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)

Tetraspanins are integral membrane proteins involved in a variety of physiological and pathological processes. In cancer, clinical and experimental studies have reported a link between tetraspanin expression levels and metastasis. Tetraspanins play a role as organizers of a molecular network of interactions, the "tetraspanin web". Here, we have performed a proteomic characterization of the tetraspanin web using a model of human colon cancer consisting of two cell lines derived from primary tumor and metastasis from the same patient. The tetraspanin complexes were isolated after immunoaffinity purification and the proteins were identified by MS using LC-ESI-MS/MS and MALDI-FTICR. The high resolution and mass accuracy of FTICR MS allowed reliable identification using mass finger printing with only two peptides. Thus, it could be used to resolve the composition of complex peptide mixtures from membrane proteins. Different types of membrane proteins were identified, including adhesion molecules (integrins, Lu/B-CAM, GA733 proteins), receptors and signaling molecules (BAI2, PKC, G proteins), proteases (ADAM10, TADG15), and membrane fusion proteins (syntaxins) as well as poorly characterized proteins (CDCP1, HEM-1, CTL1, and CTL2). Some components were differentially detected in the tetraspanin web of the two cell lines. These differences may be relevant for tumor progression and metastasis.
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PMID:Proteomic analysis of the tetraspanin web using LC-ESI-MS/MS and MALDI-FTICR-MS. 1640 22

Constitutive and PKC-regulated alpha-secretase pathways have been reported to produce the secreted form of alpha-secretase-derived APP (sAPPalpha). Here, we examined putative role of furin in the regulation of alpha-secretase activity in vitro and in vivo. Overexpression of the prodomain of furin and infection with a furin-specific inhibitor significantly reduced the levels of sAPPalpha regardless of PKC activity, whereas total APP levels remained unchanged. Furin mRNA levels in the brains of AD patients and Tg2576 mice were significantly lower than those in controls, whereas ADAM10 and TACE mRNA levels were much alike between Tg2576 and littermate mice. Moreover, the injection of furin-adenovirus into Tg2576 mouse brains markedly increased alpha-secretase activity and reduced beta-amyloid protein (Abeta) production in infected brain regions. Our results suggest that furin enhances alpha-secretase activity via the cleavage of ADAM10 and TACE, and that attenuated furin activity is connected to the production of Abeta.
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PMID:Furin is an endogenous regulator of alpha-secretase associated APP processing. 1694 50

CD30 is a transmembrane protein selectively overexpressed on many human lymphoma cells and therefore an interesting target for antibody-based immunotherapy. However, binding of therapeutic antibodies stimulates a juxtamembrane cleavage of CD30 leading to a loss of target antigen and an enhanced release of the soluble ectodomain of CD30 (sCD30). Here, we show that sCD30 binds to CD30 ligand (CD153)-expressing non-target cells. Because antibodies bind to sCD30, this results in unwanted antibody binding to these cells via sCD30 bridging. To overcome shedding-dependent damage of normal cells in CD30-specific immunotherapy, we analyzed the mechanism involved in the release. Shedding of CD30 can be enhanced by protein kinase C (PKC) activation, implicating the disintegrin metalloproteinase ADAM17 but not free cytoplasmic calcium. However, antibody-induced CD30 shedding is calcium dependent and PKC independent. This shedding involved the related metalloproteinase ADAM10 as shown by the use of the preferential ADAM10 inhibitor GI254023X and by an ADAM10-deficient cell line generated from embryonically lethal ADAM10(-/-) mouse. In coculture experiments, the antibody-induced transfer of sCD30 from the human Hodgkin's lymphoma cell line L540 to the CD30-negative but CD153-expressing human mast cell line HMC-1 was inhibited by GI254023X. These findings suggest that selective metalloproteinase inhibitors blocking antibody-induced shedding of target antigens could be of therapeutic value to increase the specificity and reduce side effects of immunotherapy with monoclonal antibodies.
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PMID:ADAM10 inhibition of human CD30 shedding increases specificity of targeted immunotherapy in vitro. 1721 Jul 15

