Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

By observing increases in the transepithelial paracellular permeability of a range of radiolabeled solutes and electron dense dyes, changes in molecular sieving caused by the cytokine, TNF (tumor necrosis factor), and the phorbol ester, TPA (12-0-tetra-decanoylphorbol-13-acetate), were characterized. Using 14C-labeled mannitol (mw 182), raffinose (mw 504), PEG (polyethylene glycol; mw 4000), and dextran (mw 10,000, 70,000 and 2,000,000), the transepithelial flux rates of these compounds were determined at the peak of the transepithelial electrical resistance (TER) changes caused by these two agents. TNF treatment resulted in increased permeability across LLC-PK1 epithelial cell sheets only to relatively small solutes, with an upper limit of approximately 4,000 mw. The low molecular weight "ceiling" for the TNF-treated epithelium is further evidence against TNF increasing transepithelial permeability by means of inducing nonspecific, microscopic "holes" in the epithelium, for which a "ceiling" would not exist. TPA treatment increases transepithelial paracellular permeability to a much broader range of solutes, extending well beyond 2 million mw. Transmission electron micrographs provide evidence that even the electron-dense dye complex, ruthenium red, can cross tight junctions of TPA-treated cell sheets. However, cationic ferritin cannot cross tight junctions of TPA-treated cell sheets. This shows that there is an upper limit to solutes able to cross TPA-treated cell sheets, but that this upper limit will include most proteins, which would then be able to cross tumor promoter-exposed (protein kinase C-activated) epithelial layers at accelerated rates. The biomedical implications for a high molecular weight cutoff in tumor promoter action in epithelial carcinogenesis, and for a low molecular weight cutoff in cytokine-induced epithelial apoptosis in inflammation, are discussed.
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PMID:Different size limitations for increased transepithelial paracellular solute flux across phorbol ester and tumor necrosis factor-treated epithelial cell sheets. 913 Apr 71

Modulation of protein kinase C (PKC) by 12-O-tetradecanoylphorbol-13-acetate (TPA) disrupts the cell-cell junctions of the epithelial cell line LLC-PK1. To examine the role of specific PKC isoforms in this process we have created modified LLC-PK1 subclones that express wild-type and dominant negative versions of PKC-alpha under control of the tetracycline-responsive expression system. Overexpression of wild-type PKC-alpha rendered the cells more sensitive to the effects of TPA on transepithelial permeability as measured by loss of transepithelial resistance across the cell sheet. Conversely, expression of a dominant negative PKC-alpha rendered the cells more resistant to the effects of TPA as measured both by loss of transepithelial resistance as well as cell scattering. The properties of both subclones could be modulated by the addition of tetracycline, which suppressed the effect of the exogenous genes. These results indicate that the alpha isoform of PKC is at least one of the isoforms that regulate tight junctions and other cell-cell junctions of LLC-PK1 epithelia.
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PMID:Protein kinase C-alpha activity modulates transepithelial permeability and cell junctions in the LLC-PK1 epithelial cell line. 916 67

The effect of chronic hypoxia on the proliferation and dedifferentiation of LLC-PK1 cells was examined. Cultures were exposed either to hypoxia (3% O2) or normoxia (18% O2), and [3H]thymidine incorporation, cell number, and sodium-dependent glucose (Na/Glc) uptakes were assessed. Cultures exposed to hypoxia for 16 h significantly increased [3H]thymidine incorporation followed by a significant increase in cell number both at 24 and 48 h in comparison with respective normoxic controls. Cultures exposed to 24 and 72 h of hypoxia exhibited significant inhibition of Na/Glc uptake when compared with their respective normoxic counterparts. Significant inhibition of cell ATP levels were observed under hypoxic conditions. Acute reoxygenation of hypoxic cells normalized cell ATP levels without any effect on the Na/Glc uptake. Hypoxia also activated protein kinase C (PKC) at 1 and 4 h followed by a subsequent return to baseline with reactivation at 24 h, which remained sustained up to 72 h, suggesting both acute and sustained activation of PKC. Furthermore, the hypoxia-induced alterations in [3H]thymidine incorporation as well as Na/Glc uptake were mitigated by inhibitors of PKC. These results indicate that chronic hypoxia induces both proliferation and dedifferentiation of LLC-PK1 cells mediated, in part, by the activation of PKC.
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PMID:Chronic hypoxia induces LLC-PK1 cell proliferation and dedifferentiation by the activation of protein kinase C. 922 43

