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)

Ten protein kinases have been assayed for their ability to phosphorylate in vitro the recombinant bovine PrP (25-242) (rbPrP). Substantial phosphorylation was observed with PKC, CK2, and two tyrosine kinases, Lyn and c-Fgr. With regard to CK2, phosphorylation occurs at Ser 154 with a stoichiometry of about 0.1 mol phosphate/mol rbPrP, which is doubled by mild heat treatment of rbPrP. Heat also reduces the overall protein ellipticity, suggesting that reversibly unfolded conformers are more susceptible to phosphorylation. Our data disclose the possibility that phosphorylation might modulate PrP biological activity.
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PMID:Susceptibility of the prion protein to enzymic phosphorylation. 1079 98

Cellular prion protein (PrP(c)) undergoes a proteolytic attack at the 110/111 downward arrow112 peptide bond, whereas the PrP isoform (PrP(res)) that accumulates in the brain tissue in Creutzfeldt-Jakob disease reveals an alternate cleavage site at about residue 90. Interestingly, the normal processing of PrP occurs inside the 106-126 amino acid region thought to be responsible for the neurotoxicity of the pathogenic prions, whereas PrP(res) cleavage preserves this potentially toxic domain. Therefore, any molecular mechanisms leading to enhanced cleavage at the 110/111 downward arrow112 peptide bond could be of potential interest. We set up TSM1 neurons and HEK293 stable transfectants overexpressing the wild-type or 3F4-tagged murine PrP(c), respectively. Both mock-transfected and PrP(c)-expressing cell lines produced an 11-12-kDa PrP fragment (referred to as N1), the immunological characterization of which strongly suggests that it corresponds to the N-terminal PrP(c) fragment derived from normal processing. We have established that the recovery of secreted N1 is increased by the protein kinase C agonists PDBu and PMA in a time- and dose-dependent manner in both cell lines. In contrast, secretion of N1 remains unaffected by the inactive PDBu analog alphaPDD and by the protein kinase A effectors dibutyryl cAMP and forskolin. Overall, our data indicate that the normal processing of PrP(c) is up-regulated by protein kinase C but not protein kinase A in human cells and murine neurons.
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PMID:Phorbol ester-regulated cleavage of normal prion protein in HEK293 human cells and murine neurons. 1095 79

The cellular prion protein (PrP(C)) is a glycosylphosphatidylinositol (GPI)-anchored protein. We investigated whether PrP(C) can move from one cell to another cell in a cell model. Little PrP(C) transfer was detected when a PrP(C) expressing human neuroblastoma cell line was cultured with the human erythroleukemia cells IA lacking PrP(C). Efficient transfer of PrP(C) was detected with the presence of phorbol 12-myristate 13-acetate, an activator of protein kinase C. Maximum PrP(C) transfer was observed when both donor and recipient cells were activated. Furthermore, PrP(C) transfer required the GPI anchor and direct cell to cell contact. However, intercellular protein transfer is not limited to PrP(C), another GPI-anchored protein, CD90, also transfers from the donor cells to acceptor cells after cellular activation. Therefore, this transfer process is GPI-anchor and cellular activation dependent. These findings suggest that the intercellular transfer of GPI-anchored proteins is a regulated process, and may have implications for the pathogenesis of prion disease.
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PMID:Intercellular transfer of the cellular prion protein. 1235 24

We have previously established that cellular prion PrP(c) elicited p53-dependent caspase 3 activation in various transfected cells and primary cultured neurons. Although we showed that PrP(c) modulates p53 expression at both transcriptional and post-transcriptional levels, it remained unclear as to whether cellular prion signals at the membrane to trigger intracellular messages or if prion proapoptotic activity necessitated its translocation into the cytoplasm. Here, we compare the processing and cell death-related functions of PrP(c) with those of a mutated PrP(c) protein (N-3F4 MoPrP(c)) in which three basic N-terminal residues responsible for PrP(c) internalization had been mutated. As expected, N-3F4 MoPrP(c) remains exclusively located at the membrane, whereas PrP(c) partitions between membrane-associated and intracellular compartments, but both, proteins undergo constitutive and protein kinase C-regulated disintegrin-mediated proteolysis, leading to N1 fragment production. Unlike PrP(c), N-3F4 MoPrP(c) expression does not induce caspase 3 activation after stimulation by staurosporine and was inert on p53 expression and promoter transactivation in both human cells and TSM1 mouse neurons. Interestingly, PrP(c)-induced caspase 3 activation was closely linked to its endocytosis. This phenotype was enhanced by proteasomal inhibition and prevented by sucrose treatment. Accordingly, immunohistochemical analysis showed that protection towards degradation increased intracellular PrP(c)-like immunoreactivity, while sucrose treatments fully abolished PrP(c) intracellular expression and co-localization with transferrin. Altogether, we, establish here, using combined biochemical, mutational and cell biology approaches, that the caspase 3 activation associated with cellular prion is closely related to its ability to undergo endocytosis. This is, to our knowledge, the first direct description of an endocytosis-dependent PrP(c)-associated function.
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PMID:Combined pharmacological, mutational and cell biology approaches indicate that p53-dependent caspase 3 activation triggered by cellular prion is dependent on its endocytosis. 1574 58

