125I]iodoproxyfan [K(d)=28+/-5 pM (n=4) and 8+/-1 pM (n=5) respectively]. In addition, H3(Deltai3, 365aa) possessed the same pharmacological profile as the H3(445aa) receptor. However, in CHO cells expressing H3(Deltai3, 365aa), H3 agonists did not inhibit forskolin-induced cAMP production, stimulate [35S]guanosine 5'-[gamma-thio]triphosphate ([35S]GTP[S]) binding or stimulate intracellular Ca(2+) mobilization. Therefore the 80-amino-acid sequence located at the C-terminal portion of i3 plays an essential role in H3 agonist-mediated signal transduction. The existence of multiple H3 isoforms with different signal transduction capabilities suggests that H3-mediated biological functions might be tightly regulated through alternative splicing mechanisms.
- Published
- 2001
3. Cloning and molecular characterization of the novel human melanin-concentrating hormone receptor MCH2
- Author
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Rodriguez M, Beauverger P, Naime I, Rique H, Ouvry C, Souchaud S, Dromaint S, Nagel N, Suply T, Audinot V, Jean Albert Boutin, and Jp, Galizzi
- Subjects
DNA, Complementary ,Base Sequence ,Sequence Homology, Amino Acid ,Molecular Sequence Data ,Chromosome Mapping ,Receptors, G-Protein-Coupled ,Humans ,Chromosomes, Human, Pair 6 ,Tissue Distribution ,Amino Acid Sequence ,RNA, Messenger ,Receptors, Pituitary Hormone ,Receptors, Somatostatin ,Cloning, Molecular ,Peptides - Abstract
Using a genomics-based approach for screening orphan G-protein-coupled receptors, we have identified and cloned a novel high-affinity, melanin-concentrating hormone (MCH) receptor. This receptor, named S643b, displays the greatest overall identity (32%) with the previously reported human SLC-1 receptor (MCH1) and to a lesser extent with the somatostatin receptor subtypes. The gene encoding the S643b receptor spans more than 23 kilobase pairs (kb) and was mapped, by radiation hybrid experiments, on chromosome 6q14.3-q15. Comparison of the S643b cDNA with human genomic sequence reveals that the 340-amino-acid receptor is encoded by five exons. Its tissue distribution, as determined by Northern blot and reverse transcription-polymerase chain reaction analysis, indicates that a 4-kb transcript is predominantly expressed in the brain. When expressed in Chinese hamster ovary (CHO) cells, the S643b receptor displays a strong, dose-dependent, transient elevation of intracellular calcium in response to MCH (EC(50) = 9.5 nM). During the present study, we isolated a splice variant, designated S643a, encoding for a receptor that was not activated by MCH in a cellular calcium mobilization assay. Comparative pharmacological studies using CHO cells stably expressing either SLC-1 or S643b receptors demonstrated that similar structural features of MCH are required to stimulate intracellular Ca(2+) mobilization at both receptors. The identification and localization of this new MCH receptor (MCH2) provides further insight into the physiological implication of MCH in modulating behavioral responses, including food intake.
- Published
- 2001
4. Characterisation of cyclooxygenase 1 and 2 expression in mouse resident peritoneal macrophages in vitro; interactions of non steroidal anti-inflammatory drugs with COX2
- Author
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Michael Spedding, Andre N, Coge F, Rique H, Tordjman C, and Bonnet J
- Subjects
Male ,Lipopolysaccharide ,Molecular Sequence Data ,Biophysics ,Prostaglandin ,Inflammation ,macromolecular substances ,Pharmacology ,Biochemistry ,Polymerase Chain Reaction ,Dinoprostone ,Electron Transport Complex IV ,chemistry.chemical_compound ,Mice ,Endocrinology ,In vivo ,medicine ,Animals ,Drug Interactions ,RNA, Messenger ,Cells, Cultured ,biology ,Base Sequence ,L-Lactate Dehydrogenase ,Zymosan ,Anti-Inflammatory Agents, Non-Steroidal ,food and beverages ,In vitro ,chemistry ,biology.protein ,Macrophages, Peritoneal ,Arachidonic acid ,Cyclooxygenase ,medicine.symptom - Abstract
