scholarly journals SN1 Transporter Regulation by Sp1 in Ammonia-Treated Mouse Cortical Astrocytes: Role of Sp1 Phosphorylation Status

2018 ◽  
Author(s):  
Katarzyna Dąbrowska ◽  
Magdalena Zielińska
Keyword(s):  
Cellular Localization ◽  
Regulatory Role ◽  
Sp1 Transcription Factor ◽  
Kinase C ◽  
Cultured Astrocytes ◽  
Pkc Isoforms ◽  
Glutamine Transport ◽  
Specificity Protein

The involvement of astrocytic SN1 (SNAT3) transporter in ammonia-induced L-glutamine retention was recently documented in mouse cultured astrocytes. Here we investigated the involvement of specificity protein 1 (Sp1) transcription factor in SN1 regulation in ammonium chloride (“ammonia”)-treated astrocytes. Sp1 expression and its cellular localization were determined using real-time qPCR, Western blot and confocal microscopy, respectively. Sp1 binding to Snat3 promoter was analyzed by chromatin immunoprecipitation. Ammonia-induced Sp1 regulatory role in SN1-mediated [3H]glutamine transport was verified using siRNA and mithramycin A. The involvement of protein kinase C (PKC) isoforms in Sp1 level/phosphorylation status was verified using siRNA technology. Sp1 translocation to the nuclei and its enhanced binding to Snat3 promoter, along with Sp1 dependence of system N-mediated [3H]glutamine transport were observed in astrocytes upon ammonia exposure. Ammonia decreased the level of phosphorylated Sp1, and the effect was reinforced by long-term incubation with PKC modulator, phorbol 12-myristate 13-acetate, a treatment likely to dephosphorylate Sp1. Furthermore,  silencing of PKCδ isoform abolished the increase of Sp1 level by ammonia. Collectively, the results demonstrate the regulatory role of Sp1 in regulation of SN1 expression and activity in ammonia-treated astrocytes and implicate altered Sp1 phosphorylation status in this capacity.

scholarly journals Silencing of Transcription Factor Sp1 Promotes SN1 Transporter Regulation by Ammonia in Mouse Cortical Astrocytes

2019 ◽  
Vol 20 (2) ◽  
pp. 234 ◽  
Author(s):  
Katarzyna Dąbrowska ◽  
Magdalena Zielińska
Keyword(s):  
Transcription Factor ◽  
Cellular Localization ◽  
Sp1 Transcription Factor ◽  
Kinase C ◽  
Cultured Astrocytes ◽  
Pkc Isoforms ◽  
Specificity Protein ◽  
Glutamine Uptake

The involvement of the astrocytic SN1 (SNAT3) transporter in ammonia-induced l-glutamine retention was recently documented in mouse-cultured astrocytes. Here we investigated the involvement of specificity protein 1 (Sp1) transcription factor in SN1 regulation in ammonium chloride (“ammonia”)-treated astrocytes. Sp1 expression and its cellular localization were determined using real-time qPCR, Western blot, and confocal microscopy. Sp1 binding to Snat3 promoter was analyzed by chromatin immunoprecipitation. The role of Sp1 in SN1 expression and SN1-mediated [3H]glutamine uptake in ammonia-treated astrocytes was verified using siRNA and mithramycin A. The involvement of protein kinase C (PKC) isoforms in Sp1 level/phosphorylation status was verified using siRNA technology. Sp1 translocation to the nuclei and its enhanced binding to the Snat3 promoter, along with Sp1 dependence of system N-mediated [3H]glutamine uptake, were observed in astrocytes upon ammonia exposure. Ammonia decreased the level of phosphorylated Sp1, and the effect was reinforced by long-term incubation with PKC modulator, phorbol 12-myristate 13-acetate, which is a treatment likely to dephosphorylate Sp1. Furthermore, silencing of the PKCδ isoform appears to enhance the ammonia effect on the Sp1 level. Collectively, the results demonstrate the regulatory role of Sp1 in regulation of SN1 expression and activity in ammonia-treated astrocytes and implicate altered Sp1 phosphorylation status in this capacity.

The role of PKMζ in the maintenance of long-term memory: a review

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Hamish Patel ◽  
Reza Zamani
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Long Term Potentiation ◽  
Global Memory ◽  
Long Term Memory ◽  
Compensatory Mechanism ◽  
Term Memory ◽  
Kinase C

