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)

A synthetic oligopeptide (QCDKRKRRKQQYQQRQSV) corresponding to a carboxyl-terminal sequence of the rat m3 receptor (amino acids 561-578) was coupled to carrier proteins and used to generate a polyclonal antiserum. This serum selectively immunoprecipitates at least 90% of the m3 receptors expressed by A9 cells transfected with the cDNA encoding the m3 muscarinic receptor but does not precipitate receptors from cells transfected with cDNA encoding m1, m2, m4, or m5 receptors. Using this m3 antiserum, the density of m3 receptors in various regions of rat brain was quantified. Areas expressing the highest density of m3 receptors are the cortex, hippocampus, striatum, and olfactory tubercle, with 232 fmol/mg, 197 fmol/mg, 140 fmol/mg, and 130 fmol/mg, respectively. Hindbrain regions (i.e., cerebellum, thalamus/hypothalamus, and pons/medulla) expressed fewer m3 receptors, both as a percentage of total muscarinic receptors (5-6%) and in terms of absolute receptor density (12-70 fmol/mg). A panel of subtype-selective antisera (m1, m2, and m3) was used to determine receptor distribution in several peripheral tissues of the rat (lung, ileum, and bladder). The m2 receptor subtype constitutes the majority of total receptors in the bladder (86%), lung (91%), and ileum (69%). The m3 receptor was found at lower densities in these tissues (5-11%), whereas the m1 receptor is present in highest amounts in the ileum (17%). Human clonal cell lines, in which regulation of muscarinic receptors has been commonly studied, were also examined. The SK-N-SH neuroblastoma line, which has been reported to express M3 receptors, on the basis of pharmacology and molecular size, was found to express a mixture of subtypes (m1 = 31%, m2 = 21%, m3 = 43%). Interestingly, SH-SY-5Y and SH-IN cells, both derived from SK-N-SH cells, exhibit predominantly m3 receptors (74% for SH-SY-5Y; 58% for SH-IN), with lower levels of m1 and m2 receptors (5% and 8% for SH-SY-5Y; 4% and 23% for SH-IN, respectively.) Another commonly studied cell line, 132-1-N1 astrocytoma cells, reportedly expressing M3 receptors, based upon mRNA measurements and second messenger linkage, also expresses a predominance of m3 receptors (91% of total). This m3-selective antiserum should prove useful not only for localizing and quantifying m3 muscarinic receptors but also for examining mechanisms involved in the regulation of receptor expression in human tissues or animal models of disease, as well as in cell culture.
Mol Pharmacol 1991 Nov
PMID:Development of an antiserum against m3 muscarinic receptors: distribution of m3 receptors in rat tissues and clonal cell lines. 194 43

The presence of the trivalent metallic cations, aluminum and boron, in the culture medium of differentiated human LAN-5 neuroblastoma cells results in increased amounts of specific isomers of microtubule-associated tau proteins. The cells were differentiated to a neuronal phenotype by the addition of retinoic acid. Six-day exposures of the differentiated cells to a 1-mM dose of aluminum or boron yielded increases in tau protein immunoreactivity to the monoclonal antibodies Tau-1 and Alz-50. Significant increases in immunoreactivity were seen at treatment levels of aluminum down to 100 microM. The increases in tau proteins were independent from increases in levels of total cell protein. Control cultures treated with the divalent cations zinc and iron showed no increases in levels of tau proteins.
Mol Chem Neuropathol 1991 Jun
PMID:Effects of aluminum on tau proteins in human neuroblastoma cells. 195 63

