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)

Atherogenesis involves an early endothelial dysfunction hallmarked by elevated free radical production and increased adhesiveness for monocytes. It was hypothesized that activation of the tissue renin angiotensin system may contribute to the endothelial alteration. To test this hypothesis, thoracic aortae were isolated from normocholesterolemic (NC; n = 6) and hypercholesterolemic (HC; n = 6; diet: 0.5% cholesterol; 6 weeks) New Zealand white rabbits, and incubated for 2 h with the angiotensin II (Ang II) receptor antagonist Sar-1,Ile-8-Ang II, the antioxidant pyrolidine dithiocarbamate (PDTC) and the protein kinase C (PKC) antagonist staurosporin. Superoxide production from aortic segments was measured by lucigenin-enhanced chemiluminescence. In comparison to the normocholesterolemic state, hypercholesterolemia led to a significant increase in superoxide production (221 +/- 44%, p < 0.02); this was reduced by ex vivo treatment of the vessel segment with Ang II-antagonist (to 130 +/- 29%; p < 0.04 vs HC), or PKC-antagonist (to 86 +/- 26%; p < 0.001 vs HC), or PDTC (to 103 +/- 27%; p < 0.02 vs HC). Monocyte-endothelial interaction was assessed by functional binding assay. When compared to normocholesterolemic rabbits, hypercholesterolemia led to a twofold increase in monocyte binding (74 +/- 13 vs 37 +/- 4 monocytoid cells per high power field (m/hpf); p < 0.03). The Ang II-antagonist and the PKC-antagonist led to a normalization of monocyte-endothelial binding (Ang II-antagonist: 37 +/- 9 m/hpf; PKC-antagonist: 41 +/- 17 m/hpf; p < 0.05). In conclusion, these results indicate that hypercholesterolemia activates the tissue renin angiotensin system, which results in an increased endothelial production of superoxide and monocyte adhesiveness. Ang II-antagonist inhibits free radical production and monocyte adhesion through a mechanism which may include PKC.
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PMID:Cholesterol-induced upregulation of angiotensin II and its effects on monocyte-endothelial interaction and superoxide production. 1178 66

We investigated the role of angiotensin II type 1 (AT(1)) receptors in angiotensin II-induced actin reorganization and the signaling pathways of the response in pleural mesothelial cells. The effects of angiotensin II on actin reorganization in pleural mesothelial cells were evaluated by dual fluorescence labeling of filamentous (F) and monomeric (G) actin with fluorescein isothiocyanate (FITC)-labeled phalloidin and Texas Red-labeled DNase I, respectively. Angiotensin II (10 microM) induced actin reorganization in the presence and the absence of extracellular Ca(2+). An angiotensin AT(1) receptor antagonist ([Sar(1),Ile(8)]angiotensin II) inhibited angiotensin II-induced actin reorganization. Pretreatment with C3 exoenzyme or tyrosine kinase inhibitors significantly reduced angiotensin II-induced actin reorganization. However, pertussis toxin, phosphatidylinositol-3-kinase and protein kinase C inhibitors had no effect on these responses. These results suggest that angiotensin II-induced actin reorganization in pleural mesothelial cells is extremely dependent on the angiotensin AT(1) receptor coupled with pertussis toxin-insensitive heterotrimeric G proteins, Rho GTPases and tyrosine phosphorylation pathways.
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PMID:Involvement of Rho and tyrosine kinase in angiotensin II-induced actin reorganization in mesothelial cells. 1183 42