In LLC-PK1 cells, urokinase-type plasminogen activator (uPA) mRNA has a short half-life of 70 min. We have previously demonstrated that most of the regulatory regions responsible for the rapid turnover of uPA mRNA in LLC-PK1 cells reside in its 3' untranslated region (3' UTR), where there are at least three regulatory sites, one of which is A+U-rich. This A+U-rich sequence mediates uPA mRNA stabilization induced by protein kinase C (PKC) down-regulation. In this work, we found that uPA mRNA is rather stable in MDA-MB-231 cells with a half-life of 17 h. We compared the stability of hybrid globin mRNA containing different parts of uPA mRNA in its 3' UTR and found that the A+U-rich sequence of uPA mRNA renders otherwise stable globin mRNA unstable in LLC-PK1 cells but not in MDA-MB-231 cells. We identified a cytoplasmic protein of 40 kDa (p40) which specifically interacts with the A+U-rich sequence. Levels of p40 activity as detected by ultraviolet cross-linking were higher in MDA-MB-231 and PKC-down-regulated LLC-PK1 cells than in untreated LLC-PK1 cells. Prior treatment of the cytoplasm with a specific antibody against heterogeneous nuclear ribonucleoprotein C (hnRNP C) significantly reduced p40 activity. These results suggest a correlation between the A+U-rich sequence-dependent uPA mRNA stabilization in vivo and the binding of hnRNP C to the A+U-rich sequence in vitro.
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PMID:Enhanced stability of urokinase-type plasminogen activator mRNA in metastatic breast cancer MDA-MB-231 cells and LLC-PK1 cells down-regulated for protein kinase C--correlation with cytoplasmic heterogeneous nuclear ribonucleoprotein C. 924 23

Although the exact mechanism of prostaglandin E2 (PGE2)-mediated cytoprotection has not been elucidated, its ability to induce cytoprotection in cell culture suggests this action occurs at the cellular level. The present studies were conducted to determine whether PGE2 induces protection against 2,3,5-(trisglutathion-S-yl)-hydroquinone [2,3,5-(trisglutathion-S-yl)-HQ]-mediated cytotoxicity in a renal proximal tubule epithelial cell line (LLC-PK1) and to delineate the cellular and molecular mechanisms associated with this response. Pretreatment of LLC-PK1 cells with 0.01-40 microM PGE2 for 24 h fully protects against a moderately toxic concentration of 2,3,5-(trisglutathion-S-yl)-HQ. PGE2-mediated cytoprotection is observed in cells pretreated at pH 7.4 but not at pH 7.8. However, cytoprotection is observed in LLC-PK1 cells pretreated with the PGE2 analog, 11-deoxy-16,16-dimethyl PGE2 (DDM-PGE2) but not with the PGE2 receptor [E-prostanoid (EP)] agonists 17-phenyltrinor PGE2 (EP1), 11-deoxy PGE1 (EP2/EP4), sulprostone (EP1/EP3), PGE1, or PGA2. 12-O-tetradecanoylphorbol-13-acetate (TPA), a potent activator of protein kinase C (PKC), also induces cytoprotection, supporting a role for this pathway in the cytoprotective response. PGE2, DDM-PGE2, and TPA all induce the binding of nuclear proteins to a TPA responsive element (TRE), whereas analogs that did not induce cytoprotection (PGE1, 17-phenyltrinor PGE2, sulprostone) were without effect. DDM-PGE2- and TPA-mediated cytoprotection and TRE binding activity are inhibited by N-(2[[3-(4-bromophenyl)-2-propenyl]-amino]-ethyl)-5-isoquinolinesulfonam ide (H-89), a PKC inhibitor. These data suggest that cytoprotection by PGE2 and DDM-PGE2 in LLC-PK1 cells is mediated by a PKC-coupled receptor, which is pharmacologically distinct from the presently classified EP receptor subtypes.
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PMID:PGE2-mediated cytoprotection in renal epithelial cells: evidence for a pharmacologically distinct receptor. 936 28