Resident peritoneal macrophages exposed to inflammatory stimuli (zymosan, lipopolysaccharide (LPS)) represent a widely used model for studying arachidonic acid metabolism and for screening of prostaglandin (PG) synthesis inhibitors. In the present study, cyclooxygenase 1 (COX1) was shown constitutively expressed in mouse adherent and non-adherent macrophages whereas expression of COX2 was observed only in adherent cells, even when cultured in minimal conditions (Ca-, Mg- and serum-free medium). The COX2 expression was amplified by arachidonic acid cascade stimulating agents (Ca, Mg, zymosan) and by LPS in a time-dependant manner; PGE2 by itself amplified LPS-induced COX2 expression. In well-defined experimental conditions of COX2 expression (LPS-stimulated adherent macrophages), we studied specific interactions of some representative anti-inflammatory drugs with COX2 enzymatic activity and expression. By contrast with dexamethasone, which reduced PGE2 release together with a strong reduction of COX2 expression (protein and mRNA), non steroidal anti-inflammatory drugs (NSAIDs) reduced PGE2 synthesis without any effect at the COX2 mRNA level. This reduction of PGE2 production by NSAIDs resulted from either an exclusive enzymatic inhibition (aspirin, NS398, 6-Methoxy naphtyl acetic acid) or an enzymatic inhibition associated with a slight decrease of COX2 protein level (indomethacin). For paracetamol and salicylic acid, two weak inhibitors of COX enzymatic activity, reduction of PGE2 synthesis appeared to be related to reduced level of COX2. These findings show that the macrophage can be used as a cellular model to study specifically COXI and COX2. In this cell type, COX2 expression is dependent on adhesion, enhanced by stimulation of arachidonic acid metabolism, and auto amplified by PGE2. Furthermore, the results indicate that known NSAIDs differ in their interaction with cyclooxygenase, being able to inhibit either COX2 enzymatic activity, and/or COX2 expression. However, further studies are required to determine the mechanism and the role of COX2 expression during inflammation in vivo, and to define more precisely the best target for new potent and safe NSAIDs.
- Published
- 1995
5. Characterisation of cyclooxygenase 1 and 2 expression in mouse resident peritoneal macrophages in vitro; interactions of non steroidal anti-inflammatory drugs with COX2
- Author
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Tordiman, C., primary, Coge, F., additional, Andre, N., additional, Rique, H., additional, Spedding, M., additional, and Bonnet, J., additional
- Published
- 1995
- Full Text
- View/download PDF
6. Structure-activity relationship studies of melanin-concentrating hormone (MCH)-related peptide ligands at SLC-1, the human MCH receptor.
- Author
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Audinot, V, Beauverger, P, Lahaye, C, Suply, T, Rodriguez, M, Ouvry, C, Lamamy, V, Imbert, J, Rique, H, Nahon, J L, Galizzi, J P, Canet, E, Levens, N, Fauchere, J L, and Boutin, J A
- Abstract
Melanin-concentrating hormone (MCH) is a cyclic nonadecapeptide involved in the regulation of feeding behavior, which acts through a G protein-coupled receptor (SLC-1) inhibiting adenylcyclase activity. In this study, 57 analogues of MCH were investigated on the recently cloned human MCH receptor stably expressed in HEK293 cells, on both the inhibition of forskolin-stimulated cAMP production and guanosine-5'-O-(3-[(35)S]thiotriphosphate ([(35)S]- GTPgammaS) binding. The dodecapeptide MCH-(6-17) (MCH ring between Cys(7) and Cys(16), with a single extra amino acid at the N terminus (Arg(6)) and at the C terminus (Trp(17))) was found to be the minimal sequence required for a full and potent agonistic response on cAMP formation and [(35)S]- GTPgammaS binding. We Ala-scanned this dodecapeptide and found that only 3 of 8 amino acids of the ring, namely Met(8), Arg(11), and Tyr(13), were essential to elicit full and potent responses in both tests. Deletions inside the ring led either to inactivity or to poor antagonists with potencies in the micromolar range. Cys(7) and Cys(16) were substituted by Asp and Lys or one of their analogues, in an attempt to replace the disulfide bridge by an amide bond. However, those modifications were deleterious for agonistic activity. In [(35)S]- GTPgammaS binding, these compounds behaved as weak antagonists (K(B) 1-4 microm). Finally, substitution in MCH-(6-17) of 6 out of 12 amino acids by non-natural residues and concomitant replacement of the disulfide bond by an amide bond led to three compounds with potent antagonistic properties (K(B) = 0.1-0.2 microm). Exploitation of these structure-activity relationships should open the way to the design of short and stable MCH peptide antagonists.
