Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.6.3.44 (P-glycoprotein)
13,344 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Previous studies of P-glycoprotein have demonstrated that its function can be modulated by phosphorylation. In the present study, inhibition of protein kinase C with calphostin C or stauroporine or prolonged treatment with the phorbol ester TPA decreased phosphorylation of P-glycoprotein, and impaired transport of vinblastine. Calphostin C also inhibited transport of actinomycin D, vincristine, rhodamine, and azidopine in SW620 Ad300 multidrug-resistant human colon carcinoma cells. Photoaffinity labeling of P-glycoprotein with azidopine was decreased by calphostin C, suggesting that dephosphorylation alters the affinity of P-glycoprotein for its substrates. Impaired transport of rhodamine in normal T lymphocytes treated with staurosporine demonstrates that modulation of P-glycoprotein function is not limited to cells selected for drug resistance in vitro. Transport of P-glycoprotein antagonists in SW620 Ad300 cells was also affected by calphostin C. Cyclosporin A transport decreased, while verapamil transport increased. Cyclosporin A in calphostin C-treated cells resulted in additive P-glycoprotein antagonism, while no additive effect could be demonstrated with verapamil, suggesting that the increase in verapamil transport makes it a poorer P-glycoprotein antagonist. These studies suggest that transport by P-glycoprotein is a dynamic process which can be modulated by phosphorylation, and that antagonists may block P-glycoprotein differently in different phosphorylation states.
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PMID:Differential modulation of P-glycoprotein transport by protein kinase inhibition. 769 Feb 50

The presence of a 'multixenobiotic resistance' [MXR] mechanism in gills of the freshwater clam Corbicula fluminea was investigated. Western blot analyses of membrane vesicles from gills, applying antibodies to vertebrate P170 multidrug resistance (MDR) protein, revealed a 135 kDa immunoreactive protein. Verapamil caused a reduction of 3H-vincristine (3H-VCR) binding onto vesicles from clam. Exposure of clams to 3H-VCR in the presence of verapamil or staurosporine (STP) enhanced the accumulation of 3H-VCR over control values. Furthermore, clams were exposed instead to VCR, to a model carcinogen, 2-acetylaminofluorene (AAF), to determine the verapamil- and STP-dependent increase of single-strand breaks (SSBs) in DNA from gills of this organism. Verapamil caused no or little increase of SSBs induced by exposure to 0.01 or 0.10 microM AAF, respectively, as measured by the alkaline elution technique. In contrast, in the presence of STP a highly significant and dose-dependent enhancement of AAF-mediated SSBs was measured already at exposure to 0.01 microM AAF. These data indicate (i) that the clam C. fluminea is provided with a P-glycoprotein-like element of the MDR-mechanism, (ii) that this system can be poisoned by chemosensitizers such as verapamil and STP, (iii) the role of protein kinase C in the regulation of MXR function and (iv) the importance of the MXR modulators for the assessment of ecotoxicological effects of pollutants.
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PMID:Increased genotoxicity of acetylaminofluorene by modulators of multixenobiotic resistance mechanism: studies with the fresh water clam Corbicula fluminea. 771 13

The phosphorylation of P-glycoprotein has been appreciated for many years, yet little is known about the factors that initiate this post-translational modification. To determine whether the activation of P-glycoprotein phosphorylation could occur in response to cellular stress and to investigate the possible signal pathways involved, we studied the effect of heat shock on the phosphorylation of P-glycoprotein in sensitive and resistant MCF-7 human breast cancer cells. Treatment of multidrug resistant MCF-7/AdrR cells with heat shock increased the phosphorylation of P-glycoprotein. The response was not seen in the sensitive MCF-7 line, which does not express this drug transporter. Phosphorylation of P-glycoprotein induced by heat shock was not dependent on synthesis of new proteins, since phosphorylation was not inhibited by cycloheximide and the content of P-glycoprotein, as measured by immunoblotting, did not change after heat shock. The activation of P-glycoprotein phosphorylation by heat shock may be initiated through activation of phospholipase C, since heat shock stimulated the activity of this enzyme, as evidenced by increased formation of inositol trisphosphate and diacylglycerol and by phosphorylation of phospholipase C-gamma. U-73122, an inhibitor of phospholipase C and staurosporine, an inhibitor of protein kinase C, both decreased the heat-shock-induced phosphorylation of P-glycoprotein. These results suggest that heat shock induces phosphorylation of P-glycoprotein through the activation of the phospholipase C/protein kinase C pathway.
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PMID:Involvement of phospholipase C in heat-shock-induced phosphorylation of P-glycoprotein in multidrug resistant human breast cancer cells. 774 43

