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

The response to initial glucocorticoid therapy in childhood acute lymphoblastic leukaemia (ALL) reliably predicts the response to multiagent chemotherapy. Patients resistant to glucocorticoids (prednisone poor responders (PPR)) have a poorer event-free survival compared to glucocorticoid-sensitive patients (prednisone good responders (PGR)). A case-control study was performed to investigate differential protein expression in leukaemic blasts from PGR and PPR childhood ALL patients. Two-dimensional gel electrophoresis (2-DE) was used for an unsupervised screening and surface enhanced laser desorption/ionisation-time of flight mass spectrometry (SELDI-TOF MS) for the characterisation of protein spots. In difference maps of average gels for the proteomes of each responder group, differentially expressed proteins were identified after tryptic digestion and spotting onto H4-SELDI-TOF-MS chips. Proteins overexpressed in PPR were Catalase, RING finger protein 22 alpha, Valosin-containing protein (VCP) and a G-protein-coupled receptor. Proteins overexpressed in PGR were protein kinase C and malate dehydrogenase. Valosin-containing protein was chosen for validation and quantification by Western blot analysis in a second case-control group of ALL patients. In this second independent cohort, median VCP expression (P25-P75) was 0.15 (0.11-0.28) in PGR and 0.34 (0.14-0.99) in PPR patients (P = 0.04). We conclude that high VCP expression is associated with poor prednisone response in childhood ALL patients.
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PMID:Unsupervised proteome analysis of human leukaemia cells identifies the Valosin-containing protein as a putative marker for glucocorticoid resistance. 1654 Nov 42

Mitochondrial ATP sensitive potassium channels (mitoK(ATP) channels) are involved in the cardioprotection afforded by ischemic preconditioning (IPC) and diazoxide, a selective mitoK(ATP) channel opener. The activation of some kinases, including phoshoprotein kinase (PKC)-epsilon and mitogen-activating protein kinases (MAPK), is involved in signal conduction of preconditioning downstream from mitoK(ATP) channel opening. Diazoxide can open mitoK(ATP) channels and activate PKC-epsilon, which will phosphorylate some substrate proteins. These proteins that exhibit altered post-translational modification via phosphorylation due to diazoxide pretreatment may be the target molecules and play an important role in cellular protection after mitoK(ATP) channel opening. To analyze and identify the phosphoproteins associated with diazoxide preconditioning, phosphoprotein enrichment and comparative two-dimensional gel electrophoresis (2D-GE) were used. Cultured adult rat ventricular myocytes were pretreated in the presence and absence of 100 micronol/1l diazoxide for 10 min and enriched phosphoproteins from control myocytes and those pretreated with 100 micromol/l diazoxide were separated by 2D-GE and stained with a silver staining kit. Phosphoproteins of interest were further identified by matrix-assisted laser desorption ionization tandem mass spectrometry (MALDI-TOF MS). Eight protein spots with different abundance were found, of which six differentially expressed proteins were identified by MALDI-TOF MS. They included 94 kDa glucose-regulated protein, calpactin I heavy chain, chaperonin containing TCP-1 zeta subunit, hypothetical protein XP_346548, ferritin light chain and ferritin light chain 2. These findings provide new clues to understanding the mechanism of ischemic preconditioning in cardiomyocytes downstream from mitoK(ATP) channel opening.
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PMID:Comparative proteomics analysis of differentially expressed phosphoproteins in adult rat ventricular myocytes subjected to diazoxide preconditioning. 1684 16