The alkylating reagent iodoacetamide, a potent inhibitor of sulfhydryl proteases, was found to stimulate the selective degradation of protein kinase C alpha (PKC alpha) isoform (80 KDa). Treatment of LLC-PK1 cells with iodoacetamide (0.5-15 mM) for 30-90 minutes at room temperature allowed by western blotting on total cell homogenate, revealed the appearance of an 50 KDa band that was still recognized with the antibody. However, iodoacetamide (15 mM) resulted in the total disappearance of the 80 KDa protein. Serine protease inhibitors, metalloprotease inhibitors and leupeptin failed to prevent the degradation of PKC alpha. The degradation persisted at 4 degrees C and in the absence of Ca2+. Iodoacetamide had no direct effect on purified PKC alpha. PKC activities in iodoacetamide-treated cells were also inhibited. In conclusion, the degradation of PKC alpha is a novel phenomenon. The degradation process could not be prevented by known protease inhibitors or in the absence of Ca2+ or by incubation at 4 degrees C and appears to involve interactions with unknown cellular intermediates.
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PMID:The ATP-depleting reagent iodoacetamide induces the degradation of protein kinase C alpha (PKC alpha) in LLC-PK1 pig kidney cells. 936 85

Intracellular signal transduction for regulation of alkaline phosphatase (ALP) activity in renal epithelial cells treated with calcitonin is not yet completely understood, although it is known that calcitonin receptors couple to cyclic AMP-dependent protein kinase (PKA) and calcium/phospholipid-dependent protein kinase (PKC). Salmon calcitonin increased the cyclic AMP content in LLC-PK1 porcine kidney cells in a concentration-dependent manner. When the confluent cells were incubated for 47 h after a 1 h-pulse exposure or continuously exposed to calcitonin and forskolin for 48 h, ALP activity in the cells was increased by calcitonin about 2-fold compared with the basal activity at the maximum level but was not dependent on the exposure time; it was markedly increased by forskolin in parallel with the exposure time. The increase in activity produced by calcitonin was abolished by a PKA inhibitor H-89, and, in contrast, potentiated by a PKC inhibitor, NA-382 to near the forskolin-induced level. These results indicate that calcitonin exerts a dual-regulation of ALP activity in LLC-PK1 cells, positively through the PKA pathway and negatively through PKC.
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PMID:Dual regulation of alkaline phosphatase activity by calcitonin in porcine kidney cells. 944 8

