Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Tumor necrosis factor-alpha (TNF) causes a spontaneously reversible increase in tight junction permeability. TNF was the only cytokine tested that produced this effect. The effect on transepithelial permeability proceeds in four distinct phases: 1) a 60- to 90-min delay from time of application of TNF, 2) a rapid decrease in transepithelial resistance, 3) a recovery of transepithelial resistance to control level within 1 h, and 4) a further increase of transepithelial resistance above control levels. The recovery of transepithelial resistance occurs with or without TNF in the culture medium. Different protein kinase inhibitors affected different phases of this overall process. The tyrosine kinase inhibitor genistein significantly blocked the TNF effect. Neither transcription nor protein synthesis was required for transepithelial permeability to increase, but were required for the recovery. After the tight junctions have opened at 2 h in response to TNF, a second application of TNF will not produce the effect again for at least 12 h. The tight junctions will, however, open in response to phorbol esters during this time frame. Electron microscopy studies using apically applied ruthenium red suggest that TNF action results in < 10% of the junctions having increased permeability at any given time during the resistance decrease. The role of epithelial barrier permeability changes in TNF action in vivo is discussed.
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PMID:Modulation of tumor necrosis factor-induced increase in renal (LLC-PK1) transepithelial permeability. 127 87

Tumor necrosis factor (TNF) is a 17-kDa protein produced by endotoxin-stimulated macrophages. We have demonstrated that recombinant human TNF activates human macrophages to kill intracellular bacteria of the Mycobacterium avium complex (MAC) in a dose-related manner. TNF also primed macrophages to produce superoxide anion (O2-) following treatment with phorbol esther PMA (0.1 micrograms/ml). To investigate the intracellular pathway involved in the TNF-mediated activation of mycobacteriostatic/mycobactericidal activity in macrophages, we used two different protein kinase C (PKC) inhibitors: H7 (10(-5)-10(7) M) and staurosporine (10(-7)-10(-9) M). Mellitin (1 and 100 mM) was used as a calmodulin inhibitor. Human peripheral blood-derived macrophages cultured for 7 days were treated with H7, mellitin, or staurosporine for 1 hr prior to incubation with TNF (10(3) U/ml). Twenty-four hours after treatment with TNF the O2- release was measured spectrophotometrically following exposure to PMA. Macrophages were infected with MAC and the viable intracellular bacilli were quantitated following 4 days of treatment with TNF. All PKC inhibitors suppressed O2- production after incubation with PMA. However, treatment with either PKC or calmodulin inhibitors did not influence the intracellular killing of M. avium by TNF-stimulated macrophages. Exposure of the macrophages to cGMP inhibitor but not to cAMP inhibitor significantly impaired the response to the stimulation with TNF. In contrast, incubation of macrophages with protein kinase A (PKA) had no effect on TNF-mediated mycobacteriostatic/mycobactericidal activity. These results suggest that the TNF-mediated mycobactericidal activity in cultured macrophages probably occurs by a PKC-independent mechanism.
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PMID:Tumor necrosis factor alpha stimulates mycobactericidal/mycobacteriostatic activity in human macrophages by a protein kinase C-independent pathway. 132 40

It has been reported that thrombomodulin (TM) expression in endothelial cells is modulated by various agents. We investigated cellular regulatory mechanisms for TM expression in human umbilical vein endothelial cells (HUVECs), incubated with agents, by measuring the time course changes in surface TM activity, total TM antigen in cell lysates, and TM mRNA levels. While dibutyryl cAMP (3 mM) increased TM mRNA levels in HUVECs and was followed by increased TM activity, dibutyryl cGMP had no effect on TM activity. Phorbol myristate acetate (PMA) induced rapid loss of surface TM activity (approximately 8 h) and later increased TM mRNA levels between 4 h and 40 h (maximum at 24 h), resulting in biphasic effects on TM activity. Tumor necrosis factor or interleukin-1 beta suppressed surface TM activity and TM mRNA levels. Internalization/degradation of TM in HUVECs incubated with PMA or cytokines was suggested by co-culture with chloroquine. The decrease in surface TM activity observed was not caused by the release of TM molecules from the cells into the conditioned media. These results suggest that TM activity in HUVECs is modulated by independent mechanisms involving cytoplasmic TM mRNA levels and internalization/degradation of TM molecules. These regulatory mechanisms may involve protein kinase A and protein kinase C-dependent mechanisms but are independent of protein kinase G.
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PMID:Regulatory mechanisms for thrombomodulin expression in human umbilical vein endothelial cells in vitro. 164 58

