Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.4.1 (phosphodiesterase)
 18,767 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

L-Histidine and imidazole (the histidine side chain) significantly increase cAMP accumulation in intact LLC-PK1 cells. This effect is completely inhibited by isobutylmethylxanthine (IBMX). Histidine and imidazole stimulate cAMP phosphodiesterase activity in soluble and membrane fractions of LLC-PK1 cells suggesting that the IBMX-sensitive effect of these agents to stimulate cAMP formation is not due to inhibition of cAMP phosphodiesterase. Histidine and imidazole but not alanine (the histidine core structure) increase basal, GTP-, forskolin-, and AVP-stimulated adenylate cyclase activity in LLC-PK1 membranes. Two other amino acids with charged side chains (aspartic and glutamic acids) increase AVP-stimulated but neither basal- nor forskolin-stimulated adenylate cyclase activity. This suggests that multiple amino acids with charged side chains can regulate selected aspects of adenylate cyclase activity. To better define the mechanism of histidine regulation of adenylate cyclase, membranes were detergent-solubilized which prevents histidine and imidazole potentiation of forskolin-stimulated adenylate cyclase activity and suggests that an intact plasma membrane environment is required for potentiation. Neither pertussis toxin nor indomethacin pretreatment alter imidazole potentiation of adenylate cyclase. IBMX pretreatment of LLC-PK1 membranes also prevents imidazole to potentiate adenylate cyclase activity. Since IBMX inhibits adenylate cyclase coupled adenosine receptors, LLC-PK1 cells were incubated in vitro with 5'-N-ethylcarboxyamideadenosine (NECA) which produced a homologous pattern of desensitization of NECA to stimulate adenylate cyclase activity. Despite homologous desensitization, histidine and imidazole potentiation of adenylate cyclase was unaltered. These data suggest that histidine, acting via an imidazole ring, potentiates adenylate cyclase activity and thereby increases cAMP formation in cultured LLC-PK1 epithelial cells. This potentiation requires an intact plasma membrane environment, occurs independent of a pertussis toxin-sensitive substrate and of products of cyclooxygenase, and is inhibited by IBMX. This IBMX-sensitive pathway does not involve either inhibition of cAMP phosphodiesterase activity or a stimulatory adenosine receptor coupled to adenylate cyclase.
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PMID:Histidine regulation of cyclic AMP metabolism in cultured renal epithelial LLC-PK1 cells. 168 53

A mutant LLC-PK1 cell line, M18, was isolated after a single treatment of the parent culture with N-methyl-N'-nitro-N-nitroso-guanidine. In contrast to LLC-PK1 cells, the mutant did not exhibit production of urokinase-type plasminogen activator (uPA) in response to the hormones calcitonin and vasopressin, but produced the expected levels of uPA upon stimulation by the receptor-independent adenylate cyclase activators forskolin and cholera toxin, as well as by the phosphodiesterase inhibitor isobutylmethylxanthine and the 8-bromo analogue of adenosine cyclic monophosphate, Br8cAMP. The patterns of activation of cAMP-dependent protein kinase were identical to those of uPA induction: calcitonin and vasopressin were without effect, but the response to all other agents was normal. In similar fashion, mutant cell homogenates displayed normal activation of adenylate cyclase upon treatment with sodium fluoride, forskolin, or the non-hydrolyzable GTP analogue guanosine 5'-[beta, gamma-imino]triphosphate, but were unresponsive to calcitonin or vasopressin. The ability of M18 cells to bind radioactively labelled calcitonin and vasopressin was measured. The mutant possessed less than 4% of the normal levels of the receptor binding activity for both hormones. Somatic cell hybrids formed between M18 and LLC-PK1 cells were found to retain normal hormone binding activity and responsiveness to hormones, indicating that the defect in M18 cells was recessive. M18 was concluded most probably to contain a single mutation impairing the function of two distinct polypeptide hormone receptors.
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PMID:Isolation of a mutant LLC-PK1 cell line defective in hormonal responsiveness. A pleiotropic lesion in receptor function. 302 58

Induction of urokinase-type plasminogen activator (uPA) in response to either reagents activating cAMP-dependent protein kinase (cAMP-PK) or the calcium ion phospholipid-dependent kinase (C-kinase) was compared in the LLC-PK1 and T47D cell lines. The two cell lines exhibited quantitatively different responses to calcitonin, to the phosphodiesterase inhibitor isobutylmethylxanthine, and to the adenylate cyclase activator forskolin. Both showed activation of cAMP-PK in response to all these reagents, with T47D cells displaying a greater extent of activation. T47D cells, however, failed to produce uPA in response to calcitonin, forskolin, or the cAMP analog 8-bromo-cAMP, whereas LLC-PK1 cells produced high levels of uPA in response to all these agents. Both cell lines responded to phorbol esters in terms of uPA induction, though to differing extents. Phorbol myristate acetate (PMA) was shown conclusively not to activate cAMP-PK in either cell line, even at concentrations 10-fold higher than those promoting maximal uPA induction. It was concluded that phorbol ester-mediated induction of uPA does not involve cAMP or cAMP-PK activation. These results are discussed in relation to proposed models concerning the role of cAMP-PK in uPA induction.
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PMID:Pathway of urokinase-type plasminogen activator induction in the T47D and LLC-PK1 cell lines. 365 59