The multidrug resistance P-glycoprotein (Pgp) transports hydrophobic drugs out of cells and has been recently associated with volume-activated chloride channels. Activation of these channels by hypotonic swelling was seen to be prevented by protein kinase C (PKC) in cells expressing high levels of Pgp by transfection. HeLa cells possess equivalent chloride currents yet they are not regulated by PKC. HeLa cells do not express Pgp as assessed by Western blotting. Following transfection of HeLa cells with cDNA encoding for Pgp, PKC-dependent suppression of volume activated chloride currents was observed. PKC regulation in transiently transfected HeLa cells was abolished by alanine replacement of the serine/threonine residues in the consensus phosphorylation sites of the linker region of Pgp. Replacement of these residues with glutamate, to mimic the effect of phosphorylation, mimicked the effects of PKC on channel activation. These results indicate that overexpression of Pgp confers PKC-regulation of endogenous volume-activated chloride channels. More generally they favour a model in which Pgp acts as a regulator of volume-activated chloride channels.
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PMID:Regulation of volume activated chloride channels by protein kinase C-mediated phosphorylation of P-glycoprotein. 775 61

The expression of protein kinase C (PKC) in 83 untreated solid human non-small cell lung carcinomas was determined and its correlation with inherent resistance to doxorubicin, with the expression of P-glycoprotein (P-170), and with the expression of glutathione S-transferase-pi (GST-pi) was analysed. Doxorubicin resistance was measured using an in vitro short-term test. The expression of PKC, P-170 and GST-pi was assessed immunohistochemically. Twenty-three tumors (= 28%) were PKC-positive, whereas 60 tumors (= 72%) were PKC-negative. Nineteen tumors (= 23%) were classified as sensitive and 64 tumors (= 77%) as resistant to doxorubicin. Thirty-nine tumors (= 47%) were P-170-positive and 51 tumors (= 61%) GST-pi-positive. Out of the PKC-positive tumors, 21 were resistant to doxorubicin and 2 were sensitive. Of the same 23 tumors, 18 were P-170-positive and 19 were GST-pi-positive. The correlations between the expression of PKC and the resistance to doxorubicin, the expression of P-170 and the expression of GST-pi were statistically significant. Corresponding results were obtained comparing the results of all tumors with the results of a subgroup of tumors having the same histology (squamous cell carcinomas). This supports the hypothesis that PKC is involved in the inherent doxorubicin-resistance of human lung cancer.
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PMID:Associated expression of protein kinase C with resistance to doxorubicin in human lung cancer. 776 22

The ability of malignant cells to develop resistance to chemotherapeutic drugs is a major obstacle to the successful treatment of clinical tumors. The phenomenon multidrug resistance (MDR) in cancer cells results in cross-resistance to a broad range of structurally diverse antineoplastic agents, due to outward efflux of cytotoxic substrates by the mdr1 gene product, P-glycoprotein (P-gp). Numerous pharmacologic agents have been identified which inhibit the efflux pump and modulate MDR. The biochemical, cellular and clinical pharmacology of agents used to circumvent MDR is analyzed in terms of their mechanism of action and potential clinical utility. MDR antagonists, termed chemosensitizers, may be grouped into several classes, and include calcium channel blockers, calmodulin antagonists, anthracycline and Vinca alkaloid analogs, cyclosporines, dipyridamole, and other hydrophobic, cationic compounds. Structural features important for chemosensitizer activity have been identified, and a model for the interaction of these drugs with P-gp is proposed. Other possible cellular targets for the reversal of MDR are also discussed, such as protein kinase C. Strategies for the clinical modulation of MDR and trials combining chemosensitizers with chemotherapeutic drugs in humans are reviewed. Several novel approaches for the modulation of MDR are examined.
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PMID:Pharmacologic circumvention of multidrug resistance. 776 25

The newly identified drug transporter MRP is functionally linked to a multiple drug resistance independent from P-glycoprotein. Resistance modifiers for this type of MDR are rare at present. We analyzed the modulating effect of the highly selective bisindolylmaleimide PKC inhibitor GF 109203X on the MRP overexpressing human MDR sublines HL60/AR and GLC4/ADR. Applying a 72 hour MTT-assay we demonstrate a complete reversal of the vincristine resistance of HL60/AR cells. Adriamycin resistance of HL60/AR, or vincristine resistance of GLC4/ADR were partially reversed. Furthermore, rhodamine 123 efflux from HL60/AR was strongly modulated by GF 109203X. Since the PKC inhibitor did not significantly influence MRP gene expression at the mRNA level which was examined by cDNA-PCR, our results suggest either a direct interaction of the compound with MRP or/and an indirect influence on MRP activity via altering the phosphorylation status of the transporter.
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PMID:The specific bisindolylmaleimide PKC-inhibitor GF 109203X efficiently modulates MRP-associated multiple drug resistance. 781 10