Abstract Long-term memories are thought to be stored in neurones and synapses that undergo physical changes, such as long-term potentiation (LTP), and these changes can be maintained for long periods of time. A candidate enzyme for the maintenance of LTP is protein kinase M zeta (PKMζ), a constitutively active protein kinase C isoform that is elevated during LTP and long-term memory maintenance. This paper reviews the evidence and controversies surrounding the role of PKMζ in the maintenance of long-term memory. PKMζ maintains synaptic potentiation by preventing AMPA receptor endocytosis and promoting stabilisation of dendritic spine growth. Inhibition of PKMζ, with zeta-inhibitory peptide (ZIP), can reverse LTP and impair established long-term memories. However, a deficit of memory retrieval cannot be ruled out. Furthermore, ZIP, and in high enough doses the control peptide scrambled ZIP, was recently shown to be neurotoxic, which may explain some of the effects of ZIP on memory impairment. PKMζ knockout mice show normal learning and memory. However, this is likely due to compensation by protein-kinase C iota/lambda (PKCι/λ), which is normally responsible for induction of LTP. It is not clear how, or if, this compensatory mechanism is activated under normal conditions. Future research should utilise inducible PKMζ knockdown in adult rodents to investigate whether PKMζ maintains memory in specific parts of the brain, or if it represents a global memory maintenance molecule. These insights may inform future therapeutic targets for disorders of memory loss.

Role of PKC isoforms in glucose transport in 3T3-L1 adipocytes: insignificance of atypical PKC

2002 ◽  
Vol 283 (2) ◽  
pp. E338-E345 ◽  
Author(s):  
Masatoshi Tsuru ◽  
Hideki Katagiri ◽  
Tomoichiro Asano ◽  
Tetsuya Yamada ◽  
Shigeo Ohno ◽  
...  
Keyword(s):  
Glucose Transport ◽  
Dominant Negative ◽  
Transport Activity ◽  
Wild Type ◽  
Atypical Pkc ◽  
Endogenous Expression ◽  
Kinase C ◽  
Pkc Isoforms ◽  
Pkc Ζ

To elucidate the involvement of protein kinase C (PKC) isoforms in insulin-induced and phorbol ester-induced glucose transport, we expressed several PKC isoforms, conventional PKC-α, novel PKC-δ, and atypical PKC isoforms of PKC-λ and PKC-ζ, and their mutants in 3T3-L1 adipocytes using an adenovirus-mediated gene transduction system. Endogenous expression and the activities of PKC-α and PKC-λ/ζ, but not of PKC-δ, were detected in 3T3-L1 adipocytes. Overexpression of each wild-type PKC isoform induced a large amount of PKC activity in 3T3-L1 adipocytes. Phorbol 12-myristrate 13-acetate (PMA) activated PKC-α and exogenous PKC-δ but not atypical PKC-λ/ζ. Insulin also activated the overexpressed PKC-δ but not PKC-α. Expression of the wild-type PKC-α or PKC-δ resulted in significant increases in glucose transport activity in the basal and PMA-stimulated states. Dominant-negative PKC-α expression, which inhibited the PMA activation of PKC-α, decreased in PMA-stimulated glucose transport. Glucose transport activity in the insulin-stimulated state was increased by the expression of PKC-δ but not of PKC-α. These findings demonstrate that both conventional and novel PKC isoforms are involved in PMA-stimulated glucose transport and that other novel PKC isoforms could participate in PMA-stimulated and insulin-stimulated glucose transport. Atypical PKC-λ/ζ was not significantly activated by insulin, and expression of the wild-type, constitutively active, and dominant-negative mutants of atypical PKC did not affect either basal or insulin-stimulated glucose transport. Thus atypical PKC enzymes do not play a major role in insulin-stimulated glucose transport in 3T3-L1 adipocytes.

The role of protein kinase C isozymes in TNF-α-induced cytotoxicity to a rat intestinal epithelial cell line

2001 ◽  
Vol 280 (4) ◽  
pp. G572-G583 ◽  
Author(s):  
Q. Chang ◽  
B. L. Tepperman
Keyword(s):  
Epithelial Cell ◽  
Caspase 3 ◽  
Cell Injury ◽  
Cell Damage ◽  
Epithelial Cell Line ◽  
Intestinal Epithelial ◽  
Kinase C ◽  
Pkc Isoforms ◽  
Tnf Α

Tumor necrosis factor (TNF)-α can induce cytotoxicity and apoptosis in a number of cell types and has been implicated in the regulation of many inflammatory processes. It has been suggested that protein kinase C (PKC) is one of the intracellular mediators of the actions of TNF-α. In the present study, the role of PKC isoforms in TNF-α-mediated cytotoxicity and apoptosis in intestinal cells was investigated using the rat epithelial cell line, IEC-18. Cells were incubated with TNF-α in the presence or absence of the transcription inhibitor actinomycin D (AMD). The extent of cell damage was enhanced when AMD was added to incubation medium, suggesting that new protein synthesis plays a role in the cytotoxic action of TNF. TNF-α also induced the translocation of PKC-α, -δ, and -ε from cytosol to the membrane fraction of the intestinal cells. Furthermore, the cytotoxic and apoptotic effects of TNF were reduced by pretreating the cells with the PKC-ε translocation inhibitor, PKC-εV1–2. In contrast, although cells incubated with the phorbol ester phorbol 12-myristate 13-acetate (PMA) also displayed an increase in cell injury, the extent of cytotoxicity and apoptosis was not enhanced by AMD. Furthermore, PMA-induced cell damage was reduced by rottlerin, a PKC-δ inhibitor. Caspase-3, an enzyme implicated in the mediation of apoptosis, was activated in cells in response to either TNF-α or PMA stimulation, and its effects on this activity were reduced by selective inhibition of PKC-ε and -δ, respectively. Furthermore, inhibition of caspase-3 activity reduced apoptosis. These data suggest that activation of selective PKC isoforms mediate the effects of TNF-α on intestinal epithelial cell injury.

