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)

Glutamate transporters (also called excitatory amino acid transporters, EAATs) bind extracellular glutamate and transport it to intracellular space to regulate glutamate neurotransmission and to maintain extracellular glutamate concentrations below neurotoxic levels. We previously showed that isoflurane, a commonly used anesthetic, enhanced the activity of EAAT3, a major neuronal EAAT. This effect required a protein kinase C (PKC) alpha-dependent EAAT3 redistribution to the plasma membrane. In this study, we prepared COS7 cells stably expressing EAAT3 with or without mutations of potential PKC phosphorylation sites in the putative intracellular domains. Here we report that mutation of threonine 5 or threonine 498 to alanine did not affect the isoflurane effects on EAAT3. However, the mutation of serine 465 to alanine abolished isoflurane-induced increase of EAAT3 activity and redistribution to the plasma membrane. The mutation of serine 465 to aspartic acid increased the expression of EAAT3 in the plasma membrane and also abolished the isoflurane effects on EAAT3. These results suggest an essential role of serine 465 in the isoflurane-increased EAAT3 activity and redistribution and a direct effect of PKC on EAAT3. Consistent with these results, isoflurane induced an increase in phosphorylation of wild type, T5A, and T498A EAAT3, and this increase was absent in S465A and S465D. Our current results, together with our previous data that showed the involvement of PKCalpha in the isoflurane effects on EAAT3, suggest that the phosphorylation of serine 465 in EAAT3 by PKCalpha mediates the increased EAAT3 activity and redistribution to plasma membrane after isoflurane exposure.
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PMID:Critical role of serine 465 in isoflurane-induced increase of cell-surface redistribution and activity of glutamate transporter type 3. 1706 70

ApoB mRNA editing involves site-specific deamination of cytidine 6666 producing an in-frame translation stop codon. Editing minimally requires APOBEC-1 and APOBEC-1 complementation factor (ACF). Metabolic stimulation of apoB mRNA editing in hepatocytes is associated with serine phosphorylation of ACF localized to editing competent, nuclear 27S editosomes. We demonstrate that activation of protein kinase C (PKC) stimulated editing and enhanced ACF phosphorylation in rat primary hepatocytes. Conversely, activation of protein kinase A (PKA) had no effect on editing. Recombinant PKC efficiently phosphorylated purified ACF64 protein in vitro, whereas PKA did not. Mutagenesis of predicted PKC phosphorylation sites S154 and S368 to alanine inhibited ethanol-stimulated induction of editing suggesting that these sites function in the metabolic regulation of editing. Consistent with this interpretation, substitution of S154 and S368 with aspartic acid stimulated editing to levels comparable to ethanol treatment in control McArdle RH7777 cells. These data suggest that phosphorylation of ACF by PKC may be a key regulatory mechanism of apoB mRNA editing in rat hepatocytes.
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PMID:Functional characterization of APOBEC-1 complementation factor phosphorylation sites. 1722 74

The phosphoprotein P of Borna disease virus (BDV) is an essential cofactor of the viral RNA-dependent RNA polymerase. It is preferentially phosphorylated at serine residues 26 and 28 by protein kinase C epsilon (PKCepsilon) and, to a lesser extent, at serine residues 70 and 86 by casein kinase II (CKII). To determine whether P phosphorylation is required for viral polymerase activity, we generated P mutants lacking either the PKCepsilon or the CKII phosphate acceptor sites by replacing the corresponding serine residues with alanine (A). Alternatively, these sites were replaced by aspartic acid (D) to mimic phosphorylation. Functional characterization of the various mutants in the BDV minireplicon assay revealed that D substitutions at the CKII sites inhibited the polymerase-supporting activity of P, while A substitutions maintained wild-type activity. Likewise, D substitutions at the PKC sites did not impair the cofactor function of BDV-P, whereas A substitutions at these sites led to increased activity. Interestingly, recombinant viruses could be rescued only when P mutants with modified PKCepsilon sites were used but not when both CKII sites were altered. PKCepsilon mutant viruses showed a reduced capacity to spread in cell culture, while viral RNA and protein expression levels in persistently infected cells were almost normal. Further mutational analyses revealed that substitutions at individual CKII sites were, with the exception of a substitution of A for S86, detrimental for viral rescue. These data demonstrate that, in contrast to other viral P proteins, the cofactor activity of BDV-P is negatively regulated by phosphorylation.
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PMID:Functional characterization of the major and minor phosphorylation sites of the P protein of Borna disease virus. 1737 20