Glutamate is the main excitatory neurotransmitter in the cerebral cortex. Altered glutamatergic transmission has been suggested as having a central role in many neurodegenerative diseases. Metabotropic glutamate receptors (mGluRs) are coupled to intracellular signal transduction via G proteins, and they mediate slower responses than ionotropic glutamate receptors. Group I mGluRs are positively coupled to phospholipase C beta1 (PLCbeta1). Creutzfeldt-Jakob disease (CJD) is a human transmissible spongiform encephalopathy associated with a dysfunction in the membrane glycoprotein PrP which is converted into an abnormal isoform, with a predominant beta-sheet structure, that is pathogenic and partially resistant to protease digestion. Proteins associated with the signal transduction of group I mGluRs were examined in the frontal cortex (area 8) of 12 cases with sCJD and four age-matched controls, by means of gel electrophoresis and Western blotting of total homogenates. Densitometric analysis of the bands demonstrated decreased expression levels of PLCbeta1 and PLCgamma, a non-related phospholipase which is a substrate of tyrosine kinase, in CJD cases when compared with controls. Novel protein kinase C delta (nPKCdelta) has also been found to be significantly decreased in CJD cases. However, no modifications in mGluR1 cPKCalpha expression levels are found in CJD when compared with controls. No modifications in PLCbeta1 solubility in PBS-, deoxycholate- and sodium dodecylsulphate-soluble fractions have been observed in CJD when compared with controls. Finally, no interactions between PLCbeta1 and PrP, as revealed by immunoprecipitation assays, have been found in CJD and controls. The present results show, for the first time, reduced expression levels of phospholipases, particularly PLCbeta1, which may interfere with group I mGluR signaling in the cerebral cortex in CJD. These abnormalities are not the result of abnormal PLC solubility or interactions with PrP. Selective involvement of group I mGluRs may have functional effects on glutamatergic transmission modulation and processing in CJD.
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PMID:Metabotropic glutamate receptor/phospholipase C pathway: a vulnerable target to Creutzfeldt-Jakob disease in the cerebral cortex. 1574 37

Alzheimer's disease, as well as most of other neurodegenerative disorders, is characterized by the deposition of insoluble proteinaceous aggregates. Hence, any intervention aimed at reducing this process could be envisioned as a therapeutic way to slow down the disease. In the case of Alzheimer's disease, the culprit protein is the 40-43 amino acid-long amyloid beta peptide (Abeta). This fragment is generated from the beta-amyloid precursor protein (betaAPP) by two distinct enzymes, namely the beta- and the gamma-secretases. In the past years, a tremendous effort has been made to develop potent and specific inhibitors of these proteolytic activities. Beside these Abeta-forming proteases, a third cleavage performed by the so-called alpha-secretase takes place in the middle of the Abeta sequence and not only precludes its formation but also generates the secreted product sAPPalpha that possesses neurotrophic and neuroprotective properties. This beneficial cleavage has been shown to be strongly upregulated by protein kinase C (PKC) agonists and to be, at least partially, triggered by ADAM proteases (A Disintegrin And Metalloprotease). Recently, a proteolytic attack with similar characteristics has been shown to occur in the middle of the "toxic" 106-126 domain of the prion protein (PrPc), which PrPsc isoform is the causative agent of transmissible spongiform encephalopathies. As both Abeta and PrP(106-126) trigger neurotoxicity and cell death, this ADAM-dependent proteolytic attack could represent a valuable therapeutic target in order to deplete cells from these endogenous "toxins"and prevent the associated aggregates usually detected in affected brains.
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PMID:ADAM proteases: protective role in Alzheimer's and prion diseases? 1597 64

The cellular prion protein (PrP(c)) is physiologically cleaved in the middle of its 106-126 amino acid neurotoxic region at the 110/111 downward arrow112 peptidyl bond, yielding an N-terminal fragment referred to as N1. We recently demonstrated that two disintegrins, namely ADAM10 and ADAM17 (TACE, tumor necrosis factor alpha converting enzyme) participated in both constitutive and protein kinase C-regulated generation of N1, respectively. These proteolytic events were strikingly reminiscent of those involved in the so-called "alpha-secretase pathway" that leads to the production of secreted sAPPalpha from betaAPP. We show here, by transient and stable transfection analyses, that ADAM9 also participates in the constitutive secretion of N1 in HEK293 cells, TSM1 neurons, and mouse fibroblasts. Decreasing endogenous ADAM9 expression by an antisense approach drastically reduces both N1 and sAPPalpha recoveries. However, we establish that ADAM9 was unable to increase N1 and sAPPalpha productions after transient transfection in fibroblasts depleted of ADAM10. Accordingly, ADAM9 is unable to cleave a fluorimetric substrate of membrane-bound alpha-secretase activity in ADAM10(-/-) fibroblasts. However, we establish that co-expression of ADAM9 and ADAM10 in ADAM10-deficient fibroblasts leads to enhanced membrane-bound and released fluorimetric substrate hydrolyzing activity when compared with that observed after ADAM10 cDNA transfection alone in ADAM10(-/-) cells. Interestingly, we demonstrate that shedded ADAM10 displays the ability to cleave endogenous PrP(c) in fibroblasts. Altogether, these data provide evidence that ADAM9 is an important regulator of the physiological processing of PrP(c) and betaAPP but that this enzyme acts indirectly, likely by contributing to the shedding of ADAM10. ADAM9 could therefore represent, besides ADAM10, another potential therapeutic target to enhance the breakdown of the 106-126 and Abeta toxic domains of the prion and betaAPP proteins.
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PMID:The disintegrin ADAM9 indirectly contributes to the physiological processing of cellular prion by modulating ADAM10 activity. 1623 9

