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J C Meunier

Publications and source records attributed to J C Meunier.

At least 37 records · Page 2Linked to original sources
PubMed1999

Distinct mechanisms for activation of the opioid receptor-like 1 and kappa-opioid receptors by nociceptin and dynorphin A.

To understand how two structurally analogous ligand-receptor systems, the nociceptin/opioid receptor-like 1 (ORL1) and dynorphin A/kappa-opioid receptor 1 (KOR1) systems, achieve selectivity, receptor chimeras were generated and analyzed. Replacing discrete domains located between the N-terminus and top of the third transmembrane helix of the KOR1 by the homologous domains of the ORL1 receptor yields hybrid receptors, which, in comparison with the parent KOR1, display up to 300-fold increased affinity but low sensitivity toward nociceptin, and unaltered (high) affinity and sensitivity toward dynorphin A. These substitutions contribute elements for binding of nociceptin but do not suppress determinants necessary for binding and potency of dynorphin A. More importantly, further replacement in these chimeras of the second extracellular loop with that of the ORL1 receptor fully restores responsiveness to nociceptin without impairing responsiveness to dynorphin A. A bifunctional hybrid receptor has thus been identified that binds and responds to both nociceptin and dynorphin A as efficiently as the ORL1 receptor does to nociceptin and the KOR1 to dynorphin A. Together, these results suggest that distinct peptide activation mechanisms operate in the two receptor systems. In particular, the second extracellular receptor loop appears to be an absolute requirement for activation of the ORL1 receptor by nociceptin, but not for activation of the KOR1 by dynorphin A.

Animals↗
PubMed1999

The transmembrane domain of hepatitis C virus glycoprotein E1 is a signal for static retention in the endoplasmic reticulum.

Hepatitis C virus (HCV) glycoproteins E1 and E2 assemble to form a noncovalent heterodimer which, in the cell, accumulates in the endoplasmic reticulum (ER). Contrary to what is observed for proteins with a KDEL or a KKXX ER-targeting signal, the ER localization of the HCV glycoprotein complex is due to a static retention in this compartment rather than to its retrieval from the cis-Golgi region. A static retention in the ER is also observed when E2 is expressed in the absence of E1 or for a chimeric protein containing the ectodomain of CD4 in fusion with the transmembrane domain (TMD) of E2. Although they do not exclude the presence of an intracellular localization signal in E1, these data do suggest that the TMD of E2 is an ER retention signal for HCV glycoprotein complex. In this study chimeric proteins containing the ectodomain of CD4 or CD8 fused to the C-terminal hydrophobic sequence of E1 were shown to be localized in the ER, indicating that the TMD of E1 is also a signal for ER localization. In addition, these chimeric proteins were not processed by Golgi enzymes, indicating that the TMD of E1 is responsible for true retention in the ER, without recycling through the Golgi apparatus. Together, these data suggest that at least two signals (TMDs of E1 and E2) are involved in ER retention of the HCV glycoprotein complex.

Amino Acid Sequence↗
PubMed1999

[Quantitative angiography: update for peripheral vascular applications].

Visual estimation of vessel diameter is unreliable and observer-dependent. With digital angiography, software for accurate quantification of arterial diameter are available. They allow: (1) selection of equipment of appropriate size (balloons, stents, etc) during revascularization procedures, (2) objective evaluation of the results of such procedures, (3) comparison between follow-up measurements and initial measurements and (4) provide reproducible data in the case of multicenter studies. The widespread availability of digital systems has been accompanied by a dramatic increase in the number of software packages for quantification. The present article reviews some of the currently available methods for quantification, presents the advantages and pitfalls of each system, and outlines their limits.

Angiography↗
PubMed1999

[MRI imaging in cortical laminar necrosis].