The cellular prion protein (PrP(c)) undergoes a physiological processing yielding the N-terminal fragment referred to as N1, the production of which can be constitutive or protein kinase C regulated. We show that activation of endogenous muscarinic receptors by carbachol and by the M1-selective agonist AF267B increases N1 recovery in an atropine-sensitive manner, in mouse embryonic primary neurons. To identify the muscarinic receptor subtype involved, we used human embryonic kidney HEK293 (HEK) cells stably overexpressing M1, M2, M3, or M4 receptor subtype. Carbachol and the selective M1 agonist AF267B dose dependently increased N1 release by HEK-M3 and HEK-M1 cells, respectively, whereas carbachol did not modify N1 production by HEK-M2 or HEK-M4 cells. We demonstrate that the increase of N1 was not attributable to modified trafficking to the membrane of either PrP(c) or the disintegrin metalloproteases ADAM10 or ADAM17. Furthermore, we establish that carbachol affects the overall phosphorylation of ADAM17 on its threonine and tyrosine but not serine residues, whereas levels of phosphorylated ADAM9 were not affected. Interestingly, carbachol also increases the hydrolysis of the fluorimetric substrate JMV2770, which mimicked the sequence encompassing the N1 site cleavage and was shown previously to behave as an ADAM protease substrate. Mutations of threonine 735 but not of tyrosine 702 of the ADAM17 cytoplasmic tail abolishes the carbachol-induced increase of N1, ADAM17 phosphorylation, and JMV2770-hydrolyzing activity in M1- and M3-expressing HEK293 cells. Thus, our data provide strong evidence that muscarinic receptor activation increases the physiological processing of PrP(c) by upregulating the phosphorylation state and activity of ADAM17 protease.
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PMID:M1 and M3 muscarinic receptors control physiological processing of cellular prion by modulating ADAM17 phosphorylation and activity. 1742 86

Alpha-secretase (alpha-secretase), cleaves the amyloid precursor protein (APP) within the amyloid-beta (Abeta) sequence, resulting in the release of a secreted fragment of APP (alphaAPPs) and precluding Abeta generation. We investigated the effects of the acetylcholinesterase inhibitor, huperzine A (Hup A), on APP processing and Abeta generation in human neuroblastoma SK-N-SH cells overexpressing wild-type human APP695. Hup A dose-dependently (0-10 microM) increased alphaAPPs release. Therefore, we evaluated two alpha-secretase candidates, a disintegrin and metalloprotease (ADAM) 10 and ADAM17 in Hup A-induced non-amyloidogenic APP metabolism. Hup A enhanced the level of ADAM10, and the inhibitor of tumor necrosis factor-alpha converting enzyme (TACE)/ADAM17 inhibited the Hup A-induced rise in alphaAPPs levels, further suggesting Hup A directed APP metabolism toward the non-amyloidogenic alpha-secretase pathway. Hup A had no effect on Abeta generation in this cell line. The steady-state levels of full-length APP and cell viability were unaffected by Hup A. Alpha-APPs release induced by Hup A treatment was significantly reduced by muscarinic acetylcholine receptor antagonists (particularly by an M1 antagonist), protein kinase C (PKC) inhibitors, GF109203X and calphostin C, and the mitogen-activated kinase kinase (MEK) inhibitors, U0126 and PD98059. Furthermore, Hup A markedly increased the phosphorylation of p44/p42 mitogen-activated protein (MAP) kinase, which was blocked by treatment with U0126 and PD98059. In addition, Hup A inhibited acetylcholinesterase activity by 20% in neuroblastoma cells. Our results indicate that the activation of muscarinic acetylcholine receptors, PKC and MAP kinase may be involved in Hup A-induced alphaAPPs secretion in neuroblastoma cells and suggest multiple pharmacological mechanisms of Hup A regarding the treatment of Alzheimer's disease (AD).
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PMID:Huperzine A regulates amyloid precursor protein processing via protein kinase C and mitogen-activated protein kinase pathways in neuroblastoma SK-N-SH cells over-expressing wild type human amyloid precursor protein 695. 1794 34