While a beta-sheet-rich form of the prion protein (PrPSc) causes neurodegeneration, the biological activity of its precursor, the cellular prion protein (PrPC), has been elusive. We have studied the effect of purified recombinant prion protein (recPrP) on rat fetal hippocampal neurons in culture. Overnight exposure to Syrian hamster or mouse recPrP, folded into an alpha-helical-rich conformation similar to that of PrPC, resulted in a 1.9-fold increase in neurons with a differentiated axon, a 13.5-fold increase in neurons with differentiated dendrites, a fivefold increase in axon length, and the formation of extensive neuronal circuitry. Formation of synaptic-like contacts was increased by a factor of 4.6 after exposure to recPrP for 7 days. Neither the N-terminal nor C-terminal domains of recPrP nor the PrP paralogue doppel (Dpl) enhanced the polarization of neurons. Inhibitors of protein kinase C (PKC) and of Src kinases, including p59Fyn, blocked the effect of recPrP on axon elongation, while inhibitors of phosphatidylinositol 3-kinase showed a partial inhibition, suggesting that signaling cascades involving these kinases are candidates for transduction of recPrP-mediated signals. The results predict that full-length PrPC functions as a growth factor involved in development of neuronal polarity.
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PMID:Recombinant prion protein induces rapid polarization and development of synapses in embryonic rat hippocampal neurons in vitro. 1631 16

Abnormalities of synapses and impaired synaptic transmission appear to be crucial in the pathogenesis of prion diseases. Excitotoxic mechanisms have been postulated as a major cause of neurodegeneration in these conditions. In this line, previous studies have shown abnormal group 1 metabotropic glutamate receptor signaling in Creutzfeldt-Jakob disease (CJD). In the present study, we have examined this pathway by western blotting in the cerebral cortex of bovine-spongiform encephalopathy (BSE)-infected bovine-PrP transgenic mice at different days post-inoculation (dpi). Activation of post-synaptic metabotropic glutamate receptor 1 (mGluR1) promotes phospholipase Cbeta1 (PLCbeta1) activation which may activate, in turn, protein kinase C (PKC), which regulates gene expression. Densitometric analysis of the western blot bands revealed no differences in the protein levels of (mGluR1) through time, but demonstrated decreased levels of PLCbeta1 and protein kinase C delta (nPKCdelta) at 270dpi, at the time when mice showed neurological deficits accompanied by neuropathological changes and PrPres deposition in the brain. The present results show, for the first time impairment of the mGluR1/PLCbeta1/PKCdelta pathway signaling with disease-progression in a murine model of BSE.
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PMID:Group I mGluR signaling in BSE-infected bovine-PrP transgenic mice. 1708 74

Changes in the composition of cell fractions, and in particular of detergent-resistant membranes (DRM) isolated from cultured rat cerebellar granule cells, were taken as possible changes in lipid raft composition during a signal transduction event. After activation of protein kinase C (PKC) with phorbol esters (PMA) or glutamate, the content of PKC and of proteins highly enriched (GAP43, Fyn, and PrP(c)) or not (MARCKS) in DRM was followed. PKC activation strongly increased its association with membranes (from 2% to 75%), causing its enrichment within DRM; the substrate GAP43, enriched in DRM, remained membrane associated, but its proportion in DRM dramatically decreased (from about 40% to 2.5%), suggesting its shift from raft to nonraft membranes, possibly as a consequence of phosphorylation by PKC. The distribution of Fyn and PrP(c) (DRM-enriched) and of MARCKS (present mainly outside DRM) did not change. PKC activation was followed by an increase of GAP43 and MARCKS phosphorylation (about 7- and 8-fold, respectively). Noteworthy was that, after cell treatment with the lipid raft-disrupting drug methyl-beta-cyclodextrin, PKC activation occurred normally, followed by MARCKS phosphorylation, but GAP43 phosphorylation did not occur. Taken altogether, these data suggest that the integrity of lipid rafts is necessary for PKC to affect GAP43 and catalyze its phosphorylation.
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PMID:Changes in the composition of detergent-resistant membrane domains of cultured neurons following protein kinase C activation. 1708 51


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