Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The mammalian bombesin (Bn)-like neuropeptide receptors gastrin-releasing peptide receptor (GRP-R) and neuromedin B receptor (NMB-R) transduce a variety of physiological signals that regulate secretion, growth, muscle contraction, chemotaxis and neuromodulation. We have used reverse transcription-polymerase chain reaction (PCR) to isolate a cDNA from human brain mRNA, GPCR/CNS, that encodes a putative G protein-coupled receptor (GPCR) based upon the presence of the paradigmatic seven heptahelical transmembrane domains in its predicted amino acid sequence. Analysis of the deduced protein sequence of GPCR/CNS reveals this putative receptor to be 98% identical to the deduced amino acid sequence of a recently reported gene product and minimally identical (approximately 23%) to both murine GRP-R and human endothelin-B (ET-B) receptor. Our deduced protein sequence differs at 12 positions, scattered throughout the open reading frame, relative to the original sequence. A 3.7 kb GPCR/CNS mRNA species is expressed in vivo in a tissue-specific manner, with highest levels detected in brain and spinal cord, lower levels found in testis, placenta and liver, but no detectable expression observed in any other tissue. Analysis of GPCR/CNS genomic clones reveals that the human gene contains one intron that is about 21 kb in length that divides the coding region into two exons and maps to human chromosome 7q31. No specific binding is observed with either a newly identified ligand (DTyr6, beta Ala11, Phe13, Nle14]Bn-(6-14)) having high affinity for all Bn receptor subtypes or Bn after GPCR/CNS is stably expressed in fibroblasts. No elevation in inositol trisphosphate is observed after the application of micromolar levels of either DPhe6, beta Ala11, Phe13, Nle14]Bn-(6-14) or Bn, a concentration of agonist known to activate all four known Bn receptor subtypes. When GPCR/CNS is expressed in Xenopus oocytes, no activation of the calcium-dependent chloride channel is detected despite the addition of micromolar levels of Bn peptide agonists. We conclude that the natural ligand for this receptor is none of the known naturally occurring Bn-like peptides and the true agonist for GPCR/CNS remains to be elucidated.
Brain Res Mol Brain Res 1998 Feb
PMID:A human gene encodes a putative G protein-coupled receptor highly expressed in the central nervous system. 952 70

The term "Bartter's syndrome" comprises a set of autosomal recessively inherited renal tubular disorders characterized by hypokalemia, metabolic alkalosis, hyperreninism, and hyperaldosteronism but normal blood pressure. Additional clinical and biochemical features led to a classification into phenotypically different tubulopathies: Gitelman's syndrome, hyperprostaglandin E syndrome (antenatal Bartter's syndrome), and classic Bartter's syndrome. Gitelman's syndrome results from mutations in the SLC12A3 gene encoding the human thiazide-sensitive sodium chloride cotransporter, leading to impaired reabsorption of sodium chloride in the distal convoluted tubule. Genetic heterogeneity of hyperprostaglandin E syndrome has been demonstrated by identification of mutations in the SLC12A1 gene as well as in the KCNJ1 gene. Mutations in SLC12A1 coding for the bumetanide-sensitive sodium potassium 2 chloride cotransporter (NKCC2) cause defective reabsorption of sodium chloride in the thick ascending limb of Henle's loop. Mutations in KCNJ1 leading to loss of function of the potassium channel ROMK disrupt potassium recycling back to the tubule lumen and inhibit thereby the NKCC2 activity. A third gene for hyperprostaglandin E syndrome has been mapped to the short arm of chromosome 1, and it remains to be evaluated whether other genes are involved in the pathogenesis of this disease. Classic Bartter's syndrome has been demonstrated to result from defective chloride transport across the basolateral membrane in the distal nephron due to mutations in the chloride channel gene CLCNKB. This article reviews the molecular genetic approach that has led to identification of genetic defects underlying the different hypokalemic tubulopathies.
J Mol Med (Berl) 1998 Apr
PMID:The molecular genetic approach to "Bartter's syndrome". 958 66