Protein Kinase C Isoform ε Negatively Regulates ADP-Induced Thromboxane Generation by Regulating cPLA2 Activation and Calcium Mobilization In Platelets

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2020-2020
Author(s):  
Yamini Bynagari ◽  
Parth Lakhani ◽  
Kamala Bhavaraju ◽  
Jianguo Jin ◽  
Mario C Rico ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Conflicts Of Interest ◽  
Arterial Injury ◽  
Calcium Mobilization ◽  
Erk Activation ◽  
Kinase C ◽  
Cpla2 Activation ◽  
Pkc Isoforms

Abstract Abstract 2020 Positive regulatory role of Protein Kinase C (PKC) isoforms in platelets have been extensively studied. However, negative regulatory roles of PKCs in platelets are poorly understood. In this study we investigated the mechanism by which PKCs negatively regulate ADP-induced thromboxane generation and identified PKC isoforms involved in this process. Pan PKC inhibition with GF109203X potentiated ADP-induced cPLA2 phosphorylation suggesting that PKCs negatively regulate thromboxane generation by regulating cPLA2 activation. Inhibition of PKCs potentiated ADP-induced ERK activation and intracellular calcium mobilization, two upstream signaling molecules of cPLA2.These data suggest that PKCs negatively regulate thromboxane by regulating ERK activation and calcium mobilization, which inturn regulate cPLA2 activation. Pan-PKC inhibition potentiated ADP-induced, P2Y1 receptor-mediated calcium mobilization in platelets independent of P2Y12-receptor. Pretreatment of platelets with GF109203X, a Pan PKC inhibitor, but not Go-6976, a classical PKC isoform inhibitor, potentiated ADP-induced thromboxane generation. Thus, we investigated the role of various novel class of PKC isoforms (nPKCs) in platelets. We have previously demonstrated that nPKC η, θ, δ positively regulates agonist-induced thromboxane generation in platelets. Thus, we investigated if the role of nPKC ε in ADP-induced thromboxane generation using PKC ε knockout mice (PKCε KO). ADP-induced thromboxane generation in PKC ε KO murine platelets was ten-fold higher than that of wild type platelets. Furthermore, PKC ε KO mice exhibited shorter times to occlusion in FeCl3-induced arterial injury model and shorter bleeding times in Tail bleeding experiments. We conclude that PKCε negatively regulates ADP-induced thromboxane generation in platelets and thereby offers protection against thrombosis. Disclosures: No relevant conflicts of interest to declare.

The interrelation of the angiotensin and endothelin systems on the modulation of NAD(P)H oxidaseThis paper is one of a selection of papers published in this Special issue, entitled Young Investigator's Forum.

2006 ◽  
Vol 84 (1) ◽  
pp. 21-28 ◽  
Author(s):  
Marc-André Laplante ◽  
Jacques de Champlain
Keyword(s):  
Superoxide Anion ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Cellular Membrane ◽  
Kinase C ◽  
Therapeutic Tools ◽  
Produce Superoxide Anion ◽  
Selection Of

The NAD(P)H oxidase is an enzyme assembled at the cellular membrane able to produce superoxide anion from NADH or NAD(P)H (nicotinamide adenine dinucleotide phosphate). It is one of the main sources of superoxide anion in cardiovascular tissues and its role in a variety of cardiovascular disorders such as atherosclerosis, cardiac hypertrophy, and endothelial dysfunction was recently proposed. Although, many factors and receptors were shown to lead to the activation of the enzyme, particulary the type 1 angiotensin receptor, the pathways involved are still widely unknown. Despite the identification of factors such as c-Src and protein kinase C implicated in the acute activation of NAD(P)H oxidase, the signalling involved in the sustained activation of the enzyme is probably far more complex than was previously envisioned. In this review, we describe the role of endothelin-1 in NAD(P)H oxidase signalling after a sustained stimulation by angiotensin II. Since most pathologies caused by an NAD(P)H oxidase overactivation develop over a relatively long period of time, it is necessary to better understand the long-term signalling of the enzyme for the development or use of more specific therapeutic tools.