PURPOSE: The goal of this study was to follow over time the MR imaging features of cortical laminar necrosis. PATIENTS AND METHODS: Six patients with cortical laminar necrosis were included. There were two women and four men aged 54-84 years, with a mean age of 68 years. In four patients, cortical laminar necrosis was caused by ischemic stroke, one case occurred after a cardiac arrest and the last patient had a meningoencephalitis. The time delay from insult to the first MR study varied between one week and 3 months. RESULTS: The MRI showed hyperintense lesions in the cerebral cortex on T1W and T2W images. The high intensity signal was still observed a few months after the insult. Cortical laminar necrosis lesions did not demonstrate hemorrhage on CT and MRI studies. CONCLUSION: MRI allowed detection of cortical laminar necrosis and could differentiate it from hemorrhage.

Aged↗
PubMed1998

[Phe1psi(CH2-NH)Gly2]nociceptin-(1-13)-NH2 is an agonist of the nociceptin (ORL1) receptor.

[Phe1psi(CH2-NH)Gly2]nociceptin-(1-13)-NH2, a pseudopeptide analog of nociceptin, has been shown to be a selective 'antagonist' of the nociceptin receptor in the isolated guinea pig ileum and mouse vas deferens preparations (Guerrini et al., 1998. Br. J. Pharmacol. 123, 163-165). However, in recombinant chinese hamster ovary cells expressing the human nociceptin receptor, we find that the pseudopeptide is a potent (IC50 = 7.5 nM) and fully efficacious inhibitor of forskolin-induced accumulation of cAMP, thus behaving as a pure 'agonist' rather than an antagonist of the receptor. The contrary behaviour of the pseudopeptide in smooth muscle and transformed cells may suggest that different nociceptin receptor types are being addressed in the two systems.

Animals↗
PubMed1998

Different domains of the ORL1 and kappa-opioid receptors are involved in recognition of nociceptin and dynorphin A.

In order to gain further insight into the functional architecture of structurally related G protein-coupled receptors, the ORL1 (nociceptin) and opioid receptors, we have constructed chimeras of ORL1 and mu-, delta- and kappa-opioid receptors, and compared their binding and functional properties with those of the parent receptors. We find in particular that a ORL1-kappa-opioid (O-K) hybrid construct has retained high affinity for non-type-selective opiate ligands, and has acquired the ability to bind and respond to enkephalins and mu- and/or delta-opioid receptor-selective enkephalins analogs, thus behaving like a 'universal' opioid receptor. Most significantly however, whilst the ORL1 and kappa-opioid receptors display high binding preference (KD 0.1 vs. 100 nM) for their respective endogenous ligands, nociceptin and dynorphin A, the O-K chimeric receptor binds both nociceptin and dynorphin A, with high affinity (KD < 1 nM). Together, these data (i) add weight to the hypothesis that the extracellular loops of opioid receptors act as a filter for ligand selection, and (ii) demonstrate that different domains of the ORL1 and kappa-opioid receptors are involved in recognition of their endogenous peptide ligands.

Animals↗
PubMed1998

Enzymatic phosphorylation of food proteins by purified and recombinant protein kinase CK2.

Protein kinase CK2 formerly called casein kinase II is a protein kinase able to phosphorylate more than 100 proteic substrates. We have purified protein kinase CK2 from the yeast Y. lipolytica to phosphorylate milk and plant reserve proteins to a significant extent. In the case of plant reserve proteins, which are polymeric substrates, not all subunits are substrate for protein kinase CK2, even if non phosphorylated subunits contain significant potent phosphorylations sites. Best substrates were soy beta-conglycinin (0.72 P/mol) and dephosphorylated caseins (0.5 P/mol). We have studied some functional properties of phosphorylated caseins. Solubility was improved for all pH values but pI. Sensitivity to calcium has also been assessed, and it is slightly improved upon phosphorylation. We have cloned the catalytic subunit of protein kinase CK2 from yeast Y. lipolytica. The recombinant catalytic subunit expressed in E. coli was active and displayed kinetic properties similar to those of the purified enzyme. The recombinant catalytic subunit was able to phosphorylate plant reserve proteins and milk proteins to a significant extent. Best substrates were soy beta-conglycinin (1.0 P/mol), and glycinin (0.59 P/mol).