Cystic fibrosis transmembrane conductance regulator (cftr) gene mutations are thought to result in cystic fibrosis due to an absence of the protein's chloride channel. Recently, the lethal intestinal blockage in the cftr knockout mouse was reversed by a single in utero dose of a recombinant adenovirus containing the human cftr gene. The rescue of these animals did not require continuous expression of the gene and the cAMP-dependent chloride channel was not permanently restored. These data suggested that cftr was required for normal development of the intestine but not for normal function of the adult organ. Phenotypic changes in the intestines and lungs of in utero cftr-treated knockout and heterozygous mice revealed that altered development was induced. The intestines of the untreated knockout mice were shown to be deficient in both intracellular calcium and UTP receptors. Both of these deficiencies were partially corrected in the rescued knockout mice, whereas treatment of heterozygous animals disrupted the normal pattern of these markers. Examination of the lungs of knockout cftr (-/-) mice with lectins showed an increase in secreted glycoconjugates containing alpha(2,6)-sialic acid and fucose as compared with control heterozygotes. The in utero-treated knockouts showed an increase in this material as well, but it was contained in intracellular vesicles. Electron microscopy of these tissues confirmed the developmental alteration of secretory cell differentiation in the lungs. These data show that cftr is required in both the lung and intestines for normal differentiation of a secretory cell population and that in its absence these cells fail to develop properly.
Mol Genet Metab 1998 Jun
PMID:Molecular pathophysiology of cystic fibrosis based on the rescued knockout mouse model. 970 35

The GABAA receptor is a ligand-gated chloride channel belonging to the superfamily of ligand-gated ion channels of which the nicotinic acetylcholine (nACh) receptor is prototypic. In the central nervous system the GABAA receptor mediates fast neuronal inhibition. To facilitate the study of this receptor, a GABAA receptor-green fluorescent protein (GABAAR-GFP) chimera was constructed by fusing green fluorescent protein (GFP) to the C-terminus region of the GABAA receptor alpha1 subunit. When expressed in Xenopus oocytes, this chimera responded in a manner indistinguishable from the wild-type GABAA receptor with respect to agonist potency, receptor desensitization, allosteric modulation, rectification, and ion selectivity of the channel. The addition of GFP to the GABAA receptor alpha1 subunit did not appear to alter the assembly or efficiency of expression of the GABAA receptor complex. The GABAAR-GFP chimera generated a strong fluorescent signal that was restricted to the animal pole of the oocyte plasma membrane. This signal was readily detectable using either epifluorescence or laser confocal microscopy. To confirm the extracellular location of the GFP portion of the chimera, non-permeabilized oocytes were immunolabeled with an anti-GFP antibody. Fluorescence microscopy showed that GFP was located extracellularly since it was accessible to the GFP antibody. These results confirm the predicted extracellular location of the C-terminus of the GABAA receptor alpha1 subunit and also demonstrate that GFP retains its fluorescent property when expressed extracellularly. The usefulness of the GABAAR-GFP chimera in receptor trafficking was investigated using non-hydrolyzable GTP analogues since GTP binding proteins participate in protein transport in oocytes. Microinjections of GTP-gamma-S but not GDP-beta-S reduced both GABA-gated chloride currents and cell surface GFP fluorescence in oocytes expressing the GABAAR-GFP chimera indicating that the chimera undergoes internalization upon stimulation of oocyte GTP-binding proteins. The results of the present study show that the GABAAR-GFP chimera is functionally similar to the wild-type GABAA receptor and can be used to study receptor trafficking in living cells. This is the first demonstration of a ligand-gated ion channel-GFP chimera for an ion channel belonging to this superfamily and also is the first example of the fusion of GFP to an extracellular domain of an integral membrane protein.
Brain Res Mol Brain Res 1998 Aug 31
PMID:Functional characterization and visualization of a GABAA receptor-GFP chimera expressed in Xenopus oocytes. 972 62