This study evaluates the role of intracellular levels of Ca2+ [Ca2+]i in cyclic GMP formation mediated by muscarinic and histamine receptors in the mouse neuroblastoma clone N1E-115. Muscarinic agonists activated the turnover of phosphoinositides with a relative maximal response similar to that observed previously for cyclic GMP formation. Carbamylcholine induced a transient increase in inositol trisphosphate with a time course similar to that of cyclic GMP formation. In cells loaded with the fluorescent Ca2+ probe fura-2/acetoxymethyl ester, carbamylcholine as well as histamine induced a rapid and transient rise in [Ca2+]i. The time course of the changes in [Ca2+]i induced by agonists as well as by ionomycin closely paralleled that of cyclic GMP formation. Chelation of [Ca2+]i by loading of N1E-115 cells with quin 2/acetoxymethyl ester inhibited cyclic GMP formation induced by agonists in a dose-dependent manner. When cyclic GMP formation induced by agonists was assayed after the cells were exposed to 3 mM ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) for 2 min, the formation of cyclic GMP was not inhibited significantly; however, it was completely abolished after 30-min exposure to EGTA. Treatment of cells with phospholipase A2 had no effect on resting [Ca2+]i and only slightly increased cyclic GMP formation, in spite of the induction of a marked release of [3H]arachidonate. Moreover, the formation of cyclic GMP induced by ionomycin was inhibited by the addition of phospholipase A2. Melittin contaminated with phospholipase A2 activity induced a rapid and sustained increase in cyclic GMP formation, as well as unesterified [3H]arachidonate release. However, after inactivation of the phospholipase A2 activity of melittin, its ability to stimulate cyclic GMP formation was enhanced. Our data indicate that receptor agonists stimulate cyclic GMP formation in N1E-115 cells by activating the formation of inositol trisphosphate, which is followed by the release of Ca2+ from intracellular stores. The evidence obtained does not support a major role for arachidonate release in receptor-mediated activation of guanylate cyclase. Conversely, it is consistent with an inhibitory role for arachidonic acid or its metabolites in this process.
Mol Pharmacol 1990 Jun
PMID:Role of intracellular Ca2+ mobilization in muscarinic and histamine receptor-mediated activation of guanylate cyclase in N1E-115 neuroblastoma cells: assessment of the arachidonic acid release hypothesis. 197 74

Unlabeled p-iodoclonidine was efficacious in attenuating forskolin-stimulated cAMP accumulation in SK-N-SH neuroblastoma cells. Maximal attenuation was 76 +/- 3%, with an EC50 of 347 +/- 60 nM. Comparable values of epinephrine were 72 +/- 3% and 122 +/- 22 nM. Responses to both agonists were abolished by 10 microM phentolamine. Therefore, p-iodoclonidine is an agonist in a cell culture model system of the neuronal alpha 2-adrenergic receptor. p-[125I]Iodoclonidine binding to membranes were measured using various regions of the rat brain. The agonist labeled a single population of sites present on cerebral cortical membranes, which was saturable (Bmax = 230 fmol/mg of protein) and possessed high affinity for the ligand (Kd = 0.6 nM). Binding was largely specific (93% at 0.6 nM). A variety of alpha 2-adrenergic agonists and antagonists were shown to compete for the binding of the radioligand. The binding of p-[125I]iodoclonidine was much less sensitive to agents that interact with alpha 1-adrenergic, serotonergic, and dopaminergic receptors. Approximately 65% of the binding was sensitive to guanine nucleotides. Association kinetics using 0.4 nM radioligand were biphasic (37% associate rapidly, with kobs = 0.96 min-1, with the remainder binding more slowly, with kobs = 0.031 min-1) and reached a plateau by 90 min at 25 degrees. Dissociation kinetics were also biphasic, with 30% of the binding dissociating rapidly (k1 = 0.32 min-1) and the remainder dissociating 50-fold more slowly (k2 = 0.006 min-1). Agonist binding is, therefore, uniquely complex and probably reflects the conformational changes that accompany receptor activation.
Mol Pharmacol 1990 Sep
PMID:p-[125I]iodoclonidine, a novel radiolabeled agonist for studying central alpha 2-adrenergic receptors. 197 27