Several hydrolase activities characteristic of the apical brush border membrane of renal proximal tubule, leucine aminopeptidase, gamma-glutamyl transpeptidase, alkaline phosphatase, maltase, and trehalase, were identified in cultures of the LLC-PK1 kidney epithelial cell line. A coordinate increase in activities of these enzymes was observed upon development of a confluent cell density and functional membrane polarization. Further large progressive increases in individual hydrolase activities were induced after the addition of compounds known as differentiation inducers. Hexamethylene bisacetamide preferentially induced increased trehalase and maltase activities. Induced trehalase activity exhibited an increased Vmax but a similar Km compared with activity in control extracts. Induction required protein synthesis and was dependent on inducer concentration and exposure time. Treatment of confluent cultures with N,N'-dimethylformamide triggered an induction of maltase, trehalase, alkaline phosphatase, and gamma-glutamyl transpeptidase activities, whereas dimethylsulfoxide induced trehalase and gamma-glutamyl transpeptidase activities. Increased leucine aminopeptidase and maltase activities were observed after addition of the phosphodiesterase inhibitor 1-methyl-3-isobutylxanthine. Induction of trehalase activity by N,N'-dimethylformamide was reversible over a 4-day period after removal of inducer, but effects of hexamethylene bisacetamide were irreversible. These results suggest that the LLC-PK1 cell line reproducibly develops differentiation-specific characteristics under defined conditions in cell culture, which can be individually modulated by chemicals known as inducers of cell differentiation.
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PMID:Induction of microvillar hydrolase activities by cell density and exogenous differentiation inducers in an established kidney epithelial cell line (LLC-PK1). 609 Apr 80

A cell line (LLC-PK1) isolated from porcine kidney increases 3'5'-cyclic adenosine monophosphate (cAMP) content when incubated with salmon calcitonin (SCT) or antidiuretic hormone (ADH). We have examined several factors which modulate the hormone-induced changes in cAMP levels in these cells. Preincubation with increasing concentrations of SCT results in a dose-dependent decrease in cAMP levels in cells retested with this hormone. Addition of 3-isobutyl-l-methylxanthine (IBMX) to cells preincubated with SCT results in 15--30-fold increases in cAMP levels compared to cells preincubated without this hormone. These observations suggest that the decrease in SCT-induced cAMP response in cells pretreated with this hormone is related at least in part to stimulation of phosphodiesterase activity. Preincubation with ADH does not affect subsequent cAMP response to either ADH or SCT, suggesting that these hormones interact with different cell surface receptors. Cell cycle and plating density also affect cAMP levels. cAMP content per cell increases with increasing cell density, which is associated with an increase in the SCT-induced cAMP response. These studies illustrate that factors other than receptor occupancy modulate cAMP responses of these cells to specific hormones.
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PMID:Factors modulating the response of a porcine renal tubular cell line to calcitonin and antidiuretic hormone. 616 77

In intact LLC-PK1 cells, occupancy of vasopressin receptors (Roy, C., and Ausiello, D. A. (1981) J. Biol. Chem. 256, 3415-3522) correlated with cell cAMP production. This relationship was observed as a function of hormone dose, incubation time, and changes in receptor affinity. However, the rate of cAMP production diminished with time in intact cells exposed to high hormone concentrations, even in the presence of a phosphodiesterase inhibitor. A rapid desensitization of adenylate cyclase activity was observed in minutes upon treatment of intact cells with high hormonal concentrations. Desensitization was dose- and time-dependent. Hypertonic sodium chloride, which increased hormonal binding and cell cAMP production, prevented desensitization. The acute decrease in hormone-stimulated adenylate cyclase activity correlated with increased occupancy of low affinity binding sites. EDTA-suspended cells, which have a homogeneous population of binding sites, did not demonstrate desensitization. A proposal is made as to the consequences of this phenomenon at physiological concentrations of vasopressin.
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PMID:Relationship of (8-lysine) vasopressin receptor transition to receptor functional properties in a pig kidney cell line (LLC-PK1). 616 46