Mutations in the ClC-1 muscle chloride channel cause either recessive or dominant myotonia congenita. Using a systematic screening procedure, we have now identified four novel missense mutations in dominant (V286A, F307S) and recessive myotonia (V236L, G285E), and have analysed the effect of these and other recently described mutations (A313T, I556N) on channel properties in the Xenopus oocyte expression system. Mutations V286A, F307S and A313T displayed a 'classical' dominant phenotype: their voltage dependence was shifted towards positive potentials and displayed a dominant-negative effect by significantly imparting a voltage shift on mutant-wild-type heteromeric channels as found in heterozygous patients. In contrast, the recessive mutation V236L also shifted the voltage dependence to positive values, but co-expression with wild-type ClC-1 gave almost wild-type currents. I556N, a mutation found in patients with benign dominant myotonia, drastically shifts the voltage dependence, but only a slight shift is seen when co-expressed with wild-type ClC-1. Thus, the voltage dependence of mutant heteromeric channels is not always intermediate between those of the constituent homomeric channel subunits, a conclusion further supported by mixing different ClC-1 mutants. These complex interactions correlate clinically with various inheritance patterns, ranging from autosomal dominant with various degrees of penetrance to autosomal recessive.
Hum Mol Genet 1998 Oct
PMID:ClC-1 chloride channel mutations in myotonia congenita: variable penetrance of mutations shifting the voltage dependence. 973 77

In order to gain a better insight into the structure and function of the regulatory domain (RD) of the cystic fibrosis transmembrane conductance regulator (CFTR) protein, 19 RD missense mutations that had been identified in patients were functionally characterized. Nine of these (I601F, L610S, A613T, D614G, I618T, L619S, H620P, G628R and L633P) resulted in aberrant processing. No or a very small number of functional CFTR proteins will therefore appear at the cell membrane in cells expressing these mutants. These mutations were clustered in the N-terminal part of the RD, suggesting that this subdomain has a folding pattern that is very sensitive to amino acid changes. Mutations that caused no aberrant processing were further characterized at the electrophysiological level. First, they were studied at the whole cell level in Xenopus laevis oocytes. Mutants that induced a whole cell current that was significantly different from wild-type CFTR were subsequently analysed at the single channel level in COS1 cells transiently expressing the different mutant and wild-type proteins. Three mutant chloride channels, G622D, R792G and E822K CFTR, were characterized by significantly lower intrinsic chloride channel activities compared with wild-type CFTR. Two mutations, H620Q and A800G, resulted in increased intrinsic chloride transport activities. Finally, T665S and E826K CFTR had single channel properties not significantly different from wild-type CFTR.
Hum Mol Genet 1998 Oct
PMID:Characterization of 19 disease-associated missense mutations in the regulatory domain of the cystic fibrosis transmembrane conductance regulator. 973 78

Glutamate transport is a primary mechanism for the synaptic inactivation of glutamate. Excitatory amino acid transporter 4 (EAAT4) is a novel glutamate transporter with properties of a ligand-gated chloride channel that was recently cloned from human brain. Here we report the cloning of rat EAAT4 (rEAAT4) cDNA from rat cerebellum. The nucleotide sequence of rEAAT4 was 88% identical to the human sequence, and the predicted peptide was 89% identical to the human protein. The transport activity encoded by rEAAT4 has high affinity for L-glutamate. In Xenopus laevis oocytes expressing rEAAT4, L-glutamate and other transporter substrates elicited a current predominantly carried by chloride ions. Like human EAAT4, the rEAAT4 mRNA was largely restricted to cerebellar Purkinje cells; the rEAAT4 protein was localized to Purkinje cell somas and dendrites.
Brain Res Mol Brain Res 1998 Dec 10
PMID:Molecular cloning and expression of the rat EAAT4 glutamate transporter subtype. 983 98