NS20Y neuroblastoma cells expressing a homogeneous population of D1-dopamine receptors were used in the present study as a model system to investigate the mechanisms of agonist-induced stimulation and desensitization of D1 receptor-coupled adenylyl cyclase activity. Membrane prepared from NS20Y cells showed a pharmacologically specific, dose-dependent increase in cAMP production in response to various dopaminergic agonists. Dopamine exhibited an EC50 of 5 microM, and at 100 microM a maximal stimulation of 3-4-fold over basal enzyme activity was observed, which could be selectively antagonized by the active stereoisomers of SCH-23390 and butaclamol. Preincubation of NS20Y cells with dopamine induced homologous desensitization of D1 receptor-coupled adenylyl cyclase activity, decreasing dopamine- but not prostaglandin-, adenosine-, or forskolin-stimulated cAMP production. Desensitization did not affect the EC50 for dopamine but resulted in an 85-90% reduction in the maximal response. Dopamine-induced desensitization of adenylyl cyclase activity was found to be both dose and time dependent. As early as 5 min after preincubation with dopamine, cAMP production was decreased by 45-50%, with maximal desensitization occurring by 90 min. Preincubation of NS20Y cells with dopamine also induced a decrease in D1 receptor ligand binding activity, as assessed with the radiolabeled antagonist [3H]SCH-23390. This decrease in binding activity occurred more slowly than the loss of enzyme activity, not achieving maximal levels until after 3 hr. [3H]SCH-23390 saturation binding isotherms in control and maximally desensitized NS20Y cell membranes revealed no change in affinity (KD); however, a 65-70% decrease in receptor number (Bmax) was observed. Because the maximal and temporal decrease in D1 receptors does not correlated with the decrease in dopamine-stimulated enzyme activity, the desensitization may involve a functional uncoupling of the D1 receptor in addition to receptor down-regulation. This is further suggested by a loss in high affinity agonist binding observed in agonist/[3H]SCH-23390 competition experiments after desensitization. Removal of dopamine after maximal desensitization/down-regulation results in recovery to control values by 24 hr. This recovery is mostly, but not completely, blocked by protein synthesis inhibitors, suggesting an involvement of receptor degradation in the desensitization process.
Mol Pharmacol 1990 Oct
PMID:Agonist-induced desensitization of D1-dopamine receptors linked to adenylyl cyclase activity in cultured NS20Y neuroblastoma cells. 197 40

A 23-kDa (p23k) rat brain protein was stereospecifically eluted from a 14 beta-bromoacetamidomorphine affinity column, purified to apparent homogeneity by reverse phase HPLC, and partially sequenced. Three degenerate oligodeoxynucleotide probes were synthesized based on this partial amino acid sequence. A rat brain cDNA library was screened using these probes, and a full-length cDNA was isolated. The deduced protein, 187 amino acids long, is rich in glutamic and aspartic acid residues, endowing p23k with a net negative charge at neutral pH. The protein lacks a signal sequence as well as any transmembrane domains. Based on predictions of secondary structure, p23k is a globular protein composed of 30% alpha-helices and 18% beta-pleated sheets. Northern blot analysis revealed p23k transcripts in rat brain, liver, and the mouse x rat neuroblastoma-glioma NG108-14 cell line. Although not an opioid receptor itself, this protein may be associated with such a receptor or be related to a protein that has been shown to be cross-linked to the opioid peptide beta-endorphin.
Mol Endocrinol 1990 Sep
PMID:Purification, cloning, and tissue distribution of a 23-kDa rat protein isolated by morphine affinity chromatography. 197 48

A cloned thymosin beta-10 cDNA was used to study modulation of thymosin beta-10 mRNA levels in the rat B104 neuroblastoma cell line in response to retinoic acid. Northern blot analysis revealed the presence of a single greater than 600-nucleotide thymosin beta-10 mRNA species that was constitutively expressed in proliferating neuroblastoma cells. Addition of retinoic acid to the culture medium induced a dose- and time-dependent increase in thymosin beta-10 mRNA abundance. Additional studies showed that although thymosin beta-4 and beta-10 are coexpressed in this cell line, the stimulatory action of retinoic acid is specific for the thymosin beta-10 gene. Serum was found to augment the stimulatory action of retinoic acid. Blockade of protein synthesis with cycloheximide abrogated the stimulatory action of retinoic acid upon thymosin beta-10 mRNA accumulation; this observation suggests that activation of the thymosin beta-10 gene in this cell line by retinoic acid is dependent upon the de novo synthesis of a labile protein. Collectively, these findings demonstrate that the developmentally regulated thymosin beta-10 gene is a target for morphogenic retinoids in cells derived from the neural crest.
J Mol Neurosci 1991
PMID:Retinoic acid and serum modulation of thymosin beta-10 gene expression in rat neuroblastoma cells. 205 65