Retinoic acid stimulates alpha-secretase processing of amyloid precursor protein (APP) and decreases beta-secretase cleavage that leads to amyloid-beta formation. Here, we investigated the effect of retinoic acid on the two putative alpha-secretases, the disintegrin metalloproteinases ADAM10 and TACE, and the beta-site cleaving enzyme BACE1, in human neuroblastoma SH-SY5Y cells. Western blot analysis showed that exposure to retinoic acid resulted in significantly increased levels of ADAM10 and TACE, suggesting that regulation of alpha-secretases causes the effects on APP processing. The presence of the phosphatidylinositol 3-kinase inhibitor LY 294002 selectively reduced the effect on ADAM10 protein levels but not on ADAM10 mRNA levels as determined by RT-PCR. On the other hand, the effect on TACE was shown to be dependent on protein kinase C, since it was completely blocked in the presence of the inhibitor bisindolylmaleimide XI. Our data indicate that different signalling pathways are involved in retinoic acid-induced up-regulation of the secretases.
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PMID:PI3-K- and PKC-dependent up-regulation of APP processing enzymes by retinoic acid. 1798 85

Accumulation of amyloid beta-peptides (Abeta) in the brain is believed to contribute to the development of Alzheimer disease (AD). Abeta, a 40-42 amino acid-comprising proteolytical fragment of the amyloid precursor protein (APP), is released from APP by sequential cleavages via beta- and gamma-secretases. However, the predominant route of APP processing consists of successive cleavages by alpha- and gamma-secretases. Alpha-secretase attacks APP inside the Abeta sequence, and therefore prevents formation of neurotoxic Abeta. After cleavage by alpha-secretase, the soluble N-terminal domain of APP, which possesses neurotrophic and neuroprotective properties, is released. In AD patients, a decrease in alpha-secretase processing of APP has been found and therefore, strategies to improve alpha-secretase activity are obvious. Several years after descriptive reports on alpha-secretase, the responsible enzymes have been identified to belong to the family of A Disintegrin And Metalloproteinase (ADAM). Three of these membrane-anchored zinc-dependent metalloproteinases, ADAM10 as well as ADAM17 and presumably also ADAM9 display alpha-secretase activity. Since the individual knock-out of these proteinases in neither case completely prevented alpha-secretase processing of APP, it seems likely that different ADAMs are compensating mutually, and under different conditions may contribute to alpha-secretase cleavage of APP. In addition to ADAMs, perhaps other membrane-associated metalloproteinases contribute to the shedding of APP. Stimulation of alpha-secretase activities can be achieved via several signaling cascades including phospholipase C, phosphatidylinositol 3-kinase and serine/threonine-specific kinases such as protein kinases C, and mitogen activated protein kinases. Direct activation of protein kinase C and stimulation of distinct G protein-coupled receptors are known to increase alpha-secretase processing of APP. Agonists for M1 muscarinic acetylcholine receptors and serotonin 5-HT4 receptors are currently in clinical trials to test their efficiency in the treatment of AD.
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PMID:A closer look at alpha-secretase. 1839 3

Vascular endothelial (VE)-cadherin is the major adhesion molecule of endothelial adherens junctions. It plays an essential role in controlling endothelial permeability, vascular integrity, leukocyte transmigration, and angiogenesis. Elevated levels of soluble VE-cadherin are associated with diseases like coronary atherosclerosis. Previous data showed that the extracellular domain of VE-cadherin is released by an unknown metalloprotease activity during apoptosis. In this study, we used gain-of-function analyses, inhibitor studies, and RNA interference experiments to analyze the proteolytic release of VE-cadherin in human umbilical vein endothelial cells (HUVECs). We found that VE-cadherin is specifically cleaved by the disintegrin and metalloprotease ADAM10 in its ectodomain, releasing a soluble fragment and generating a carboxyl-terminal membrane-bound stub, which is a substrate for a subsequent gamma-secretase cleavage. This ADAM10-mediated proteolysis could be induced by Ca(2+) influx and staurosporine treatment, indicating that ADAM10-mediated VE-cadherin cleavage contributes to the dissolution of adherens junctions during endothelial cell activation and apoptosis, respectively. In contrast, protein kinase C activation or inhibition did not modulate VE-cadherin processing. Increased ADAM10 expression was functionally associated with an increase in endothelial permeability. Remarkably, our data indicate that ADAM10 activity also contributes to the thrombin-induced decrease of endothelial cell-cell adhesion. Moreover, knockdown of ADAM10 in HUVECs as well as in T cells by small interfering RNA impaired T-cell transmigration. Taken together, our data identify ADAM10 as a novel regulator of vascular permeability and demonstrate a hitherto unknown function of ADAM10 in the regulation of VE-cadherin-dependent endothelial cell functions and leukocyte transendothelial migration.
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PMID:ADAM10 regulates endothelial permeability and T-Cell transmigration by proteolysis of vascular endothelial cadherin. 1849 10