Although exposure of LLC-PK1 epithelial cell sheets to phorbol esters (TPA) causes a near immediate and total decrease of transepithelial electrical resistance (TER), continuation of exposure for 3 to 4 days results in a tachyphylactic response as TER begins to return to control levels. Recovery of TER is maximal by 5 to 6 days, but reaches only 70 to 80% of control level. A reciprocal change in the transepithelial flux of D-mannitol indicates that the TER decrease is indicative of an increase in tight junction permeability. Exposure of cell sheets to TPA for several days also results in the appearance of multilayered polyp-like foci (PLFs) across the otherwise one cell layer thick cell sheets. The pattern of penetration of the electron dense dye, ruthenium red, from the apical surface, across the tight junction and into the lateral intercellular space indicates that the tight junctions of the cell sheet become uniformly leaky after acute exposure to TPA. However, when exposure is continued for several days, only the junctions of cells in the PLFs manifest leakiness. The decrease in TER following acute TPA exposure correlates with the translocation of protein kinase C-alpha (PKC alpha) into a membrane-associated compartment. With exposure of several days, only a trace of PKC alpha is visible by Western immunoblot, and this is in the membrane-associated compartment. Immunofluorescent microscopy indicates that the trace of PKC alpha seen in the Western immunoblots is ascribable distinctly to cells of the PLFs. Monolayer areas between PLFs show no discernible immunofluorescent signal. The data therefore indicate that tight junction barrier function may be restored in certain areas by the down regulation of PKC alpha from the membrane-associated compartment. Failure to down regulate may result in the paracellular leakiness and abnormal cell architecture of the PLFs. Possible implications of this model for in vivo epithelial tumor promotion are discussed.
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PMID:Transepithelial paracellular leakiness induced by chronic phorbol ester exposure correlates with polyp-like foci and redistribution of protein kinase C-alpha. 945 Apr 79

LLC-PK1, an epithelial cell line derived from the kidney proximal tubule, was used to study the ability of the G protein alpha-subunit, G alpha q, to regulate cell differentiation. A constitutively active mutant protein, alpha qQ209L, was expressed using the LacSwitch-inducible mammalian expression system. Induction of alpha qQ209L expression with isopropyl-beta-D-thiogalactopyranoside (IPTG) enhanced phospholipase C activity maximally by 6- to 7.5-fold. Increasing concentrations of IPTG progressively inhibited the activity of two differentiation markers, Na(+)-dependent hexose transport and alkaline phosphatase activity. Induction of alpha qQ209L expression also caused a change from an epithelial to a spindle-shaped morphology. The effects of alpha qQ209L expression on cell differentiation were similar to those observed with 12-O-tetradecanoylphorbol 13-acetate (TPA) treatment. However, protein kinase C (PKC) levels were downregulated in TPA-treated cells but not in alpha qQ209L-expressing cells, suggesting that the regulation of PKC by G alpha q may be different from regulation by TPA. Interestingly, the PKC inhibitor GF-109203X did not inhibit the effect of IPTG on the development of Na(+)-dependent hexose transport in alpha qQ209L-expressing cells. These data implicate PKC delta and PKC epsilon in the pathway used by G alpha q to block the development of Na(+)-dependent hexose transport in IPTG-treated cells.
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PMID:Inhibition of cell differentiation by G alpha q in the renal epithelial cell line LLC-PK1. 957

Studies have shown that the renal tubular epithelium adapts to alterations in the sulfur amino acid composition of the diet. The renal adaptive response has been described in man, mouse, rat, dog, and pig. The observed phenomenon involves increased or decreased initial rate activity of the NaCl-dependent taurine transporter at the brush border membrane surface of the proximal tubule following dietary manipulation of taurine. A cDNA encoding a taurine transporter has been isolated from LLC-PK1 cells, designated pTAUT, and its functional properties have been examined in Xenopus laevis oocytes. The nucleotide sequence of the clone predicts a 621-amino acid protein with about 90% homology to other cloned taurine transporter cDNAs. When expressed in oocytes the transporter displays a Km of 25 microM and is dependent on the presence of external sodium and chloride, characteristics similar to taurine uptake by LLC-PK1 cells. The abundance of pTAUT mRNA and protein were up-regulated in cells cultured in taurine-free medium as compared with cells cultured in medium containing 500 microM taurine. Activation of PKC by PMA had no effect on adaptive regulation of pTAUT mRNA and protein, indicating that down-regulation of LLC-PK1 cell taurine transport activity by PMA occurs at the post-translational level.
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PMID:Molecular cloning and functional expression of an LLC-PK1 cell taurine transporter that is adaptively regulated by taurine. 963 40


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