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

1. The present study was carried out to identify the specific protein kinase C (PKC) isoform involved in regulatory volume decrease (RVD) responses, and to investigate the signal transduction pathways underlying the activation of volume-sensitive chloride channels in human cervical cancer HT-3 cells. The role of Ca2+ in RVD and in the activation of chloride currents was also studied. 2. The time course of RVDs was prolonged by microinjection of PKC-alpha antibody but not by PKC-beta or PKC-gamma antibody, and also by exposure to Ca2+-free medium, in particular when combined with microinjection of EDTA. Immunofluorescence staining showed that hypotonic superfusion evoked the translocation of PKC-alpha to the cell membrane, whereas PKC-beta or PKC-gamma remained unaffected. The translocation of PKC-alpha was observed a few minutes after hypotonic stress, reaching peak intensity at 30 min, and returned to the cytoplasm 60 min after hypotonic exposure. Western blot analyses showed an increased PKC-alpha level in terms of intensity and phosphorylation in the cell membrane, while neither PKC-beta nor PKC-gamma was activated upon hyposmotic challenge. 3. Whole-cell patch-clamp studies demonstrated that neomycin and PKC blockers such as staurosporine and H7 inhibited volume-sensitive chloride currents. The inhibitory effect of neomycin on chloride currents can be reversed by the PKC activator phorbol 12-myristate, 13-acetate (PMA). Moreover, the PKC inhibitor and PKC-alpha antibody, but not PKC-beta or PKC-gamma antibody, significantly attenuated the chloride currents. The activation of volume-sensitive chloride currents were insensitive to the changes of intracellular Ca2+ but required the presence of extracellular Ca2+. 4. Our results suggest the involvement of PKC-alpha and extracellular Ca2+ in RVD responses and the activation of volume-sensitive chloride channels in HT-3 cells.
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PMID:Involvement of PKC-alpha in regulatory volume decrease responses and activation of volume-sensitive chloride channels in human cervical cancer HT-3 cells. 976 33

The absence of learning-related redistribution of hippocampal protein kinase C (PKC) has been correlated with impairment of learning performance induced by developmental lead (Pb) exposure. This study was designed to examine whether the properties of brain PKC are altered by chronic Pb exposure during development. Two-tenth percent Pb acetate was administered to pregnant and lactating dams and then administered to weanlings in drinking water until postnatal day (PN) 56. Effects of Pb on translocation of PKC were studied in brain slices prepared from hippocampus. When the slices were treated with 0.33 microM phorbol-12, 13-dibutyrate (PDBu) for 15 min, a significant increase in PKC activity was observed in the membrane fraction of hippocampal slices from Pb-exposed rats, suggesting that chronic Pb exposure potentiates PDBu-activated PKC translocation. Data obtained from saturation binding assays in the frontal cortices of Pb-exposed rats showed a decrease in the dissociation constant (KD) in both membrane and cytosolic PKC. A decrease in the total binding sites (Bmax) of [3H]PDBu binding was only observed in membrane PKC. Furthermore, developmental Pb exposure decreased PKC-gamma, but not PKC-alpha, -betaII, and -epsilon in the membrane fraction of the hippocampus and the frontal cortex. These results indicate that chronic Pb exposure during development increases phorbol ester binding affinity, enhances phorbol ester-induced translocation of PKC, and down-regulates membrane PKC, mainly PKC-gamma.
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PMID:Protein kinase C in rat brain is altered by developmental lead exposure. 1021 16