The neural cell adhesion molecule N-CAM has been identified in a number of species and comprises at least three major cell surface polypeptides of different molecular structures and tissue distributions. We report here the isolation and characterization of cDNA clones encoding two of the three major forms of N-CAM from a human neuroblastoma cDNA library. One of the clones, NII-6, provides the first complete sequence of a small cytoplasmic domain (140 kDa) form of the molecule in humans and differs in a number of respects from cDNA clones derived from human muscle. These differences include the presence of a 30-bp insert in the fourth immunoglobulin-like domain of N-CAM, a 3-bp insert in the extracellular portion of the molecule, and an additional 6 bp in the middle of the membrane-spanning segment. Based on the analysis of a genomic DNA clone spanning these regions of N-CAM, the first two differences arise by alternate splicing of RNA and occur in some, but not all clones; the additional 6 bp may reflect a genetic polymorphism. A second cDNA clone, NI-10, encodes the complete sequence of a segment that is specific to the large cytoplasmic domain (180 kDa) polypeptide of human N-CAM and is very similar to corresponding segments of mouse, chicken, and rat N-CAM. This sequence also arises by alternative splicing of RNA. In addition, we have identified a genomic DNA segment encoding sequences specific to the third, small surface domain (120 kDa) polypeptide of N-CAM. The data presented here and previously define the DNA sequences of the membrane-bound forms and known variants of human N-CAM. From these sequences, a wide variety of probes can be generated for investigating the expression of particular N-CAM polypeptides in normal and pathological tissues.
J Mol Neurosci 1990
PMID:Characterization of cDNA clones defining variant forms of human neural cell adhesion molecule N-CAM. 207 78

Transfected gene constructs comprising the long terminal repeat (LTR) sequence of the human immunodeficiency virus (HIV) genome spliced to an assayable reporter gene have made possible the evaluation of a lipid mediator, platelet-activating factor (PAF), as a potential HIV transcriptional regulatory molecule. We assessed the activation of the HIV LTR promoter sequence linked to the chloramphenicol acetyltransferase (CAT) reporter gene (HIV-CAT) by PAF in both a human neural (SH-SY5Y neuroblastoma) and a human leukocytic (MOLT-4 T-lymphocyte) cell line. PAF activated expression of the HIV-CAT construct in both the SH-SY5Y and MOLT-4 T-cell lines. PAF-induced CAT activity was approximately six to seven times higher in the SH-SY5Y cells than in the MOLT-4 cells. Preincubation of cells with the specific PAF antagonist BN 52021 completely inhibited CAT expression in both cell lines. The biologically inactive PAF precursor lyso-PAF did not activate CAT expression. Assays for CAT mRNA demonstrated an increase after PAF treatment, an effect that was completely inhibited by BN 52021, and which was not elicited by lyso-PAF. These results show that PAF represents a potential cellular mediator evoking the expression of the HIV genome.
J Mol Neurosci 1990
PMID:Platelet-activating factor activates HIV promoter in transfected SH-SY5Y neuroblastoma cells and MOLT-4 T lymphocytes. 207 79

In vitro differentiation of murine neuroblastoma N1E-115 cells induced by low serum (0.5%) and dimethyl sulfoxide (1.5%) increased the uptake of 45Ca2+ as well as basal and forskolin-stimulated adenylate cyclase activity. Associated with these biochemical indices of differentiation was an increase in the density of binding sites for the angiotensin II (Ang II) receptor agonist 125I-[Sar1]-Ang II and the antagonist 125I-[Sar1,Ile8]-Ang II (125I-SARILE). This up-regulation was apparent within 24 hr and was maximal at 72 hr. Other manipulations that independently increased intracellular cAMP or Ca2+ levels produced a qualitatively similar up-regulation of Ang II receptors. In vitro differentiation did not diminish the specificity of these receptors for Ang-II related peptides. Sarcosine-substituted Ang II receptor antagonists such as [Sar1,Gly8]-Ang II, [Sar1,Thr8]-Ang II, or SARILE itself competed for 125I-SARILE in a monophasic fashion, whereas the competition displayed by the agonists Ang II, angiotensin III, and Crinia-Ang II for 125I-SARILE-labeled sites was biphasic, consisting of distinct high and low affinity components. Moreover, in vitro differentiation predominantly increased the density of high affinity sites for angiotensin III and Crinia-Ang II, but the lower affinity site for Ang II, and in all three cases the majority of this increased binding was insensitive to guanine nucleotides. Collectively, these results demonstrate that the expression of Ang II receptors on neuron-like cells is regulated by the biochemical events accompanying differentiation and suggest that the biphasic nature of the binding of some angiotensin agonists may be indicative of multiple receptor subtypes.
Mol Pharmacol 1990 Dec
PMID:Up-regulation of angiotensin II receptors by in vitro differentiation of murine N1E-115 neuroblastoma cells. 212 21


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