Deregulation of protein kinase-mediated signaling events is one of the major causes to malignant transformation. In this work, we tried to purify protein kinase inhibitory activity and antitumor activity from ethanol extracts of the seeds of Livistona chinensis R. Brown (LC), a traditional herb used for the treatment of nasopharyngeal carcinoma (NPC). Both activities were found to be co-purified in various chromatography steps, and a highly purified fraction, LC-X, was obtained and its biological effects were characterized further. LC-X inhibited the activities of various protein kinases in vitro, including PAK2, PKA, PKC, GSK-3alpha, CK2, mitogen-activated protein kinase (MAPK), and JNK1, with IC(50) between approximately 1 and 40microg/ml. The proliferation of two NPC (NPC-TW02 and -TW04) and one breast cancer (MCF-7) cell lines, but not the epidermoid (A431) and cervical (HeLa) carcinoma cell lines, were significantly blocked by LC-X at the dose of >50microg/ml. Cell cycle arrested at G(2)/M phase and apoptosis were detected in NPC-TW02 cells treated with LC-X for 24h. Further studies revealed that epidermal growth factor (EGF)-induced activation of epidermal growth factor receptor (EGFR) and MAPK could be potently inhibited by LC-X in both NPC-TW02 and A431cells in a dose-dependent manner. More interestingly, the level of EGFR protein detected by Western blot decreased drastically in LC-X-treated A431 and NPC-TW02 cells in a dose- and time-dependent fashion. Further analysis of the plasma membrane and cytosolic fractions from LC-X-treated and untreated A431 cells showed that a 170kDa protein selectively disappeared from the plasma membrane of LC-X-treated cells. The protein was identified as EGFR by MALDI-TOF mass spectrometry, indicating EGFR as a selective target for LC-X. Moreover, the electrophoretic mobility of purified EGFR in SDS-PAGE was altered dramatically post LC-X treatment, suggesting that LC-X may chemically modify EGFR. In conclusion, the active components with both antitumor and protein kinases inhibitor activities were highly purified from LC, which can inhibit the EGF signaling events mainly through EGFR modification. Blockage of the functions of EGFR may account for the antitumor activity of these active components.
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PMID:Selective downregulation of EGF receptor and downstream MAPK pathway in human cancer cell lines by active components partially purified from the seeds of Livistona chinensis R. Brown. 1691 67

Recently, uridine adenosine tetraphosphate (Up(4)A) was described as a strong vasoconstrictor released from endothelial cells after stimulation with mechanical stress. In this study, we isolated and identified Up(4)A from kidney tissue, and we characterized the essential varying effects of Up(4)A on the afferent and efferent arterioles. Porcine and human kidney tissue was fractionated by size exclusion chromatography, affinity chromatography, anion exchange chromatography and reverse phase chromatography. In fractions purified to homogeneity, Up(4)A was identified by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS), MALDI-LIFT fragment mass spectrometry (MALDI-TOF/TOF MS), retention-time comparison and enzymatic cleavage analysis. We analysed the release of Up(4)A from cultivated renal proximal tubule cells after stimulation of protein kinase C with oleoyl-2-acetyl-sn-glycerol (OAG). Up(4)A was identified in renal tissue, and the effect of Up(4)A on the vascular tone of isolated perfused afferent and efferent arterioles was tested. Stimulation of tubule cells with OAG increased the release rate of Up(4)A from tubule cells about tenfold. Up(4)A acts as a strong vasoconstrictive mediator on afferent arterioles, but has no significant effect on the tone of efferent arterioles, suggesting a functional role of Up(4)A as an autocrine hormone for glomerular perfusion. Because of the predominant effect of the Up(4)A on afferent arterioles, we assume that Up(4)A may decrease glomerular perfusion, intra-glomerular pressure and, hence, glomerular filtration rate. The release of Up(4)A from renal tubular cells may be an additional mechanism whereby tubular cells could affect renal perfusion. Up(4)A release may further contribute to renal vascular autoregulation mechanisms. In conclusion, as Up(4)A occurs in renal tissue and has marked effects on afferent but not efferent arterioles, Up(4)A may play a role in renal hemodynamics and possibly blood pressure regulation.
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PMID:Uridine adenosine tetraphosphate acts as an autocrine hormone affecting glomerular filtration rate. 1825 Sep 97

The purpose of this study was to find protein kinase C (PKC) isozyme-specific peptides. A peptide library containing 1772 sequences was designed using Scansite and screened by MALDI-TOF MS and kinase activity assays for PKC isozyme-specificity. A peptide (Alphatomega; H-FKKQGSFAKKK-NH(2)) with high specificity for PKC alpha relative to other isozymes was identified. The peptide was phosphorylated to a greater extent by tissue lysates from B16 melanoma, HepG2, and human breast cancer, which had higher levels of activated PKC alpha, when compared to normal skin, liver, and human breast tissue lysates, respectively. Moreover, addition of Ro-31-7549, an inhibitor with great specificity for PKC alpha, to the phosphorylation reaction caused a dose-dependent reduction in phosphorylation, but no inhibition was identified with the addition of rottlerin and H-89. These results show that this peptide has great potential as a PKC alpha-specific substrate.
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PMID:A short peptide is a protein kinase C (PKC) alpha-specific substrate. 1842 34