Legumain (asparaginyl endopeptidase) was purified to homogeneity from bovine kidneys. The molecular mass of the purified enzyme was calculated to be 34000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis in the presence of beta-mercaptoethanol. The enzyme rapidly hydrolyzed the substrate Z-Ala-Ala-Asn-MCA and was strongly inhibited by N-ethylmaleimide, p-chloromercuribenzene-sulfonic acid, Hg(2+) and Cu(2+). The amino acid sequence of the first 26 residues of the enzyme was Gly-Gly-Lys-His-Trp-Val-Val-Ile-Val-Ala-Gly-Ser-Asn-Gly-Gln-Tyr-Asn-Tyr-Arg-His-Gln-Ala-Phe-Ala-Asp-His-. This sequence is highly homologous to the sequences in the N-terminal of pig kidney legumain. We screened a bovine kidney cortex cDNA library using a DNA probe that originated from rat legumain, and we determined the bovine kidney cDNA structure and deduced the amino acid sequence. The cDNA is composed 1934 bp and encodes 433 amino acids in the coding region. The enzyme was strongly stained in the proximal tubules of the rat kidney in an immunohistochemical study. Vitamin D-binding protein which is known to be a ligand to megalin existing in the proximal tubules, was cleaved in a limited proteolytic manner by bovine kidney legumain. These results suggested that legumain contributes to the processing of macromolecules absorbed by proximal tubule cells. The enzyme also cleaved an N-terminal synthetic peptide of bovine annexin II (Gly(24)-Ser-Val-Lys-Ala-Tyr-Thr(30)-Asn-Phe-Asp-Ala-Glu(35)-Arg-Asp(37)) at a position between Asn(31) and Phe(32). The amino-terminal domain of annexin II has p11 subunit binding sites and phosphorylation sites for both pp60(src) and protein kinase C. This suggests that legumain plays an important role in inactivation and degradation of annexin II, which is abundant in the receptor-recycling compartments of endosomes/lysosomes.
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PMID:Legumain from bovine kidney: its purification, molecular cloning, immunohistochemical localization and degradation of annexin II and vitamin D-binding protein. 1198 26

We previously cloned Na(+)/Ca(2+) exchanger (NCX1) from mesangial cells of salt-sensitive (SNCX = NCX1.7) and salt-resistant (RNCX = NCX1.3) Dahl/Rapp rats. The abilities of these isoforms to regulate cytosolic Ca(2+) concentration ([Ca(2+)](i)) were assessed in fura 2-loaded OK cells expressing the vector (VOK), RNCX (ROK), and SNCX (SOK). Baseline [Ca(2+)](i) was 98 +/- 20 nM (n = 12) in VOK and was significantly lower in ROK (44 +/- 5 nM; n = 12) and SOK (47 +/- 13 nM; n = 12) cells. ATP at 100 microM increased [Ca(2+)](i) by 189 +/- 55 nM (n = 12), 21 +/- 9 nM (n = 12), and 69 +/- 18 nM (n = 12) in VOK, ROK, and SOK cells, respectively. ATP (1 mM) or bradykinin (0.1 mM) caused large increases in [Ca(2+)](i) and ROK but not SOK cells were much more efficient in reducing [Ca(2+)](i) back to baseline levels. Parental Sprague-Dawley rat mesangial cells express both RNCX (SDRNCX) and SNCX (SDSNCX). SDRNCX and RNCX are identical at every amino acid residue, but SDSNCX and SNCX differ at amino acid 218 where it is isoleucine in SDSNCX and not phenylalanine. OK cells expressing SDSNCX (SDSOK) reduced ATP (1 mM)-induced [Ca(2+)](i) increase back to baseline at a rate equivalent to that for ROK cells. PKC downregulation significantly attenuated the rate at which ROK and SDSOK cells reduced ATP-induced [Ca(2+)](i) increase but had no effect in SOK cells. The reduced efficiency of SNCX to regulate [Ca(2+)](i) is attributed, in part, to the isoleucine-to-phenylalanine mutation at amino acid 218.
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PMID:Impaired ability of the Na+/Ca2+ exchanger from the Dahl/Rapp salt-sensitive rat to regulate cytosolic calcium. 1252 51

The sodium-calcium exchanger, NCX1, is a ubiquitously expressed membrane protein essential in calcium homeostasis for many cells including those in mammalian heart and brain. The function of NCX1 depends on subcellular ("local") factors, the phosphorylation state of NCX1, and the subcellular location of NCX1 within the cell. Here we investigate the molecular organization of NCX1 within the cardiac myocyte. We show that NCX1 is dynamically phosphorylated by protein kinase A (PKA)-dependent phosphorylation in vitro. We also provide evidence that the regulation of this phosphorylation is attributed to the existence of an NCX1 macromolecular complex. Specifically, we show that the macromolecular complex includes both the catalytic and regulatory subunits of PKA. However, only the RI regulatory subunit is found in this macromolecular complex, not RII. Other critical regulatory enzymes are also associated with NCX1, including protein kinase C (PKC) and two serine/threonine protein phosphatases, PP1 and PP2A. Importantly, the protein kinase A-anchoring protein, mAKAP, is found and its presence in the macromolecular complex suggests that these regulatory enzymes are coordinately positioned to regulate NCX1 as has been found in diverse cells for a number of channel proteins. Dual immunocytochemical staining showed the colocalization of NCX1 protein with mAKAP and PKA-RI proteins in cardiomyocytes. Finally, leucine/isoleucine zipper motifs have been identified as possible sites of interaction. Our finding of an NCX1 macromolecular complex in heart suggests how NCX1 regulation is achieved in heart and other cells. The existence of the NCX1 macromolecular complex may also provide an explanation for recent controversial findings.
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PMID:Sodium/calcium exchanger (NCX1) macromolecular complex. 1275 2