Tumor necrosis factor (TNF) production following X-irradiation has been implicated in the biological response to ionizing radiation. Protein kinase C (PKC) is suggested to participate in TNF transcriptional induction and X-ray-mediated gene expression. We therefore studied radiation-mediated TNF expression in HL-60 cells with diminished PKC activity produced by either pretreatment with protein kinase inhibitors or prolonged 12-O-tetradecanoylphorbol-13-acetate treatment. Both treatments resulted in attenuation of radiation-mediated TNF induction. Consistent with these results, we found no detectable induction of TNF expression following X-irradiation in the HL-60 variant deficient in PKC-mediated signal transduction. The rapid activation of PKC following gamma-irradiation was established using an in vitro assay measuring phosphorylation of a PKC specific substrate. A 4.5-fold increase in PKC activity occurred 15 to 30 s following irradiation, which declined to baseline at 60 s. Two-dimensional gel electrophoresis of phosphoproteins extracted from irradiated cells demonstrated in vivo phosphorylation of the PKC specific substrate Mr 80,000 protein at 45 s following X-irradiation. These findings indicate that signal transduction via the PKC pathway is required for the induction of TNF gene expression by ionizing radiation.
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PMID:Tumor necrosis factor gene expression is mediated by protein kinase C following activation by ionizing radiation. 187 1

Protein phosphorylation is central to multiple regulatory processes in cells. Tumor necrosis factor (TNF), a cytokine synthesized by macrophages, effects polymorphonuclear leukocyte (neutrophil) chemotaxis, induces superoxide anion generation, and mediates neutrophil adhesion to endothelial cells. Although protein phosphorylation is almost certainly involved in many TNF-mediated neutrophil functions, little is known about TNF's impact on neutrophil protein phosphorylation. Therefore, we studied human recombinant TNF-alpha-induced protein phosphorylation in human neutrophils. Neutrophils were preincubated with 32PO(4)2- and treated with a variety of stimulatory agents. One- and two-dimensional polyacrylamide gel electrophoresis was used to analyze phosphorylated proteins. Phosphoaminoacids were identified by two-dimensional thin layer chromatography electrophoresis. The findings were as follows: (1) TNF induces the phosphorylation of two 16-kD proteins (pI = 5.9 and 6.1) by 5- to 6-fold, and a 57-kD protein (pI = 5.8) by 3- to 4-fold compared with untreated neutrophils; (2) these proteins are phosphorylated as early as 15 min after stimulation with TNF, and phosphorylation is induced by concentrations of TNF as low as 1 ng/ml (10 U/ml); (3) TNF induces the phosphorylation of proteins at either serine or threonine residues and not at tyrosine; (4) TNF-stimulated neutrophils show a unique pattern of protein phosphorylation when compared to neutrophils treated with formylmethionylleucylphenylalanine; (5) lipopolysaccharide does not induce protein phosphorylation in neutrophils; (6) a 16-kD protein is phosphorylated in response to TNF in neutrophils but not in mononuclear cells; and (7) protein kinase inhibitors appear to have no effect on TNF-induced protein phosphorylation. Thus, the mechanism of action of TNF on neutrophils may involve protein phosphorylation.
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PMID:Tumor necrosis factor-induced protein phosphorylation in human neutrophils. 191 Aug 14

Tumor necrosis factor (TNF) and interleukin-1 (IL-1) enhanced the phosphorylation of identical cytosolic 65 kDa protein (P65 or l-plastin) and 74 kDa protein (P74) at serine residues in human peripheral blood mononuclear cells (PBMC). The isoelectric points of P65 and P74 were 5.6 and 4.7 to 5.0, respectively. The phosphorylation of these proteins increased with a few minutes and reached maximal levels of approximately 3 times the unstimulated levels by 10 minutes. The phosphorylation of P65 and P74 was extensively enhanced by a potent protein kinase C (PKC) activator, PMA. However, there was no translocation of PKC from cytosol to membrane in PBMC that was stimulated with either TNF or IL-1, which suggests that PKC does not participate in TNF or IL-1 signal transduction. cAMP dependent protein (PKA) activators, forskolin and PGE2, failed to increase the phosphorylation, which is in agreement with the data showing that neither TNF nor IL-1 increased cAMP levels in PBMC. These results suggest that induction of phosphorylation of P65 and P74 by TNF and IL-1 is not mediated by PKC and PKA but may be mediated by another protein kinase and result in overlapping of biological activities between TNF and IL-1.
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PMID:Enhanced phosphorylation of 65 and 74 kDa proteins by tumor necrosis factor and interleukin-1 in human peripheral blood mononuclear cells. 196 46

Ceramide produced by sphingomyelinases (SMases) has been recognized as an important second messenger in growth factor receptor signaling. Tumor necrosis factor (TNF), through binding to the 55 kDa TNF receptor (TNF-R55), rapidly activates two distinct types of SMase, a membrane-associated neutral (N-)SMase, and an endosomal acidic (A)-SMase. N-SMase and A-SMase are activated independently by different cytoplasmic domains of TNF-R55. Each type of SMase specifically couples to select pathways of TNF signaling. Ceramide generated by N-SMase directs the activation of proline-directed serine/threonine protein kinase(s) and phospholipase A2. In contrast, A-SMase triggers the activation of NF-kappa B. No apparent crosstalk was detected between N-SMase and A-SMase pathways, indicating that ceramide action depends on the topology of its production. These results suggest that N-SMase and A-SMase control important yet dissociable and nonoverlapping pathways of TNF receptor signal transduction.
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PMID:Functional dichotomy of neutral and acidic sphingomyelinases in tumor necrosis factor signaling. 792 51