The cystic fibrosis transmembrane conductance regulator is a cAMP-regulated chloride channel. We used molecular modelling to predict 3-D models for the CFTR membrane domain. Hydropathy and residue conservation in all CFTRs as well as in other proteins suggested that the membrane domain is a 12-helix bundle. If the domain is enclosing a channel for chloride, it could be made of five helices. We propose two structural models in which both lumenal and cytoplasmic entrances to the chloride pore have a ring of positively charged residues. The inner surface of the channel is covered with neutral polar plus one or two charged residues. Helices that are not directly involved in the chloride channel could organise to form a second channel; a dimeric symmetrical structure is proposed. Analysis raised interest for helix 5: this hydrophobic fragment is conserved in all CFTRs and aligns with segments present in several different ion channels and transporters. The existence of an FFXXFFXXF motif is proposed. Helix 5 could be an important domain of CFTRs. The models agree with available data from pathological mutations but does not account for the membrane insertion of a hydrophilic fragment of NBDI.
J Mol Graph Model 1998 Apr
PMID:Topological model of membrane domain of the cystic fibrosis transmembrane conductance regulator. 987 57

Hyperekplexia (startle disease) results from mutations in the glycine receptor chloride channel that disrupt inhibitory synaptic transmission. The Q266H missense mutation is the only hyperekplexia mutation located in the transmembrane domains of the receptor. Using recombinant expression and patch-clamping techniques, we have investigated the functional properties of this mutation. The ability of glycine and taurine to open the channel was reduced in the mutated channel, as shown by a 6-fold shift in the concentration-response curve for both agonists. This was not accompanied by similar changes in agonist displacement of strychnine binding, suggesting that the mutation affects functions subsequent to ligand binding. Taurine was also converted to a weak partial agonist and antagonized the actions of glycine, consistent with changes in its channel gating efficacy. Because the Q266H mutation is within the pore-forming second transmembrane domain, we tested for a direct interaction with permeating ions. No change in either the cation/anion selectivity ratio or in single channel conductance levels was observed. No differential effects of Zn++, pH, and diethylpyrocarbonate were observed, implying that the histidine side chain is not exposed to the channel lumen. Single-channel recordings revealed a significant reduction in open times in the mutant receptors, at both high and low agonist concentrations, consistent with the open state of the channel being less stable. This study demonstrates that residues within the second transmembrane domain of ligand-gated ion channel receptors, even those whose side chains do not directly interact with permeating ions, can affect the kinetics of channel gating.
Mol Pharmacol 1999 Feb
PMID:The startle disease mutation Q266H, in the second transmembrane domain of the human glycine receptor, impairs channel gating. 992 32

Dent's disease, which is a renal tubular disorder characterized by low molecular weight proteinuria, hypercalciuria and nephrolithiasis, is associated with inactivating mutations of the X-linked chloride channel, CLC-5. However, the manner in which a functional loss of CLC-5 leads to such diverse renal abnormalities remains to be defined. In order to elucidate this, we performed studies to determine the segmental expression of CLC-5 in the human kidney and to define its intracellular distribution. We raised and characterized antisera against human CLC-5, and identified by immunoblotting an 83 kDa band corresponding to CLC-5 in human kidney cortex and medulla. Immunohistochemistry revealed CLC-5 expression in the epithelial cells lining the proximal tubules and the thick ascending limbs of Henle's loop, and in intercalated cells of the collecting ducts. Studies of subcellular human kidney fractions established that CLC-5 distribution was associated best with that of Rab4, which is a marker of recycling early endosomes. In addition, confocal microscopy studies using the proximal tubular cell model of opossum kidney cells, which endogenously expressed CLC-5, revealed that CLC-5 co-localized with the albumin-containing endocytic vesicles that form part of the receptor-mediated endocytic pathway. Thus, CLC-5 is expressed at multiple sites in the human nephron and is likely to have a role in the receptor-mediated endocytic pathway. Furthermore, the functional loss of CLC-5 in the proximal tubules and the thick ascending limbs provides an explanation for the occurrences of low molecular weight proteinuria and hypercalciuria, respectively. These results help to elucidate further the patho-physiological basis of the renal tubular defects of Dent's disease.
Hum Mol Genet 1999 Feb
PMID:Intra-renal and subcellular distribution of the human chloride channel, CLC-5, reveals a pathophysiological basis for Dent's disease. 993 32


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