- Published
- 2001
- Full Text
- View/download PDF
7. Molecular identification of the long isoform of the human neuropeptide Y Y5 receptor and pharmacological comparison with the short Y5 receptor isoform.
- Author
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Rodriguez M, Audinot V, Dromaint S, Macia C, Lamamy V, Beauverger P, Rique H, Imbert J, Nicolas JP, Boutin JA, and Galizzi JP
- Subjects
- Alternative Splicing, Animals, Arginine pharmacology, Base Sequence, Brain metabolism, COS Cells drug effects, Cloning, Molecular, Codon, Initiator, Humans, Molecular Sequence Data, Neuropeptide Y metabolism, Neuropeptide Y pharmacology, Protein Biosynthesis, Protein Isoforms drug effects, Protein Isoforms genetics, RNA, Messenger metabolism, Receptors, Neuropeptide Y antagonists & inhibitors, Receptors, Neuropeptide Y metabolism, Transcription, Genetic, Trinucleotide Repeats genetics, 5' Untranslated Regions, Arginine analogs & derivatives, Receptors, Neuropeptide Y drug effects, Receptors, Neuropeptide Y genetics
- Abstract
The neuropeptide Y Y5 receptor gene generates two splice variants, referred to here as Y5(L) (long isoform) and Y5(S) (short isoform). Y5(L) mRNA differs from Y5(S) mRNA in its 5' end, generating a putative open reading frame with 30 additional nucleotides upstream of the initiator AUG compared with the Y5(S) mRNA. The purpose of the present work was to investigate the existence of the Y5(L) mRNA. The authenticity of this transcript was confirmed by isolating part of its 5' untranslated region through 5' rapid amplification of cDNA ends and analysing its tissue distribution. To study the initiation of translation on Y5(L) mRNA, we cloned the Y5(L) cDNA and two Y5(L) cDNA mutants lacking the first or the second putative initiation start codon. Transient expression of the three plasmids in COS-7 cells and saturation binding experiments using (125)I-labelled polypeptide YY (PYY) as a ligand showed that initiation of translation on Y5(L) mRNA could start at the first AUG, giving rise to a Y5(L) receptor with an N-terminal 10-amino-acid extension when compared with the Y5(S) receptor. The human Y5(L) and Y5(S) receptor isoforms displayed similar affinity constants (1.3 nM and 1.5 nM respectively). [(125)I]PYY binding to COS-7 cells expressing either the Y5(L) or the Y5(S) isoform was inhibited with the same rank order of potency by a selection of six chemically diverse compounds: PYY>neuropeptide Y>pancreatic polypeptide>CGP71683A>Synaptic 34>Banyu 6. Comparison of the tissue distribution of Y5(L) and Y5(S) mRNAs, as determined by reverse transcription-PCR analysis, indicated that expression of Y5(L) mRNA occurs in a tissue-specific manner. Finally, we have shown that the two AUG triplets contained in the 5' untranslated region of Y5(L) mRNA did not affect receptor expression.
- Published
- 2003
- Full Text
- View/download PDF
8. Structure and expression of the human histamine H4-receptor gene.
- Author
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Cogé F, Guénin SP, Rique H, Boutin JA, and Galizzi JP
- Subjects
- 3' Untranslated Regions genetics, 5' Untranslated Regions genetics, Base Sequence, Blotting, Southern, Brain metabolism, Genomic Library, Humans, Leukocytes metabolism, Liver metabolism, Lung metabolism, Molecular Sequence Data, Organ Specificity, Promoter Regions, Genetic genetics, RNA, Messenger biosynthesis, RNA, Messenger genetics, Radiation Hybrid Mapping, Receptors, Histamine H4, Reverse Transcriptase Polymerase Chain Reaction, Sequence Analysis, DNA, Spleen metabolism, Physical Chromosome Mapping, Receptors, G-Protein-Coupled, Receptors, Histamine genetics
- Abstract
We report the characterization by genomics-based approach of the human H4-receptor gene structure. The H4-receptor gene have been mapped by radiation hybrid experiments (Gene Bridge 4) on chromosome 18q11.2, between the AFMBB11WH5 and CHLC.GATA85D10 markers. The H4-receptor gene spans more than 21 kbp and contains three exons separated by two large introns (>7 kbp). RT-PCR analysis showed that the H4-receptor gene encoded a 3.7 kb mRNA which did not seem to be alternatively spliced within its coding region. The H4-receptor transcripts were found to be highly expressed in peripheral tissues implicated in inflammatory responses such as leukocytes, spleen, lung, and liver. In addition, low expression level of the H4-receptor mRNA was also detected in several human brain regions. Analysis of the 5'-flanking region of the H4-receptor gene did not reveal the existence of canonical TATA or CAAT-box. However, several putative regulatory elements mediating TNFalpha or IL-6-stimulated transcriptional activation were detected. The uteroglobin promoter binding factor, known to mediate anti-inflammatory response of uteroglobin, in the lung, was also found in this region. Thus, the description of the H4-receptor gene promoter region will facilitate the elucidation of its transcriptional control by factors secreted during inflammatory responses., (Copyright 2001 Academic Press.)