Signaling through the B cell receptor (BCR) of normal splenic B cells, as well as B cell lymphoma lines, can abrogate Fas-mediated apoptosis. Using the B lymphoma line A20.2J, BCR signaling immediately inhibited Fas-induced apoptosis upstream of caspase-8 activation, as determined by Ile-Glu-Thr-Asp-(IETD)ase activity and cleavage of the caspase-8 substrate Bid. Furthermore, following overexpression of a human Fas:FLICE construct, which directly induces caspase activation in a death-inducing signaling complex-independent manner, cells could not be protected through BCR stimulation.Co-incubation with cycloheximide partially reversed protection from apoptosis and increased Fas-stimulated initiator and effector caspase activation, suggesting new protein synthesis is necessary to induce protection upstream of caspase activation. Furthermore, co-incubation with a broad-spectrum protein kinase C (PKC) inhibitor, such as bisindolylmaleimide (Bis), also partially reversed protection from apoptosis, and examination of a panel of PKC inhibitors suggested a role for atypical isozymes in protection. Bis also acted to increase initiator and effector caspase activation upon anti-IgG and anti-Fas treatment. These data suggest that BCR-induced protection is being mediated upstream of initiator caspase activation, and is partially dependent upon both PKC family members and new protein synthesis.
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PMID:B cell receptor signaling mediates immediate protection from Fas-induced apoptosis upstream of caspase activation through an atypical protein kinase C isozyme and de novo protein synthesis. 1293 25

We characterized the tracheal and bronchial relaxation caused by proteinase-activated receptor-2 (PAR-2) activation in ddY mice and/or in wild-type and PAR-2-knockout mice of C57BL/6 background. Ser-Leu-Ile-Gly-Arg-Leu-amide (SLIGRL-NH(2)) and Thr-Phe-Leu-Leu-Arg-amide, PAR-2- and PAR-1-activating peptides, respectively, caused relaxation in the isolated ddY mouse trachea and main bronchus. The relaxation was abolished by specific inhibitors of cyclooxygenase (COX)-1, COX-2, mitogen-activated protein kinase kinase (MEK), and p38 MAP kinase. The MEK and p38 MAP kinase inhibitors did not affect prostaglandin E(2)-induced relaxation. Inhibitors of cytosolic Ca(2+)-dependent phospholipase A(2) (PLA), Ca(2+)-independent PLA(2), diacylglycerol lipase, tyrosine kinase, and protein kinase C exhibited no or only minor inhibitory effects on the PAR-mediated relaxation. Trypsin, a PAR-2 activator, and 2-furoyl-Leu-Ile-Gly-Arg-Leu-amide, a potent PAR-2-activating peptide, in addition to SLIGRL-NH(2), caused airway relaxation in wild-type C57BL/6 mice, as in ddY mice. In PAR-2-knockout mice, the peptide effects were absent and the potency of trypsin decreased. Desensitization of PAR-2 and/or PAR-1 greatly suppressed the relaxant effect of trypsin. The bronchial and tracheal tissues displayed distinct sensitivities toward trypsin and the PAR-2-activating peptides. Our data indicate an involvement of both COX-1 and COX-2, and the MEK-extracellular signal-regulated kinase and p38 MAP kinase signaling pathways in the PAR-2- and PAR-1-triggered relaxation of mouse airway tissue, and substantiate a role for PAR-2 in regulating both the trachea and bronchial responsiveness in the mouse lung.
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PMID:Proteinase-activated receptor-2-mediated relaxation in mouse tracheal and bronchial smooth muscle: signal transduction mechanisms and distinct agonist sensitivity. 1519 93