Cultured pig kidney cells designated LLC-PK1, previously shown to acquire Na+-dependent concentrative transport of hexoses as the cells become growth arrested, also show Na+-dependent concentrative uptake of the amino acid analogs alpha-aminoisobutyric acid (AIB) and (methyl) meAIB. This A system-like transport is most active in sparse, growing cultures and becomes stepped down at confluence. The cell/medium equilibrium distribution ratio of the lipophilic cation tetraphenylphosphonium ion (TPP+) decreases in parallel fashion, suggesting that a decrease in membrane potential may be a major factor in the stepdown. Differentiation inducers (hexamethylene bisacetamide) and phosphodiesterase inhibitors (theophylline, methylisobutyl xanthine) accelerate the stepdown, but even in the presence of these compounds addition of the tumor promoter 12-0-tetradecanoylphorbol-13-acetate (TPA) results in the maintenance of a high level of AIB and meAIB uptake. In all these respects the changes in A system-like amino acid transport are the reciprocal of those seen for concentrative hexose transport, although the driving force appears to be the same for both systems. The TPA analogs phorbol and 4-0-methyl TPA which are inactive in tumor promotion are inactive in this system as well. In confluent, already stepped-down cultures, addition of TPA leads to a rapid (2-6 hour) stimulation of AIB and meAIB uptake. The enhancement is sensitive to cycloheximide and actinomycin D. The ouabain-sensitive fraction of meAIB uptake is not markedly changed in the TPA-enhanced uptake, nor is the TPP+ distribution ratio elevated in TPA-treated cells, making it unlikely that the TPA effect is through an alteration in the membrane potential.
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PMID:Growth-dependent AIB and meAIB uptake in LLC-PK1 cells: effects of differentiation inducers and of TPA. 618 10

LLC-PK1 cells in culture do not concentrate alpha-methylglucoside (alpha-meG) during their early growth phase but develop the capacity to concentrate this hexose as the growth rate decreases in confluent cultures. The concentrating ability is dependent on the Na+ electrochemical gradient and is inhibited by phlorizin with KI,0.5 approximately 0.2 microM. The development of the concentrative capacity can be accelerated by the Friend cell inducer hexamethylene bisacetamide (HMBA) and by the phosphodiesterase inhibitors dibutyryl cAMP, theophylline, and 1-methyl-3-isobutylxanthine (MIX). In cultures treated with any of these differentiation-accelerating chemicals, the development of alpha-meG concentrating capacity is severely inhibited by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) but not by inactive (in tumor promotion) analogs of TPA. In all cases, an early event in the development of alpha-meG accumulating capacity is an elevated intracellular cAMP concentration; however the results suggest that this increase in cAMP may be necessary but not sufficient to induce the differentiated hexose-accumulating capacity.
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PMID:Development of Na+-dependent hexose transport in a cultured line of porcine kidney cells. 627

LLC-PK1 is an established porcine renal cell line with epithelial characteristics. Upon hormonal stimulation by vasopressin, LLC-PK1 cells release adenosine 3',5'-cyclic monophosphate (cAMP) into the medium. Release of cAMP is inhibited by the organic anion transport inhibitor probenecid and by cold phosphodiesterase inhibitors and iodoacetate but not by prostaglandins A1 or E1. The kinetics of release are first order, and cAMP analogues do not induce the release of cAMP. When grown on cellulose filters, monolayers of LLC-PK1 have morphological characteristics of transporting epithelia (apical microvilli and intercellular tight junctions) and maintain a transepithelial potential difference. Stimulation of such monolayers by vasopressin elicits probenecid-sensitive release of cAMP into the medium bathing the apical surface. Smaller quantities of cAMP are released from the basolateral surface, but release in this direction is not inhibited by probenecid. In contrast, release of cAMP from the nonepithelial cell line BHK is symmetrical and is symmetrically inhibited by probenecid. Probenecid-sensitive release of cAMP from LLC-PK1 is thus a function of the apical (brush-border) membrane.
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PMID:Release of cAMP from a renal epithelial cell line. 632 92

Previous studies demonstrated that, upon attaining confluence, a clone of the renal epithelial cell, LLC-PK1, expressed progressively binding sites for the lectin Dolichos biflorus agglutinin (DBA) at the apical cell surface. Activation of cAMP-dependent protein kinase enhanced surface expression dramatically. The goal of this study was to define the process leading to surface expression of DBA binding sites and to investigate further the role of cAMP-dependent protein kinase in modulating surface expression. Both subconfluent and confluent cells exhibited intracellular DBA binding sites (50-70% of total cellular binding sites) in a perinuclear vesicular compartment which was disrupted by Brefeldin A treatment. Both total cellular content and the proportion of DBA binding sites at the cell surface increased modestly after confluence was attained. A 48 h treatment of cells with 1-methyl-3-isobutyl xanthine, a phosphodiesterase inhibitor, dramatically increased the level of cellular DBA binding sites as well as the proportion of DBA binding sites at the cell surface. Analysis of two mutants of this cell line suggests that the effect of 1-methyl-3-isobutyl xanthine requires cAMP-dependent protein kinase activity but is not due to cAMP-dependent protein kinase-mediated activation of gene transcription.
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PMID:cAMP-dependent protein kinase modulates expression and subcellular localization of Dolichos biflorus agglutinin binding sites in renal epithelial cells. 769 30


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