The reinforcing effects and long-term consequences of cocaine self-administration have been associated with brain regions of the mesolimbic dopamine pathway, namely the nucleus accumbens (NAc). Studies of cocaine-induced biochemical adaptations in rodent models have advanced our knowledge; however, unbiased detailed assessments of intracellular alterations in the primate brain are scarce, yet essential, to develop a comprehensive understanding of cocaine addiction. To this end, two-dimensional difference in gel electrophoresis (2D-DIGE) was used to compare changes in cytosolic protein abundance in the NAc between rhesus monkeys self-administering cocaine and controls. Following image normalization, spots with significantly differential image intensities (P<0.05) were identified, excised, trypsin digested and analyzed by matrix-assisted laser-desorption ionization time-of-flight time-of-flight (MALDI-TOF-TOF). In total, 1098 spots were subjected to statistical analysis with 22 spots found to be differentially abundant of which 18 proteins were positively identified by mass spectrometry. In addition, approximately 1000 protein spots were constitutively expressed of which 21 proteins were positively identified by mass spectrometry. Increased levels of proteins in the cocaine-exposed monkeys include glial fibrillary acidic protein, syntaxin-binding protein 3, protein kinase C isoform, adenylate kinase isoenzyme 5 and mitochondrial-related proteins, whereas decreased levels of proteins included beta-soluble N-ethylmaleimide-sensitive factor attachment protein and neural and non-neural enolase. Using a complimentary proteomics approach, the differential expression of phosphorylated proteins in the cytosolic fraction of these subjects was examined. Two-dimensional gel electrophoresis (2DGE) was followed by gel staining with Pro-Q Diamond phosphoprotein gel stain, enabling differentiation of approximately 150 phosphoprotein spots between the groups. Following excision and trypsin digestions, MALDI-TOF-TOF was used to confirm the identity of 15 cocaine-altered phosphoproteins. Significant increased levels were detected for gamma-aminobutyric acid type A receptor-associated protein 1, 14-3-3 gamma-protein, glutathione S-transferase and brain-type aldolase, whereas significant decreases were observed for beta-actin, Rab GDP-dissociation inhibitor, guanine deaminase, peroxiredoxin 2 isoform b and several mitochondrial proteins. Results from these studies indicate coordinated dysregulation of proteins related to cell structure, signaling, metabolism and mitochondrial function. These data extend and compliment previous studies of cocaine-induced biochemical alterations in human postmortem brain tissue, using an animal model that closely recapitulates the human condition and provide new insight into the molecular basis of the disease and potential targets for pharmacotherapeutic intervention.
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PMID:Integrative proteomic analysis of the nucleus accumbens in rhesus monkeys following cocaine self-administration. 1850 25

To identify the correlation between the phosphorylation ratios by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-ToF MS) analysis and enzyme kinetics (Km, Vmax, and Vmax/Km) is important to understand whether MALDI-TOF MS can be applied for monitoring the properties of peptides that are substrates of protein kinases. The correlation between phosphorylation ratios and enzyme kinetics was examined using peptides for protein kinase C (PKC) and for 60 kDa phosphoprotein, encoded by the cellular sarcoma gene (c-Src). Phosphorylation ratios, analyzed by MALDI-ToF MS, showed higher correlation coefficient (r = or > +0.7) for Vmax/Km compared with that (r = or < -/+0.6) for Km or Vmax. For ion modes, a higher correlation coefficient between phosphorylation ratios and Vmax/Km was identified in the positive mode (r = or > +0.7) compared to that in the negative mode (r = or < +0.5). These results suggest that MALDI-ToF MS is a useful tool to evaluate Vmax/Km of peptides for protein kinases.
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PMID:Letter: correlation between phosphorylation ratios by matrix-assisted laser desorption/ionization time-of-flight mass spectrometric analysis and enzyme kinetics. 1875 24