Calcium↗
PubMed1998

Enzymatic phosphorylation of soy globulins by the protein kinase CK2. Determination of the phosphorylation sites of beta-conglycinin alpha subunit by mass spectrometry.

Beta-conglycinin alpha subunit has been phosphorylated using a cAMP-independent protein kinase (CK2) purified from the yeast Yarrowia lipolytica. CK2 is known to phosphorylate serines and threonines in the consensus sequence Ser/Thr-X-X-Asp/Glu. Only 0.5 to 1 mol P/mol alpha subunit was incorporated although seven consensus sequences are present. Phosphorylated beta-conglycinin alpha subunit (P-alpha) was digested by trypsin. The resulting peptides were analysed by RP-HPLC coupled to electrospray ionisation mass spectrometry (LC-ESMS). Two phosphopeptides were identified corresponding to 70-89 and 116-127 sequences with Ser 75 and Ser 117 phosphorylated respectively. Ser 75 is one of the predicted phosphorylation sites according to the consensus sequence criteria. Ser 117 is inside a very acidic peptide but does not belong to a previously described consensus sequence.

Antigens, Plant↗
PubMed1998

Expression and characterization of the recombinant catalytic subunit of casein kinase II from the yeast Yarrowia lipolytica in Escherichia coli.

The alpha catalytic subunit of casein kinase II from the yeast Yarrowia lipolytica has been cloned and overexpressed using the pT7-7 expression vector in Escherichia coli. Casein kinase activity is found in the bacterial extracts. The catalytic subunit is partially expressed in a soluble and active form, which is purified to electrophoretic homogeneity. Most of the enzyme was found in inclusion bodies. In this form, the enzyme, which is almost pure, exhibits a low specific activity. We have focused our efforts on methods to activate the protein from the inclusion bodies. We have studied the renaturation of urea-denaturated CKII catalytic subunit. We have tried different renaturation buffers and found that renaturation by dilution was more efficient than renaturation by dialysis. Treatment of the enzyme found in the inclusion bodies with different nondetergent sulfobetaines (NDSB) led to a time-dependent activation. NDSB195 is a V-type activator of the recombinant catalytic subunit of casein kinase II. The NDSB195-activated enzyme remained active at the room temperature for weeks. Kinetic properties of the recombinant casein kinase II subunit are similar to those of the purified holoenzyme (low Km for ATP and inhibition by heparin). Kinetic study indicates that the beta subunit could interact with the alpha subunit at the level of the catalytic site to enhance activity and to modify the kinetic behavior of the enzyme.

Base Sequence↗
PubMed1998

Characterization of an exocellular protein phosphatase with dual substrate specificity from the yeast Yarrowia lipolytica.

In previous work, the major endocellular protein phosphatase activity has been identified in the secretory yeast Yarrowia lipolytica as a PP2A. The aim of the present work was to seek the presence of one protein phosphatase excreted in the exocellular medium and to study its activity during yeast growth in media supplemented or not supplemented with inorganic phosphate. Protein phosphatase was purified and activity was assayed by following the dephosphorylation of three substrates, [32P]casein, phosphotyrosine and a synthetic tyrosine-phosphorylated peptide. Phosphatase activity recovered in the medium after 25 h culture was greatly enhanced by Pi-deficiency. After several purification steps, the enzyme preparation presents an apparent electrophoretic homogeneity on SDS-PAGE with associated phosphoseryl/threonyl and phosphotyrosyl activities. The kinetic properties exclude contamination by a copurified protein and it is concluded that the two activities are carried by the same single proteic species. It was characterized by gel filtration as a 33 kDa protein with one single subunit demonstrated by SDS-PAGE. An absolute requirement for reducing-agents is observed suggesting that the enzyme contains at least one essential reactive cysteinyl residue. Optimum pH value is 6.1, apparent K(m) for phosphotyrosine was calculated to be 760 microM and Hill coefficient 3.2 indicating a rather high cooperativity. These results showed that the involvement of alkaline and/or acid phosphatase was unlikely. In conclusion, a protein phosphatase distinct from endocellular PP2A is secreted by Yarrowia lipolytica and characterized as a phosphotyrosine protein phosphatase with associated phosphoseryl/threonyl activity.