- Published
- 2001
- Full Text
- View/download PDF
9. Truncated isoforms inhibit [3H]prazosin binding and cellular trafficking of native human alpha1A-adrenoceptors.
- Author
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Cogé F, Guenin SP, Renouard-Try A, Rique H, Ouvry C, Fabry N, Beauverger P, Nicolas JP, Galizzi JP, Boutin JA, and Canet E
- Subjects
- Amino Acid Sequence, Animals, Biological Transport, Blotting, Western, COS Cells, Cloning, Molecular, DNA, Complementary, Humans, Liver metabolism, Molecular Sequence Data, Prazosin metabolism, Protein Binding, Protein Isoforms chemistry, Protein Isoforms genetics, Radioligand Assay, Receptors, Adrenergic, alpha-1 chemistry, Receptors, Adrenergic, alpha-1 genetics, Sequence Homology, Amino Acid, Subcellular Fractions metabolism, Tritium, Prazosin antagonists & inhibitors, Protein Isoforms metabolism, Receptors, Adrenergic, alpha-1 metabolism
- Abstract
We have identified from human liver eight alpha(1A)-adrenoceptor (alpha(1A)-AR) splice variants that were also expressed in human heart, prostate and hippocampus. Three of these alpha(1A)-AR isoforms (alpha(1A-1)-AR, alpha(1A-2a)-AR and alpha(1A-3a)-AR) gave rise to receptors with seven transmembrane domains (7TMalpha(1A)-AR). The other five (alpha(1A-2b)-AR, alpha(1A-2c)-AR, alpha(1A-3c)-AR, alpha(1A-5)-AR and alpha(1A-6)-AR) led to truncated receptors lacking transmembrane domain VII (6TMalpha(1A)-AR). The 7TMalpha(1A)-AR isoforms transiently expressed in COS-7 cells bound [(3)H]prazosin with high affinity (K(d) 0.2 nM) and mediated a noradrenaline (norepinephrine)-induced increase in cytoplasmic free Ca(2+) concentration, whereas the 6TMalpha(1A)-AR isoforms were incapable of ligand binding and signal transduction. Immunocytochemical studies with N-terminal epitope-tagged alpha(1A)-AR isoforms showed that the 7TMalpha(1A)-AR isoforms were present both at the cell surface and in intracellular compartments, whereas the 6TMalpha(1A)-AR isoforms were exclusively localized within the cell. Interestingly, in co-transfected cells, each truncated alpha(1A)-AR isoform inhibited [(3)H]prazosin binding and cell-surface trafficking of the co-expressed 'original' 7TMalpha(1A-1)-AR. However, there was no modification of either the [(3)H]prazosin-binding affinity or the pharmacological properties of alpha(1A-1)-AR. Immunoblotting experiments revealed that co-expression of the alpha(1A-1)-AR with 6TMalpha(1A)-AR isoforms did not impair alpha(1A-1)-AR expression. Therefore the expression in human tissues of many truncated isoforms constitutes a new regulation pathway of biological properties of alpha(1A)-AR.