Tumor necrosis factor (TNF) and interleukin-1 (IL-1) are cytokines with pleiotropic biological activities, exerting a broad range of overlapping biological functions. The redundancy of TNF and IL-1 activities may be based on the utilization of shared key components of intracellular signaling pathways. Two lipid second messengers have been found to transmit TNF and IL-1 intracellular signals: 1,2-diacylglycerol (DAG), generated by a phosphatidylcholine-specific phospholipase C, and ceramide, generated by sphingomyelinase (SMase). DAG is a well established activator of the important signaling system protein kinase C (PKC), which appears to mediate various cellular responses to TNF or IL-1. In addition, it is obvious that DAG also activates other enzyme systems like acidic sphingomyelinase. SMases have been implicated in a number of TNF responses, including stimulation of cell growth and differentiation, as well as triggering cytotoxicity and apoptosis. The metabolic active cleavage product of SMase, ceramide, is a novel multifunctional lipid second messenger capable of inducing various signaling systems. Both cytokines, TNF and IL-1, stimulate a neutral,plasma membrane-associated SMase that leads to stimulation of a protein kinase and eventually to activation of the mitogen-activated protein (MAP) kinase cascade and phospholipase A2. Ceramide is also capable of stimulating a cytosolic protein phosphatase. PKC plays a role in activation of the nuclear transcription factor AP-1, and the DAG-regulated acidic SMase is involved in transducing TNF signals to the cell nucleus via activation of the nuclear transcription factor NF-kappa B.
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PMID:The role of diacylglycerol and ceramide in tumor necrosis factor and interleukin-1 signal transduction. 796 60

Tumor necrosis factor (TNF) binds two distinct cell surface receptors designated p60 and p80. Our previous studies indicate that a protein kinase from U-937 cells binds to and phosphorylates the p60 receptor. While the p80 receptor is phosphorylated in vivo, no association of a protein kinase has been described. We employed a fusion protein comprising of glutathione S-transferase and the cytoplasmic domain of the p80 receptor (GST-p80CD) to identify cellular proteins that might associate with this receptor. From 35S- and 32P-labeled cells, a protein of 59 kDa bound specifically to GST-p80CD. In vitro kinase reactions indicated that serine/threonine protein kinase activity associated with GST-p80CD and causes its phosphorylation. Additionally, a 59-kDa phosphoprotein was also identified after kinase reactions of proteins bound to GST-p80CD. This kinase activity required either Mg2+ or Mn2+ for optimal activity, and it phosphorylated myelin basic protein, histone H2B, and also the cytoplasmic domain of the p60 receptor. Treatment of cells with TNF increased the p80 receptor-associated kinase activity by 200%. In summary, our results provide evidence of a novel ligand-activated serine/threonine protein kinase that associates with the cytoplasmic domain of the p80 receptor and causes the phosphorylation of both forms of the TNF receptor. This p80 TNF receptor-associated protein and the associated kinase described here are referred to as p80-TRAP and p80-TRAK, respectively.
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PMID:Physical and functional association of a serine-threonine protein kinase to the cytoplasmic domain of the p80 form of the human tumor necrosis factor receptor in human histiocytic lymphoma U-937 cells. 805 Oct 45

Tumor necrosis factor (TNF) has been shown to bind two distinct receptors, designated p60 and p80, with high affinity, resulting, within minutes, in phosphorylation of several proteins. The receptors themselves do not exhibit protein kinase activity nor have any associated proteins been identified. We employed the glutathione-S-transferase (GST) fusion protein system consisting of the cytoplasmic domain of p60 (GST-p60CD delta 1) as a probe to help us identify receptor-associated proteins from human histiocytic lymphoma U-937 cells. We found that a protein of approximately 52 kDa (pp52) bound to GST-p60CD delta 1 from [35S]methionine- and 32P-labeled cells. The associated protein was phosphorylated on serine and threonine residues. Furthermore, we identified serine/threonine kinase activity associated with p60CD delta 1 that required either Mn2+ or Mg2+ for optimal activity. The preferred substrates for this kinase, in addition to p60CD delta 1, included casein and histone H1, but not histone H2B, myelin basic protein, enolase, or the cytoplasmic domain of p80. As was the case in U-937 cells, p60CD delta 1-associated kinase activity was also identified in human breast adenocarcinoma MCF-7 cells and human foreskin fibroblasts. TNF stimulation of MCF-7 and foreskin fibroblasts for 5-15 min induced approximately 50 and 240% increases in phosphorylation of p60CD delta 1, respectively. Thus, our results provide the first evidence for protein kinase activity that is specifically associated with the cytoplasmic domain of the p60 form of the TNF receptor and causes its phosphorylation. This p60 TNF receptor-associated protein and the associated kinase described here are referred to as p60-TRAP and p60-TRAK, respectively.
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PMID:Identification of a protein kinase associated with the cytoplasmic domain of the p60 tumor necrosis factor receptor. 805 Nov 24


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