Amino Acid Sequence↗
PubMed1998

Molecular modelling of the ORL1 receptor and its complex with nociceptin.

The opioid receptor like (ORL1) receptor is a G-protein coupled receptor superfamily, and regulates a plethora of neurophysiological functions. The structural requirements for receptor activation by its endogenous agonist, nociceptin (FGGFTGARKSARKLANQ), differ markedly from those of the kappa-opioid receptor and its putative peptide agonist, dynorphin A (YGGFLRRIRPKLKWDNQ). In order to probe the functional architecture of the ORL1 receptor, a molecular model of the receptor has been built, including the TM domain and the extra- and intracellular loops. An extended binding site able to accommodate nociceptin-(1-13), the shortest fully active analogue of nociceptin, has been characterized. The N-terminal FGGF tetrapeptide is proposed to bind in a highly conserved region, comprising two distinct hydrophobic pockets in a cavity formed by TM helices 3, 5, 6 and 7, capped by the acidic second extracellular (EL2) loop controlling access to the TM elements of the peptide binding site. The nociceptin conformation provides for the selective preference of the ORL1 receptor for nociceptin over dynorphin A, conferred by residue positions 5 and 6 (TG versus LR), and the favourable interaction of its highly positively charged core (residues 8-13) with the EL2 loop, thought to mediate receptor activation. The functional roles of the EL2 loop and the conserved N-terminal tetrapeptide opioid 'message' binding site are discussed in the context of the different structural requirements of the ORL1 and kappa-opioid receptors for activation.

Amino Acid Sequence↗
PubMed1998

Nociceptin-induced apparent hyperalgesia in mice as a result of the prevention of opioid autoanalgesic mechanisms triggered by the stress of an intracerebroventricular injection.

The effects on nociperception of nociceptin/Orphanin FQ (noc/OFQ), the endogenous ligand of the ORL1 (opioid receptor like 1) receptor, have been evaluated in mice upon intracerebroventricular injection of 10 to 10,000 ng doses of the peptide. In the hot plate test (55 degrees C) the licking, rearing and jump latencies were significantly reduced by noc/OFQ (100-250 ng). Noc/OFQ (100-1000 ng) also reduced the latency to tail withdrawal in the tail flick test. In the formalin test (injection in a hind paw of a formalin solution), noc/OFQ (100 ng) increased significantly the duration of paw licking and/or biting at the earliest period of observation. In the writhing test, the number of writhes evoked by intraperitoneal administration of dilute acetic acid was not modified by noc/OFQ at doses in the range of 10-1000 ng, but was decreased by 10,000 ng. The reduction in jump latency in the hot plate test was observed even when mice were pretreated with morphine (2 mg/kg, s.c.). The analgesic effect of acetorphan (5 mg/kg, i.v.) was also reduced by nociceptin (100 ng); on the other hand the hyperalgesic effect of naloxone (4.5 mg/kg, s.c.) was not additive with that of nociceptin (100 ng). Comparing in various tests the nociceptive thresholds of uninjected mice to that of saline i.c.v. injected mice, it appeared that the latter injection induced an increase in these thresholds which was prevented by nociceptin. It is suggested that nociceptin displays hyperalgesic effects by preventing autoanalgesic (opioidergic) mechanisms triggered by the stress elicited by intracerebroventricular injection.