- Published
- 1999
10. Cloning and characterization of the 5' flanking region of the human uncoupling protein 3 (UCP3) gene.
- Author
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Acín A, Rodriguez M, Rique H, Canet E, Boutin JA, and Galizzi JP
- Subjects
- Base Sequence, Blotting, Northern, Cloning, Molecular, DNA, Complementary, Humans, Ion Channels, Mitochondrial Proteins, Molecular Sequence Data, RNA, Messenger genetics, Reverse Transcriptase Polymerase Chain Reaction, Transcription, Genetic, Uncoupling Protein 3, Carrier Proteins genetics
- Abstract
Uncoupling protein 3 (UCP3), a member of the UCP family, mainly expressed in skeletal muscle could be responsible for thermogenesis in humans. Since little is known about its regulation, we studied the 5' flanking region of the human UCP3 (hUCP3) gene, which potentially contains the promoter sequences. We report the hUCP3 transcription initiation on a G located 764 nucleotides upstream the A contained in the first translated codon. Therefore, hUCP3 first exon has 669 bases of untranslated sequence. We also report the cloning and sequencing of seven kilobases from the gene 5' end and analyze the features of the potential proximal promoter. The MyoD family binding motif, called E-box, is the most abundant on this region. Other muscle-specific motives present in the potential proximal promoter include a MEF2 site as well as binding sequences for ubiquitous factors such as GC box and two CAAT boxes. Additionally, three putative peroxisome proliferator and one thyroid hormone response elements (PPRE and TRE, respectively) are found, which suggest a potential role for the peroxisome proliferator-activated receptor (PPAR) and thyroid hormone in human UCP3 gene expression. The description of the promoter region of the UCP3 gene will facilitate the elucidation of its transcriptional control., (Copyright 1999 Academic Press.)
- Published
- 1999
- Full Text
- View/download PDF
11. Characterisation of cyclooxygenase 1 and 2 expression in mouse resident peritoneal macrophages in vitro; interactions of non steroidal anti-inflammatory drugs with COX2.
- Author
-
Tordjman C, Coge F, Andre N, Rique H, Spedding M, and Bonnet J
- Subjects
- Animals, Anti-Inflammatory Agents, Non-Steroidal chemistry, Base Sequence, Cells, Cultured, Dinoprostone analysis, Drug Interactions, Electron Transport Complex IV antagonists & inhibitors, Electron Transport Complex IV genetics, L-Lactate Dehydrogenase analysis, Male, Mice, Molecular Sequence Data, Polymerase Chain Reaction, RNA, Messenger analysis, Anti-Inflammatory Agents, Non-Steroidal pharmacology, Electron Transport Complex IV biosynthesis, Macrophages, Peritoneal enzymology
- Abstract
Resident peritoneal macrophages exposed to inflammatory stimuli (zymosan, lipopolysaccharide (LPS)) represent a widely used model for studying arachidonic acid metabolism and for screening of prostaglandin (PG) synthesis inhibitors. In the present study, cyclooxygenase 1 (COX1) was shown constitutively expressed in mouse adherent and non-adherent macrophages whereas expression of COX2 was observed only in adherent cells, even when cultured in minimal conditions (Ca-, Mg- and serum-free medium). The COX2 expression was amplified by arachidonic acid cascade stimulating agents (Ca, Mg, zymosan) and by LPS in a time-dependant manner; PGE2 by itself amplified LPS-induced COX2 expression. In well-defined experimental conditions of COX2 expression (LPS-stimulated adherent macrophages), we studied specific interactions of some representative anti-inflammatory drugs with COX2 enzymatic activity and expression. By contrast with dexamethasone, which reduced PGE2 release together with a strong reduction of COX2 expression (protein and mRNA), non steroidal anti-inflammatory drugs (NSAIDs) reduced PGE2 synthesis without any effect at the COX2 mRNA level. This reduction of PGE2 production by NSAIDs resulted from either an exclusive enzymatic inhibition (aspirin, NS398, 6-Methoxy naphtyl acetic acid) or an enzymatic inhibition associated with a slight decrease of COX2 protein level (indomethacin). For paracetamol and salicylic acid, two weak inhibitors of COX enzymatic activity, reduction of PGE2 synthesis appeared to be related to reduced level of COX2. These findings show that the macrophage can be used as a cellular model to study specifically COX1 and COX2. In this cell type, COX2 expression is dependent on adhesion, enhanced by stimulation of arachidonic acid metabolism, and auto amplified by PGE2. Furthermore, the results indicate that known NSAIDs differ in their interaction with cyclooxygenase, being able to inhibit either COX2 enzymatic activity, and/or COX2 expression. However, further studies are required to determine the mechanism and the role of COX2 expression during inflammation in vivo, and to define more precisely the best target for new potent and safe NSAIDs.
- Published
- 1995
- Full Text
- View/download PDF
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