The mechanisms of neuronal degeneration following hypoxia/ischemia remain undefined, but the processes include increases in neurotransmitter release, elevation of cytosolic-free calcium concentration, and changes in signal transduction pathways. Activation of the multigene family of protein kinase C (PKC) has been associated with the release of neurotransmitter and the survival of neurons. Therefore, to understand which PKC isozymes are involved in hypoxia/ischemia-induced neuronal degeneration, we examined PKC isozymes after chemical hypoxia (i.e., KCN exposure) in PC12 cells. Cell toxicity, as measured by lactate dehydrogenase (LDH) release, was increased significantly by KCN in glucose-free DMEM and was exaggerated by acute 12-O-tetradecanoyl phorbol-13-acetate (TPA) pretreatment. Under parallel conditions, KCN elevated cytosolic-free calcium ([Ca2+]i) in glucose-free but not in glucose containing DMEM, and TPA pretreatment did not exaggerate KCN's effect on [Ca2+]i. Thus, increases in [Ca2+]i are not sufficient for the synergistic toxic effect of KCN and TPA. In the glucose-free DMEM, selective PKC isozyme inhibitor Go 6976 at 10 nM completely inhibited KCN-induced LDH release and at higher concentrations (1 microM) inhibited the basal levels of LDH release. The protein levels of PKCs in the nuclear, membrane, and cytosolic fractions were measured by Western blot analysis using antibodies against specific isoforms. Two Ca2+-dependent (-alpha, -gamma) and four Ca2+-independent (-delta, -epsilon, -zeta, and -lambda) isozymes were identified and two isozymes (-beta and -theta) were not detected in the subcellular fractions of PC12 cells. Treatment of the cells with TPA significantly activated translocation of conventional PKC-gamma from the cytosol to the membrane and nuclear fractions and other PKC isozymes (-alpha, -delta, and -epsilon) from the cytosol to the membrane, but not atypical PKC-zeta and -lambda. Although only the levels in the nuclear PKC-gamma but not other PKC isozymes were increased significantly following KCN, the levels of cPKC-alpha and -gamma in the membrane mainly- and those and PKC-epsilon in the nucleus-were increased when KCN was combined with TPA. In addition, this condition (TPA + KCN) did not affect the TPA insensitive atypical isozymes, PKC-zeta or -lambda. Taking the results together, differential activation/translocation of PKC isozymes by KCN and TPA is important in the regulation of chemical hypoxia-induced cell injury in PC12 cells.
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PMID:Selective subcellular redistributions of protein kinase C isoforms by chemical hypoxia. 1037 22

The newly discovered gamma-PKC-related-protein of human leukocytes (gamma-rp) crossreacts with a polyclonal antibody preparation originally designed to be specific for PKC-gamma (gammaMb-Ab). As this antibody is currently the only suitable probe for gamma-rp, we sought to characterize the binding of the two proteins. We determined that the gamma Mg-Ab does not recognize the native form of gamma-rp. However, with denaturing immunoblots of gamma-rp, we found that 1) the crossreactive gamma-rp epitope differs somewhat from that of classic rat brain PKC-gamma, but probably only to the degree of the rat/human PKC species difference; 2) the previously reported doublet bands of gamma-rp represent a single protein with cell-stimulus inducible modifications; 3) antibodies present in the gammaMg-Ab pool bind to two separate sites within the gamma-rp epitope; 4) access to one binding site is conformationally restricted, even after protein denaturation; 5) agonist-induced modification of gamma-rp does not significantly affect the total amount of gamma Mg-Ab that it can bind, but 6) does significantly affect the rate of antibody binding to one site. This investigation defines the appropriate experimental use of our antibody, and the significance of these findings for the future study and cloning of gamma-rp is discussed.
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PMID:Analysis of the PKC-gamma-related immunocrossreactive region of a novel leukocyte protein gamma-rp. 1044

Protein kinase C has been implicated in long-term regulation of cellular functions including induction and maintenance of hippocampal long-term potentiation. In the present study the time-course of long-term potentiation-induced translocation of Ca(2+)-dependent protein kinase C isoenzymes (PKCalpha/beta and PKCgamma) was investigated. Quantitative immunoblot analysis was used to measure translocation of these isoenzymes between cytosolic, membrane-associated and membrane-inserted fraction at 5, 15 and 60 min after induction of long-term potentiation in the dentate gyrus in vivo. To investigate the involvement of metabotropic glutamate receptors in protein kinase C regulation during long-term potentiation induction, additional animals were treated before tetanization with (R,S)-alpha-methyl-4-carboxyphenylglycine, an antagonist of metabotropic glutamate receptors. Brief tetanic stimulation of the perforant path resulted in a 100-150% increase in the population spike amplitude in response to test stimuli 5, 15 or 60 min after stimulation in both untreated and (R,S)-alpha-methyl-4-carboxyphenylglycine-treated animals. Only those rats showing clear potentiation were selected for further biochemical analysis of the potentiated dentate gyrus. Five minutes after high-frequency stimulation the subcellular distribution of all studied protein kinase C isoenzymes was unchanged compared with controls. PKC-gamma translocated into the cytosol 15 min after tetanization and this redistribution was blocked by (R,S)-alpha-methyl-4-carboxyphenylgly-cine pretreatment. By contrast, PKC alpha/beta levels increased in the cytosolic fraction only 60 min after tetanization, but in a (R,S)-alpha-methyl-4-carboxyphenylglycine-independent manner. In an additional set of experiments it was shown that (R,S)-alpha-methyl-4-carboxyphenylglycine alone applied intraventricularly had no effect on the subcellular distribution of the studied isoenzymes. The data suggest that PKCalpha/beta and PKCgamma are activated during different post-tetanic phases and metabotropic glutamate receptor activation might be essential for tetanus-induced translocation of postsynaptic PKCgamma only.
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PMID:Transient translocation of protein kinase Cgamma in hippocampal long-term potentiation depends on activation of metabotropic glutamate receptors. 1050 52