Analgesia↗
PubMed1998

A retention signal necessary and sufficient for endoplasmic reticulum localization maps to the transmembrane domain of hepatitis C virus glycoprotein E2.

The hepatitis C virus (HCV) genome encodes two envelope glycoproteins (E1 and E2). These glycoproteins interact to formin a noncovalent heterodimeric complex which is retained in the endoplasmic reticulum (ER). To identify whether E1 and/or E2 contains an ER-targeting signal potentially involved in ER retention of the E1-E2 complex, these proteins were expressed alone and their intracellular localization was studied. Due to misfolding of E1 in the absence of E2, no conclusion on the localization of its native form could be drawn from the expression of E1 alone. E2 expressed in the absence of E1 was shown to be retained in the ER similarly to E1-E2 complex. Chimeric proteins in which E2 domains were exchanged with corresponding domains of a protein normally transported to the plasma membrane (CD4) were constructed to identify the sequence responsible for its ER retention. The transmembrane domain (TMD) of E2 (C-terminal 29 amino acids) was shown to be sufficient for retention of the ectodomain of CD4 in the ER compartment. Replacement of the E2 TMD by the anchor signal of CD4 or a glycosyl phosphatidylinositol (GPI) moiety led to its expression on the cell surface. In addition, replacement of the E2 TMD by the anchor signal of CD4 or a GPI moiety abolished the formation of E1-E2 complexes. Together, these results suggest that, besides having a role as a membrane anchor, the TMD of E2 is involved in both complex formation and intracellular localization.

Amino Acid Sequence↗
PubMed1998

[Cloning of prepronociceptin has led to the discovery of other biologically active peptides].

Among the opioid receptors family, the cloning of the mu, kappa and delta receptors was followed by that of another member, named ORL1 (Opiate Receptor Like 1). In spite of obvious homologies with the mu, kappa and delta receptors, ORL1 does not display a relevant affinity for the endogenous ligands of these former receptors (beta endorphin, enkephalins, dynorphin A...). This observation has prompted to search for an endogenous ligand of ORL1. A heptadecapeptide which fulfils this function, with a nanomolar affinity, has been found. It was named either nociceptin or orphanin FQ. It demonstrates, according either to the dose or to the route of administration, hyperalgesic, allodynic, antiopioidergic or even analgesic effects. It displays also many behavioural effects, modifying especially locomotion, exploratory behaviour, motivation, anxiety, memory, food intake. Nociceptin results from the cleavage of a large precursor protein, prepronociceptin (PPNOC). In this latter, nociceptin is flanked on its C-terminal region by another peptide which may be regarded either as a heptadecapeptide (NocII), or a bidecapeptide (NocIII) according to the inclusion or not of a fragment constituted by 3 arginine residues. Investigating the functions modulated by NocII, we observed that it stimulates locomotor activity of mice and shortens the forepaws licking latency in the hot plate test (55 degrees C); these effects are not shared by NocIII. The simultaneous administration of NocII and nociceptin resulted in animals put on the hot plate to the appearance of their respective effects, not modified by the presence of the other. A 41 amino acid peptide flanks nociceptin on its N-terminal region in PPNOC. It may be cleaved to generate a heptadecapeptide, named nocistatin on account of its antagonist effect on the hyperalgesia/allodynia induced by nociceptin. Thus, the discovery of ORL1 has led to that of nociceptin, that of its precursor PPNOC, and thereby to that of NocII/NocIII and nocistatin. The functions modulated by these peptides are being investigated whereas their receptors are yet unknown. These multiple targets allow to expect new strategies to modulate their functions.

Animals↗
PubMed1997

Nociceptin/orphanin FQ and the opioid receptor-like ORL1 receptor.