Failure of oligodendrocyte precursor cell (OPC) differentiation contributes significantly to failed myelin sheath regeneration (remyelination) in chronic demyelinating diseases. Although the reasons for this failure are not completely understood, several lines of evidence point to factors present following demyelination that specifically inhibit differentiation of cells capable of generating remyelinating oligodendrocytes. We have previously demonstrated that myelin debris generated by demyelination inhibits remyelination by inhibiting OPC differentiation and that the inhibitory effects are associated with myelin proteins. In the present study, we narrow down the spectrum of potential protein candidates by proteomic analysis of inhibitory protein fractions prepared by CM and HighQ column chromatography followed by BN/SDS/SDS-PAGE gel separation using Nano-HPLC-ESI-Q-TOF mass spectrometry. We show that the inhibitory effects on OPC differentiation mediated by myelin are regulated by Fyn-RhoA-ROCK signalling as well as by modulation of protein kinase C (PKC) signalling. We demonstrate that pharmacological or siRNA-mediated inhibition of RhoA-ROCK-II and/or PKC signalling can induce OPC differentiation in the presence of myelin. Our results, which provide a mechanistic link between myelin, a mediator of OPC differentiation inhibition associated with demyelinating pathologies and specific signalling pathways amenable to pharmacological manipulation, are therefore of significant potential value for future strategies aimed at enhancing CNS remyelination.
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PMID:Myelin-mediated inhibition of oligodendrocyte precursor differentiation can be overcome by pharmacological modulation of Fyn-RhoA and protein kinase C signalling. 1920 90

Legionella pneumophila, the agent of Legionnaires' disease, replicates intracellularly within specialized phagosomes of human macrophages and amoebae. In this study, we have developed a protocol for the isolation of Legionella-containing phagosomes from Dictyostelium discoideum. Cell fractionation, two-dimensional gel electrophoresis and MALDI-TOF MS combined with genomic data identified 157 phagosome host proteins. In addition to proteins with an evident role in phagosome maturation, we identified proteins for which a function remains to be elucidated. Possible interactions of coronin with cytosolic NADPH oxidase components and protein kinase C inhibitors which together may lead to an inhibition of phagosomal superoxide generation are discussed. Comparative proteomics of phagosomes containing highly virulent L. pneumophila Corby versus less virulent L. hackeliae revealed distinctive protein expression patterns, e.g., an abundance of RhoGDI in L. hackeliae degrading phagosomes versus little RhoGDI in L. pneumophila Corby replicative phagosomes. We present a kinetic dissection of phagosome maturation including the complex alterations of the phagosome protein composition. A reference flow chart suggests so far unrecognized consequences of infection for host cell physiology, actin degradation on phagosomes, and a putative cysteine proteinase inhibitor interference with lysosomal enzyme sorting and activation processes.
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PMID:Proteomic analysis of Legionella-containing phagosomes isolated from Dictyostelium. 1948 47

Extracellular proteins secreted/released by protozoan parasites are key mediators of the host-parasite interaction. To characterise the profile of proteins secreted/released by Leishmania (Viannia) braziliensis promastigotes, a proteomic approach combining two-dimensional electrophoresis (2DE), tandem matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF/TOF) mass spectrometry, and data mining was carried out. The 2DE map revealed a set of 270 secreted protein spots from which 42 were confidently identified and classified into 11 categories according to Gene Ontology (GeneDB database) and KEEG Ontology annotation of biological processes. Parasite promastigotes were able to secrete/release proteins involved in immunomodulation, signal transduction, and intracellular survival, such as HSP70, acid phosphatase, activated protein kinase C receptor (LACK), elongation factor 1beta, and tryparedoxin peroxidase. Data mining showed that approximately 5% of identified proteins present a classical secretion signal whereas approximately 57% were secreted following non-classical secretion mechanisms, indicating that protein export in this primitive eukaryote might proceed mainly by unconventional pathways. This study reports a suitable approach to identify secreted proteins in the culture supernatant of L. braziliensis and provides new perspectives for the study of molecules potentially involved in the early stages of infection.
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PMID:Proteomic characterization of the released/secreted proteins of Leishmania (Viannia) braziliensis promastigotes. 1970 3


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