The senescence-accelerated P8 mouse (SAMP8) is a well-characterized model for the age-related decline in acquisition and retention. Calcium-dependent protein kinase C (PKC) and calcium-calmodulin-dependent protein kinase (CAM K) have been implicated in these processes in the hippocampus. Therefore, the expression of hippocampal PKC and CAM K was determined in SAMP8 mice aged 4, 8, and 12 months. As measured by Western blotting, total hippocampal PKC-gamma protein declined linearly with age. In addition, the distribution of the PKC-gamma also changed with age. The amount of PKC in the particulate fraction declined linearly with age relative to the soluble PKC. The decline in total PKC and particulate PKC correlated with the previously reported decline in retention but not with the decline in acquisition. Western blotting revealed no consistent change in CAM KII protein levels. In addition to protein levels, Ca-dependent protein kinase activity may also be affected by changes in intracellular Ca concentration. Therefore, the levels of calbindin and the plasma membrane Ca pump, two proteins involved in maintaining low levels of intracellular Ca, were measured in the hippocampus. Calbindin protein declined progressively with age, but there was no significant change in total plasma membrane Ca pump expression. These studies demonstrate a decrease in the amount and distribution of hippocampal PKC-gamma in the SAMP8 between 4 and 12 months that is associated with decreased retention.
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PMID:Effect of age on calcium-dependent proteins in hippocampus of senescence-accelerated mice. 1052 25

Family of protein kinase C (PKC) isozymes play a key role in transducing a vast number of signals into the cells. The members of classical PKC family are activated by binding of various lipid ligands to one of the several cysteine-rich domains of the enzyme. Second cysteine-rich (Cys2) domain of PKC-gamma was expressed in Escherichia coli as a fusion protein with glutathione-S-transferase (GST) using the cDNA sequence from rat brain. The Cys2 protein after cleavage from GST was purified to homogeneity using glutathione-agarose and Mono-S cation exchanger column. In order to investigate the interaction of lipids and calcium with Cys2 protein we used UW spectroscopy. The UV spectrum of Cys2 protein exhibited a maximum at 205 nm. Exposition of Cys2 protein to phosphatidylserine (PS) vesicles resulted in significant decrease in the absorbance in the 210 nm region. Changes in UW spectrum of Cys2 protein induced by phorbol 12,13-dibutyrate (PDB) were smaller than those induced by PS, and addition of PDB with PS had no effect on the PS induced changes in UV spectrum of Cys2. Neither phosphatidylcholine (PC) nor phosphatidylethanolamine (PE) affected UV spectrum of Cys2 but in the presence of phosphatidylinositol 4,5 bisphosphate (PIP2) or phosphatidyliinositol 4-phosphate (PIP) vesicles some changes were observed. Calcium ions alone or in the presence of PS had no effect on the UV spectrum of Cys2 protein. These data indicate that PS comparing to PDB, interacts with a larger area of Cys2 protein, and that the binding sites for these two molecules are at least overlapping. The site of PIP and PIP2 interaction with PKC-gamma is distinct from that of phorbol ester binding site.
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PMID:The ultraviolet studies on protein-lipid interaction of a protein kinase C-gamma phorbol-binding domain. 1054 41