Human epidermoid KB cell lines resistant to high levels of adriamycin, C-A90, C-A120, C-A500, and C-A1000, were isolated in selection medium containing increasing concentrations of adriamycin, 1 microgram/ml of cepharanthine, a multidrug-resistance (MDR) reversing agent, and 100 nM of mezerein, a protein kinase C activating agent. One of the adriamycin-resistant KB cell lines, C-A500, was cross-resistant to drugs that typify the classical multidrug resistance phenotype, such as vincristine, actinomycin D, VP-16, and colchicine. The accumulation of adriamycin and vincristine was decreased in C-A500 cells and the efflux of adriamycin from C-A500 was enhanced compared with parental KB-3-1 cells. These adriamycin-resistant KB cells did not contain detectable levels of P-glycoprotein or overexpress MDR1. Multidrug-resistance-associated protein (MRP) and MRP mRNA were expressed in the adriamycin-resistant KB cells, C-A120, C-A500, and C-A1000, but not in parental KB-3-1 and revertant C-AR cells. The MRP gene was amplified in all the MDR cells that overexpressed MRP mRNA. DNA topoisomerase II levels were markedly decreased in C-A500 and C-A1000 cells but only slightly decreased in C-A120 cells. These results indicate that MRP overexpressed in the resistant cells may be responsible for the reduced accumulation of adriamycin and vincristine and that both the increased expression of MRP and decreased levels of topoisomerase II underlie the drug resistance in C-A120, C-A500, and C-A1000 cell lines.
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PMID:Non-P-glycoprotein-mediated multidrug-resistant human KB cells selected in medium containing adriamycin, cepharanthine, and mezerein. 782 64

The multidrug resistance P-glycoprotein (P-gp), which transports hydrophobic drugs out of cells, is also associated with volume-activated chloride currents. It is not yet clear whether P-gp is a channel itself, or whether it is a channel regulator. Activation of chloride currents by hypotonicity in cells expressing P-gp was shown to be regulated by protein kinase C (PKC). HeLa cells exhibited volume-activated chloride currents indistinguishable from those obtained in P-gp-expressing cells except that they were insensitive to PKC. HeLa cells did not express detectable P-gp but, following transient transfection with cDNA encoding P-gp, the volume-activated channels acquired PKC regulation. PKC regulation was abolished when serine/threonine residues in the consensus phosphorylation sites of the linker region of P-gp were replaced with alanine. Replacement of these residues with glutamate, in order to mimic the charge of the phosphorylated protein, also mimicked the effects of PKC on channel activation. These data demonstrate that PKC-mediated phosphorylation of P-gp regulates the activity of an endogenous chloride channel and thus indicate that P-gp is a channel regulator.
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PMID:Protein kinase C-mediated phosphorylation of the human multidrug resistance P-glycoprotein regulates cell volume-activated chloride channels. 782 97

P-glycoprotein is phosphorylated in cells, and it has been suggested that phosphorylation may regulate the drug transport activity of P-glycoprotein. Domain mapping, utilizing a combination of cyanogen bromide digestion and immunoblot analysis, was used to reveal the major phosphorylation sites in murine mdr1b P-glycoprotein. After labeling of J7.V1-1 cells with [32P]Pi, or labeling membranes with [gamma-32P]ATP and either protein kinase A or protein kinase C, it was found that the majority of the label was contained within a single cyanogen bromide fragment (amino acid 627-682) that encompassed the majority of the linker region. The in vitro protein kinase C phosphorylation sites within this fragment were analyzed by a combination of fast atom bombardment mass spectrometry (FABMS) and two-dimensional phosphopeptide mapping. FABMS analysis of a protein kinase C-phosphorylated synthetic peptide, corresponding to a segment of the linker region of P-glycoprotein, identified serine 669 as the single site of phosphorylation. Comparison of two-dimensional tryptic phosphopeptide maps prepared from synthetic peptide and P-glycoprotein, both of which were phosphorylated in vitro with protein kinase C, revealed that serine 669 was also the major phosphorylation site in the intact glycoprotein. The in vitro protein kinase A phosphorylation site was identified as serine 681 by site-directed mutagenesis. Inspection of the gene organization and the deduced amino acid sequence of mdr1b P-glycoprotein revealed that the linker region, although shorter than the R domain (55 versus 241 amino acids), fits the operational definition of the R domain of cystic fibrosis conductance regulator. Like the R domain, the linker region is encoded by a single exon, is highly charged with alternating acidic and basic side chains, and contains several protein kinase A/protein kinase C consensus phosphorylation sites. Since the R domain is believed to be involved in the regulation of cystic fibrosis conductance regulator function by phosphorylation, it is possible that the linker region plays a similar regulatory role in P-glycoprotein function.
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PMID:Identification of the major phosphorylation domain of murine mdr1b P-glycoprotein. Analysis of the protein kinase A and protein kinase C phosphorylation sites. 790 Dec 20


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