Specificity of phosphorylation is critical to signal transduction. Recent emphasis on colocalization of substrate and kinase has eclipsed emphasis on peptide specificity, i.e., kinase preference for particular amino acids surrounding the phosphorylation site. We describe an approach to determining peptide specificity by using positional scanning of biotinylated oriented peptide libraries and insights emerging from those determinations. We accurately determine preference (or disfavor) for residues at a given substrate position (such as P+2) by comparison of in vitro phosphorylation of peptide libraries differing by a single residue at that position. By analysis of all positions near the phosphorylation site, position-specific scoring matrices are generated and used both to understand the basis of specificity and to predict phosphorylation. PKC-delta and -zeta predictions have been validated rigorously by comparisons with measured phosphorylation. The results demonstrate specificity and sensitivity (80-90%) much better than the previous predictive method. These predictions can be accessed at http://mpr.nci.nih.gov. The accuracy of the specificity determination allows identification of an important difference in peptide specificity between these closely related kinases; Ile/Leu at the P-1 position is disfavored by PKC-zeta but not PKC-delta. Our findings and visual representation of peptide specificity highlight the importance of disfavored residues. Finally, analysis of 124 experimentally determined PKC sites from the literature demonstrates a very strong role of peptide specificity in many of those sites. Thus, position-specific scoring matrices generated by this method provide a foundation for quantitative analyses of kinase specificity and improved predictions of previously determined physiologically relevant phosphorylation sites.
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PMID:Kinase peptide specificity: improved determination and relevance to protein phosphorylation. 1535 39

Prepro-vasoactive intestinal peptide (VIP) mRNA codes for two neuropeptides: VIP and peptide histidine isoleucine (PHI). Two VIP receptors, shared with a similar affinity by pituitary adenylate cyclase-activating polypeptide (PACAP), have been cloned: VPAC(1) and VPAC(2). PHI binds to these receptors with a lower affinity. VPAC receptors are classically associated with a cAMP-dependent pathway, although other pathways, including calcium mobilization and protein kinase C activation have been described. We previously showed that intracerebral administration of the glutamate agonist ibotenate to postnatal day 5 mice induces white matter lesions mimicking human periventricular leukomalacia. In this model, coinjection of VIP protects against white matter lesions. This neuroprotection is independent from cAMP and is mediated by protein kinase C. Using this model, this study aimed to determine the receptor involved in VIP-induced neuroprotection. VIP effects were mimicked with a similar potency by VPAC(2) agonists and PHI but not by VPAC(1) agonists, PACAP 27, or PACAP 38. VIP neuroprotective effects were lost in mice lacking VPAC(2) receptor. In situ hybridization confirmed the presence of VPAC(2) mRNA in the postnatal day 5 white matter. When analyzed between embryonic life and adulthood, VIP-specific binding site density peaked at postnatal day 5. These data suggest that, in this model, VIP-induced neuroprotection is mediated by VPAC(2) receptors. The pharmacology of this VPAC(2) receptor seems unconventional because 1) PACAP does not mimic VIP effects, 2) PHI acts with a comparable potency, and 3) PACAP 27 modestly inhibited the VIP-specific binding, whereas for PHI or VIP, inhibition was complete.
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PMID:VPAC2 receptors mediate vasoactive intestinal peptide-induced neuroprotection against neonatal excitotoxic brain lesions in mice. 1587 42

Like other proteins involved in neurotransmitter transport, serotonin transporter (SERT) activity is regulated by multiple intracellular signal transduction pathways. The second intracellular loop (IL2) of SERT contains consensus sequences for cGMP-dependent protein kinase and protein kinase C. A 24-residue region of SERT including IL2, from Ile-270 through Ser-293, was analyzed by cysteine-scanning mutagenesis and chemical modification. 2-(Aminoethyl)methanethiosulfonate hydrobromide (MTSEA) failed to inhibit serotonin transport or binding of the cocaine analog 2beta-carbomethoxy-3beta-(4-[125I]iodophenyl)tropane (beta-CIT) in intact cells expressing these mutants, but it inactivated beta-CIT binding in membrane preparations. From the pattern of sensitivity, IL2 appears to extend from Trp-271 through Ile-290, a significantly longer region than that initially predicted by hydropathy analysis. Six mutants reacted with MTSEA much faster than the others, and the pattern of the more reactive mutations suggested that IL2 is in an alpha-helical conformation. Some of the mutants had significantly elevated transport rates, suggesting a possible mechanism for the regulation of SERT activity.
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PMID:Cysteine-scanning mutagenesis of serotonin transporter intracellular loop 2 suggests an alpha-helical conformation. 1599 10


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