Studies on the mode of action of basic fibroblast growth factor (bFGF) identified an essential role of heparan sulfate and heparin-like molecules in the formation of distinct bFGF-heparan sulfate-bFGF-receptor complexes that are required for bFGF-induced signal transduction. In coronary smooth muscle cells that express 6-8 ng bFGF mg(-1) cell protein, the heparan sulfate chains of membrane-associated proteoheparan sulfate are implicated in bFGF signaling and thus are involved in the regulation of proliferation and differentiation of vascular smooth muscle cells. We studied the mode of action of a synthetic non-sulfated heparin-mimicking compound termed RG-13,577 (poly-4-hydroxyphenoxy acetic acid, Mr approximately 5 kD) and found a dose-dependent antiproliferative effect that was characterized by a block of G(1)/S-phase transition indicated by a marked (80%) reduction of [3H]thymidine incorporation at a concentration of 5 microg ml(-1) RG-13,577. Cell cycle analysis showed a block of cell division in the G(1)-phase. In response to RG-13,577 the cells were converted into a hypertrophic growth status within 72 h as judged from a doubling of the cellular protein content and measurement of cell and nucleus size. The increased cell protein content resulted from a de novo synthesis and was also associated with an increase in the incorporation of [35S]sulfate into cell-associated proteoglycans, including the proteoheparan sulfate coreceptor of bFGF. In contrast, the compound-induced G(1)-phase arrest was associated with an extensive downregulation of the cellular and pericellular bFGF level. The reduced bFGF content was accompanied by downregulation of the bFGF signaling-involved protein kinase C-alpha and MAP kinase, abrogation of MAP kinase phosphorylation and overexpression of protein kinase C-gamma. RG-13,577 failed to elicit apoptotic reactions at a concentration range of 0.5-10 microg ml(-1) and its effect was reversible upon removal of the compound. It appears that RG-13,577 induces a phenotype transformation of coronary SMC into a metabolically active hypertrophic status that could promote repair processes after balloon angioplasty (PTCA) without stimulating cell proliferation. Development of non-toxic polyanionic compounds may provide an effective strategy to inhibit cell proliferation associated with restenosis following balloon angioplasty and coronary artery bypass surgery.
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PMID:Differentiation of coronary smooth muscle cells to a cell cycle-arrested hypertrophic growth status by a synthetic non-toxic heparin-mimicking compound. 1055 25

Prenatal ethanol exposure can cause a number of physiological deficits known as fetal alcohol syndrome (FAS). Because protein kinase C (PKC) regulates the cell cycle and has been linked to growth, we examined the effect of ethanol on PKC isoform expression in a developing chick brain. Ethanol exposure causes decreased head weight in chickens at day 5 in a dose-dependent manner and a decreased brain weight at days 7 and 10 at an ethanol concentration of 1.0 g/kg. Antibodies specific for PKC-alpha, beta, gamma, delta, epsilon, iota, lambda, mu and zeta were used to examine ethanol's effect on PKC expression in the growth-suppressed brain at days 5, 7 and 10 of development. Only four of the PKC isoforms tested are expressed in the chick brain prior to day 10: alpha, gamma, epsilon, and iota. PKC-alpha, gamma, and epsilon are developmentally increased during the time period studied. Ethanol causes a decreased expression of PKC-alpha on days 5, 7 and 10 and a decreased expression of PKC-gamma on days 7 and 10. Ethanol causes a decreased expression of PKC-epsilon only on day 7. PKC-iota expression is unchanged over the developmental times studied and ethanol exposure has no effect on PKC-iota expression. These data suggest that only specific PKC isoforms are developmentally expressed in the embryonic chick brain and that ethanol may inhibit the expression of those PKC isoforms that are developmentally regulated.
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PMID:Ethanol-induced decrease of developmental PKC isoform expression in the embryonic chick brain. 1056 37

Systemic hyperglycemia and hypercapnia severely aggravate ischemic brain damage when instituted prior to cerebral ischemia. An aberrant cell signaling following ischemia has been proposed to be involved in ischemic cell death, affecting protein kinase C (PKC) and the calcium calmodulin kinase II (CaMKII). Using a cardiac arrest model of global brain ischemia of 10 min duration, we investigated the effect of hyperglycemia (20 mM) and hypercapnia (pCO(2) 300 mmHg) on the subcellular redistribution of PKC (alpha, beta, gamma) and CaMKII to synaptic membranes and to the microsomes, as well as the effect on PKC activity. We confirmed the marked translocation of PKC and CaMKII to cell membranes induced by ischemia, concomitantly with a decrease in the PKC activity in both the membrane fraction and cytosol. Hyperglycemia and hypercapnia markedly enhanced the translocation of PKC-gamma to cell membranes while other PKC isoforms were less affected. There was no effect of acidosis on PKC activity, or on translocation of CaMKII to cell membranes. Our data strongly suggest that the enhanced translocation of PKC to cell membranes induced by hyperglycemia and hypercapnia may contribute to the detrimental effect of tissue acidosis on the outcome following ischemia.
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PMID:Acidosis enhances translocation of protein kinase C but not Ca(2+)/calmodulin-dependent protein kinase II to cell membranes during complete cerebral ischemia. 1059 93


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