The cellular prion protein (PrP(c)) undergoes a physiological cleavage between amino acids 111 and 112, thereby leading to the secretion of an amino-terminal fragment referred to as N1. This proteolytic event is either constitutive or regulated by protein kinase C (PKC) and is operated by the disintegrins ADAM9/ADAM10 or ADAM17 respectively. We recently showed that the stimulation of the M1/M3 muscarinic receptors potentiates this cleavage via the phosphorylation and activation of ADAM17. We have examined the contribution of various PKC isoforms in the regulated processing of PrP(c). First we show that the PDBu- and carbachol-stimulated N1 secretions are blocked by the general PKC inhibitor GF109203X. We establish that HEK293 and human-derived rhabdhomyosarcoma cells over-expressing constitutively active PKCalpha, PKCdelta or PKCepsilon, but not PKCzeta, produce increased amounts of N1 and harbor enhanced ability to hydrolyze the fluorimetric substrate of ADAM17, JMV2770. Conversely, over-expression of the corresponding dominant negative proteins abolishes PDBU-stimulated N1 secretion and restores N1 to levels comparable to constitutive production. Moreover, deletion of PKCalpha lowers N1 recovery in primary cultured fibroblasts. Importantly, mutation of threonine 735 of ADAM17 significantly lowers the PDBu-induced N1 formation while transient over-expression of constitutively active PKCalpha, PKCdelta or PKCepsilon, but not PKCzeta, induced both the phosphorylation of ADAM17 on its threonine residues and N1 secretion. As a corollary, T735A mutation concomitantly reversed PKCalpha-, PKCdelta- and PKCepsilon-induced ADAM17 phosphorylation and N1 recovery. Finally, we established that PKCepsilon-dependent N1 production is fully prevented by ADAM17 deficiency. Altogether, the present results provide strong evidence that the activation of PKCalpha, delta and epsilon, but not zeta, isoforms leads to increased N1 secretion via the phosphorylation and activation of ADAM17, a process that likely accounts for M1/M3 muscarinic receptors-mediated control of N1 production.
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PMID:Isoform-specific contribution of protein kinase C to prion processing. 1872 32

Deprenyl is a selective B-type monoamine oxidase inhibitor and a neuroprotective agent that has been used to slow the progress of Alzheimer's disease for many years. We previously demonstrated that deprenyl could stem amyloid precursor protein processing (APP) toward the non-amyloidogenic pathway through mitogen activated protein kinase (MAPK) and protein kinase C (PKC)-dependent signaling pathways [Yang, H.Q., Ba, M.W., Ren, R.J., Zhang, Y.H., Ma, J.F., Pan, J., Lu, G.Q., Chen, S.D., 2007a. Mitogen activated protein kinase and protein kinase C mediated promotion of sAPPalpha by deprenyl. Neurochem. Int. 50, 74-82.]. The experiment here further showed that deprenyl could increase alpha-secretase activity in a dose-dependent manner in PC12 cells. Deprenyl increased alpha-secretase activity can be partially blocked by pretreatment with brefeldin A, an intracellular protein transport inhibitor, suggesting involvement of protein trafficking in deprenyl regulated alpha-secretase activity. In accordance with this, the experiment showed that brefeldin A also decreased sAPPalpha release induced by deprenyl. Deprenyl promoted ADAM10 transported to the membrane fraction, and this effect was blocked by pretreatment with brefeldin A. The immunocytochemistry staining revealed that deprenyl promoted colocalization of ADAM10 with PKCalpha and PKCepsilon isoforms. These data suggest a novel pharmacological mechanism in which deprenyl increased alpha-secretase activity via protein trafficking related mechanism.
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PMID:Involvement of protein trafficking in deprenyl-induced alpha-secretase activity regulation in PC12 cells. 1932 34


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