1,25-Dihydroxyvitamin D3 and TPA activate phospholipase D in Caco-2 cells: role of PKC-α

1999 ◽  
Vol 276 (4) ◽  
pp. G993-G1004 ◽  
Author(s):  
Sharad Khare ◽  
Marc Bissonnette ◽  
Beth Scaglione-Sewell ◽  
Ramesh K. Wali ◽  
Michael D. Sitrin ◽  
...  
Keyword(s):  
Phospholipase D ◽  
Dihydroxyvitamin D3 ◽  
Dihydroxyvitamin D ◽  
1,25 Dihydroxyvitamin D ◽  
Kinase C ◽  
Pkc Isoforms ◽  
Gf 109203X ◽  
Pkc Β ◽  
Stimulation Of

1,25-Dihydroxyvitamin D3[1,25(OH)2D3] and 12- O-tetradecanoylphorbol 13-acetate (TPA) both activated phospholipase D (PLD) in Caco-2 cells. GF-109203x, an inhibitor of protein kinase C (PKC) isoforms, inhibited this activation by both of these agonists. 1,25(OH)2D3activated PKC-α, but not PKC-β1, -βII, -δ, or -ζ, whereas TPA activated PKC-α, -β1, and -δ. Chronic treatment with TPA (1 μM, 24 h) significantly reduced the expression of PKC-α, -βI, and -δ and markedly reduced the ability of 1,25(OH)2D3or TPA to acutely stimulate PLD. Removal of Ca2+ from the medium, as well as preincubation of cells with Gö-6976, an inhibitor of Ca2+-dependent PKC isoforms, significantly reduced the stimulation of PLD by 1,25(OH)2D3or TPA. Treatment with 12-deoxyphorbol-13-phenylacetate-20-acetate, which specifically activates PKC-βI and -βII, however, failed to stimulate PLD. In addition, the activation of PLD by 1,25(OH)2D3or TPA was markedly reduced or accentuated in stably transfected cells with inhibited or amplified PKC-α expression, respectively. Taken together, these observations indicate that PKC-α is intimately involved in the stimulation of PLD in Caco-2 cells by 1,25(OH)2D3or TPA.

Abstract 932: Temporal Changes In The Contribution Of Protein Kinase C Isoforms To Myogenic Constriction In Rat Posterior Cerebral Arteries

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Toshihide Kashihara ◽  
Koichi Nakayama ◽  
Tomohisa Ishikawa
Keyword(s):  
Initial Phase ◽  
Cerebral Arteries ◽  
Ruthenium Red ◽  
Intraluminal Pressure ◽  
Myogenic Tone ◽  
Kinase C ◽  
Pkc Isoforms ◽  
Pkc Inhibitors ◽  
Protein Kinase C Isoforms

Background Cerebral arteries respond to an increase in intraluminal pressure with vasoconstriction, being referred to as myogenic tone. A number of studies have postulated the mechanisms involved in the development of myogenic tone; however, less information is available about how myogenic tone is maintained. The present study thus investigated the mechanisms underlying sustained myogenic constriction in isolated rat posterior cerebral arteries. Methods and Results Long-term elevation of intraluminal pressure from 5 to 60 mmHg for 1 hr caused sustained constriction and [Ca 2+ ] i elevation. In the presence of rottlerin, a PKCδ inhibitor, the pressure-induced constriction and [Ca 2+ ] i elevation were gradually declined, and nearly abolished at the end of the 1-hr stimulation (sustained phase). In contrast, Gö 6976, a cPKC inhibitor, significantly inhibited the constriction for up to 5 min after the start of stimulation (initial phase), but had no effects on the [Ca 2+ ] i elevation. The pressure stimulation induced small [Ca 2+ ] i elevation even in the presence of nicaripine. Ruthenium red (RuR), a TRPV inhibitor, significantly inhibited the nicardipine-resistant [Ca 2+ ] i elevation in the initial phase. However, slowly developing [Ca 2+ ] i elevation was still observed in the sustained phase. In contrast, rottlerin had little effect on the initial phase, but significantly inhibited the sustained one. Moreover, the combination of RuR and rottlerin nearly abolished the nicardipine-resistant [Ca 2+ ] i elevation. PKCα, γ, δ, and ε, but not PKCβ, were detected by immunohistochemisty in smooth muscle cells of rat posterior cerebral arteries. Conclusions PKC isoforms have different roles in the development and maintain of the myogenic constriction in rat posterior cerebral arteries: cPKC (α and/or γ) mediates Ca 2+ sensitization in the initial phase, whereas PKCδ mediates [Ca 2+ ] i elevation via the activation of RuR-resistant cation channels in the sustained phase. Table. Effects of PKC inhibitors on pressure-induced constriction and [Ca 2+ ] i elevation

Sign in / Sign up

Export Citation Format

Share Document