RGS5 is a member of regulators of G protein signaling (RGS) proteins that attenuate heterotrimeric G protein signaling by functioning as GTPase-activating proteins (GAPs). We investigated phosphorylation of RGS5 and the resulting change of its function. In 293T cells, transiently expressed RGS5 was phosphorylated by endogenous protein kinases in the basal state. The phosphorylation was enhanced by phorbol 12-myristate 13-acetate (PMA) and endothelin-1 (ET-1), and suppressed by protein kinase C (PKC) inhibitors, H7, calphostin C and staurosporine. These results suggest involvement of PKC in phosphorylation of RGS5. In in vitro experiments, PKC phosphorylated recombinant RGS5 protein at serine residues. RGS5 protein phosphorylated by PKC showed much lower binding capacity for and GAP activity toward Galpha subunits than did the unphosphorylated RGS5. In cells expressing RGS5, the inhibitory effect of RGS5 on ET-1-induced Ca(2+) responses was enhanced by staurosporine. Mass spectrometric analysis of the phosphorylated RGS5 revealed that Ser166 was one of the predominant phosphorylation sites. Substitution of Ser166 by aspartic acid abolished the binding capacity to Galpha subunits and the GAP activity, and markedly reduced the inhibitory effect on ET-1-induced Ca(2+) responses. These results indicate that phosphorylation at Ser166 of RGS5 by PKC causes loss of the function of RGS5 in G protein signaling. Since this serine residue is conserved in RGS domains of many RGS proteins, the phosphorylation at Ser166 by PKC might act as a molecular switch and have functional significance.
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PMID:Phosphorylation of Ser166 in RGS5 by protein kinase C causes loss of RGS function. 1754 Apr 11

Cerebral ischemia can induce both the increase of excitation and the decrease of inhibition, which leads to neuronal excitotoxicity. Since glutamatergic and GABAergic transmissions work by each counterbalancing the function of the other, enhancing GABAergic activity should balance excessive glutamatergic excitation. But the potential mechanisms underlying these effects are obscure. Here, we used two GABA agonists, muscimol and baclofen, and performed immunoblotting, immunoprecipitation and histology analysis to evaluate the neuroprotective effects by stimulating GABA receptors in rat four-vessel occlusion (4-VO) ischemic model, and to investigate the potential mechanism. Our results indicate that whether in global cerebral ischemia in vivo, or in oxygen glucose deprivation (OGD) in vitro, coapplication of muscimol with baclofen can protect neurons from neuronal death through down-regulating the function of N-methyl-d-aspartic acid (NMDA) receptors via attenuating the tyrosine phosphorylation of NR2A subunit. We further elucidate that the phosphorylation level of Src kinase and the interaction among Src, post-synaptic density protein 95 and NR2A were also suppressed by coapplication of muscimol with baclofen. Both MK-801, a specific antagonist of NMDA receptors, and chelerythrine, an inhibitor of protein kinase C (PKC), could down-regulate the phosphorylation of NR2A via inhibiting the activation of Src and PKC respectively. These results suggest that the modified pattern of dynamic balance between excitation and inhibition by coactivation of the GABA receptors in cerebral ischemia can attenuate the excitatory NMDAR via inhibiting a novel postsynaptic NMDAR/Src-mediated signal amplification, the 'NMDAR-Ca(2+) --> PKC --> Src --> NMDAR-Ca(2+)' cycle.
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PMID:Activation of GABA receptors attenuates neuronal apoptosis through inhibiting the tyrosine phosphorylation of NR2A by Src after cerebral ischemia and reperfusion. 1802 28

The human polyomavirus BK (BKV) genome encodes the capsid proteins VP1 to VP3 and the three regulatory proteins, large and small tumor-antigen and the agnoprotein. Agnoprotein is a phospho-protein, but phosphorylation sites, protein kinases that mediate phosphorylation, and the biological importance of phosphorylation for the life-cycle of BK virus remain unknown. Here, we show that protein kinase C phosphorylates BKV agnoprotein at serine-11. Replacing serine-11 by either non-phosphorylable alanine or phospho-mimicking aspartic acid reduced the ability of these mutants to propagate compared to wildtype virus. Moreover, both these mutants displayed altered expression of viral proteins, which resulted from changed transrepressive property and stability of the mutated agnoprotein. Our results indicate that BKV propagation is controlled by phosphorylation of the agnoprotein and may suggest that specific inhibition of protein kinases may be used as a therapeutic strategy to hamper BK virus infection.
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PMID:Phosphorylation of human polyomavirus BK agnoprotein at Ser-11 is mediated by PKC and has an important regulative function. 1863 45

We previously reported that novel protein kinase C (nPKC) epsilon and N-methyl-d-aspartic acid (NMDA) receptors participated in morphine preconditioning (MP)-induced neuroprotection. In this study, we used Western blot analysis, 2,3,5-triphenyltetrazolium chloride (TTC) staining and lactate dehydrogenase (LDH) leakage assay to determine the involvement of conventional PKC isoforms (cPKC) in MP-induced neuroprotection against oxygen-glucose deprivation (OGD). Hippocampus slices (400-microm thickness) from healthy male BALB/c mice exposed to OGD for 5-45 min to mimic mild, moderate and severe ischemia in the presence of MP pretreatment. We found that OGD-induced damage in neuronal cell survival rate and LDH leakage could be improved by MP pretreatment (3 microM) within 20 min of OGD, which was abolished by concomitant incubation with non-selective opioid receptor antagonist naloxone (Nal, 50 microM). The results of Western blot analysis showed that only cPKCgamma membrane translocation, not alpha, betaI and betaII, increased under the condition of OGD 10 min and 2h reperfusion (OGD/2h), and this increment of cPKCgamma membrane translocation was inhibited by MP pretreatment. To further elucidate the role of cPKCgamma in MP-induced neuroprotection, we found that cPKCgamma membrane translocation inhibitor, Go6983 (6 nM) did not affect MP-induced neuroprotection while Go6983 alone exhibited a significant inhibition on OGD-induced increment in LDH leakage and decrease in cell survival rate. These phenomena were defined by the results that Go6983 could restore OGD-induced cPKCgamma membrane translocation, but had no further effect on MP-induced inhibition of cPKCgamma membrane translocation. These results demonstrated that MP can reduce OGD-induced neuronal injuries, and the down-regulation of cPKCgamma membrane translocation might be involved in the neuroprotection.
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PMID:Inhibition of PKCgamma membrane translocation mediated morphine preconditioning-induced neuroprotection against oxygen-glucose deprivation in the hippocampus slices of mice. 1870 78