Homology cloning and, more recently, the sequencing of whole genomes, have identified many open reading frames encoding proteins of unknown function, in particular putative G protein-coupled membrane receptors. Identification of orphan receptors in this way has marked the advent of 'reverse pharmacology' to identify the corresponding physiological ligands. This approach has led to the discovery of the ORL1 (Opioid Receptor-Like 1) receptor, and of its natural ligand, nociceptin/orphanin FQ (noc/oFQ), the basic components of a new peptide-based signalling pathway in the nervous system. Based on genetic criteria, the ORL1 and opioid receptors belong to the same family, as do noc/oFQ and opioid peptides. The marked structural analogy between the ORLI and opioid receptors, especially the kappa-opioid receptor, and the noc/oFQ and opioid peptides, particularly dynorphin A, is not reflected anatomically since noc/oFQ and opioid peptides appear to be located in separate neuronal circuits. Noc/oFQ triggers the same G protein-mediated signalling pathways as do opioids, however, to produce pharmacological effects that sometimes differ from, and even oppose, those of opioids. Noc/oFQ stimulates an outward K+ current and/or inhibits voltage-gated Ca2+ channels, thereby reducing synaptic efficacy, i.e. neuronal activity. In the rat, noc/oFQ is endowed with supraspinal pronociceptive/anti-opioid properties (it suppresses opioid-mediated analgesia), while convergent electrophysiological and behavioural data indicate that the peptide is a spinal analgesic. Noc/oFQ has not yet been found to precipitate withdrawal in morphine-tolerant rats. Nor does it elicit motivational effects, suggesting it lacks abuse liability. Also, by acting supraspinally, noc/oFQ impairs motor performance, suppresses spatial learning, induces feeding, and regulates basal and stress-induced release of pituitary hormones. Noc/oFQ is also active when administered intravenously, exhibiting potent smooth muscle relaxant, diuretic, and antinatriuretic properties. Last but not least, noc/oFQ appears to regulate stimulated immune function, and to be involved in neuronal differentiation. The discovery of noc/oFQ, a neuropeptide with multiple functions, will certainly improve our knowledge of brain physiology, and may find therapeutic applications, for example in the management of pain or hyponatremic and water-retaining diseases. However, given the wide distribution of noc/oFQ and its receptor, the pharmacological profile of noc/oFQ is likely to be incomplete, and other as yet unknown functions of the peptide remain to be discovered. Most helpful in this respect will be the identification of new ligands of the ORL1 receptor, particularly antagonists. If research on noc/oFQ carries on unabated at the present pace, potentially clinically interesting new compounds could become available in the not too distant future.

Amino Acid Sequence↗
PubMed1997

Comparison of the structure-activity relationships of nociceptin and dynorphin A using chimeric peptides.

The aim of the present study was to delineate the functional domains of nociceptin (noc), a neuropeptide which is structurally related to dynorphin A (dyn). The binding and biological potencies towards the nociceptin (ORL1) and dynorphin A (kappa-opioid) receptors of twenty dyn/noc and noc/dyn hybrid peptides were compared with those of the parent heptadecapeptides. Replacement of as many as eleven residues in the C-terminus of dynorphin by the corresponding nociceptin sequence has no significant effect on binding and biological activity towards the kappa-opioid receptor. In marked contrast, replacement of as few as six residues (RKLANQ) in the C-terminus of nociceptin by the corresponding dynorphin sequence (LKWDNQ) dramatically impairs both affinity and activity towards the ORL1 receptor. This clearly indicates that the two neuropeptides have different functional architectures, despite the dual structural homology of both ligands and receptors. Moreover, the recombinant peptide approach led us to identify hybrids whose sequences differ only at positions 5 and 6 and displaying opposite or no receptor selectivity. One contains the dynorphin Leu5-Arg6 sequence and prefers the kappa-opioid receptor, whereas the other comprises the nociceptin Thr5-Gly6 sequence and prefers the ORL1 receptor. A third, containing the mixed dynorphin/nociceptin Leu5-Gly6 sequence, does not discriminate between the two types of receptor.

Amino Acid Sequence↗