Phosphorylation of the nicotinic acetylcholine receptor (nAChR) is believed to play a critical role in its nicotine-induced desensitization and up-regulation. We examined the contribution of a consensus PKC site in the alpha4 M3/M4 intracellular loop (alpha4S336) on the desensitization and up-regulation of alpha4beta2 nAChRs expressed in oocytes. Position alpha4S336 was replaced with either alanine to abolish potential phosphorylation at this site or with aspartic acid to mimic phosphorylation at this same site. Mutations alpha4S336A and alpha4S336D displayed a threefold increase in the ACh-induced response and an increase in ACh EC(50). Epibatidine binding revealed a three and sevenfold increase in surface expression for the alpha4S336A and alpha4S336D mutations, respectively, relative to wild-type, therefore, both mutations enhanced expression of the alpha4beta2 nAChR. Interestingly, the EC(50)'s and peak currents for nicotine activation remained unaffected in both mutants. Both mutations abolished the nicotine-induced up-regulation that is normally observed in the wild-type. The present data suggest that adding or removing a negative charge at this phosphorylation site cannot be explained by a simple straightforward on-and-off mechanism; rather a more complex mechanism(s) may govern the functional expression of the alpha4beta2 nAChR. Along the same line, our data support the idea that phosphorylation at multiple consensus sites in the alpha4 subunit could play a remarkable role on the regulation of the functional expression of the alpha4beta2 nAChR.
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PMID:Contribution of position alpha4S336 on functional expression and up-regulation of alpha4beta2 neuronal nicotinic receptors. 1881 99

Glutamate and norepinephrine (NE) are believed to mediate the long-lasting synaptic plasticity in the accessory olfactory bulb (AOB) that underlies pheromone recognition memory. The mechanisms by which these neurotransmitters bring about the synaptic changes are not clearly understood. In order to study signals that mediate synaptic plasticity in the AOB, we used AOB neurons in primary culture as a model system. Because induction of pheromone memory requires coincident glutamatergic and noradrenergic input to the AOB, and requires new protein synthesis, we reasoned that glutamate and NE must induce gene expression in the AOB. We used a combination of agonists that stimulate alpha1 and alpha2 adrenergic receptors in combination with N-methyl-d-aspartic acid and tested expression of the immediate-early gene (IEG) c-Fos. We found that the glutamatergic and noradrenergic stimulation caused significant induction of c-Fos mRNA and protein. Induction of c-Fos was significantly reduced in the presence of inhibitors of protein kinase C, mitogen-activated protein kinase (MAPK) and phospholipase C. These results suggest that glutamate and NE induce gene expression in the AOB through a signaling pathway mediated by protein kinase C and MAPK.
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PMID:Signal transduction and gene expression in cultured accessory olfactory bulb neurons. 1884 4

HuR is predominantly nuclear but following exposure to stress and mitogens, it can translocate to the cytoplasm where it stabilizes target mRNAs and/or modulates their translation. Several phosphorylation sites in a central 'hinge" region of HuR have been reported to affect its nucleocytoplasmic shuttle: phosphorylation by PKC at serine (S)221 and by Cdk1 at S202. Here, we investigated if there are additional putative phosphorylation sites within the HuR hinge region capable of influencing its cytoplasmic abundance. We systematically mutated all seven serine residues within the shuttling hinge domain to the nonphosphorylatable residue alanine (A), S197A, S202A, S221A, S229A, S232A, S241A and S242A. Using HeLa cells as the study system, we found that the HuR(S242A) mutant was more abundant in the cytoplasm in both untreated cells and in cells treated with short-wavelength ultraviolet light or with an inhibitor of Cdk1. Conversely, mutation of S242 to aspartic acid (D), rendered the phosphomimetic HuR(S242D) nuclear under all treatment conditions. S242 mutations did not influence HuR stability, but HuR(S242A) showed increased association with target cyclin A2 and cyclin B1 mRNAs. Accordingly, expression of HuR(S242A) led to increased cyclin mRNA stability and heightened cell proliferation rates. Our findings suggest that HuR phosphorylation at S242 hinders its cytoplasmic localization, its function as a posttranscriptional regulator, and its proliferative influence.
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PMID:Modification at HuR(S242) alters HuR localization and proliferative influence. 1894 43


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