scholarly journals Fever-like temperature modification differentially affects in vitro signaling of bradykinin B1 and B2 receptors

2011 ◽  
Vol 392 (11) ◽  
Author(s):  
Jasmin Leschner ◽  
Larisa Ring ◽  
Jens Feierler ◽  
Klaus Dinkel ◽  
Marianne Jochum ◽  
...  
Keyword(s):  
Mapk Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Temperature Sensitive ◽  
Hek 293 ◽  
Mapk Activity ◽  
Mitogen Activated Protein ◽  
Surface Receptor ◽  
B2 Receptors ◽  
Stimulation Of

Abstract The bradykinin (BK) B2 and B1 receptors (B2R, B1R) belong to the rhodopsin-like G protein-coupled receptors (GPCRs) and are involved in (patho)physiological processes such as blood pressure regulation or inflammation. They mediate the effects of the pro-inflammatory peptides bradykinin/kallidin and desArg9-BK/desArg10-kallidin, respectively. Whereas the B2R is constitutively expressed and gets internalized upon activation, the B1R is especially induced by inflammatory mediators and responds to stimulation with increased surface receptor numbers. Stimulation of both receptors activates phospholipase Cβ (PLCβ) and mitogen activated protein kinase (MAPK) signaling. Because inflammatory processes are characterized by heat (fever), we analyzed the effect of increased temperature (41°C vs. 37°C) on B1R and B2R signaling in HEK 293 and IMR 90 cells. Our results show that signaling of both receptors is temperature-sensitive, however to a different extent and with regard to the investigated pathways. Comparing PLCβ activity and Ca2+-regulated signals, a temperature-dependent increase was only observed for B1R but not for B2R activation, whereas MAPK activities were doubled at 41°C for both receptors. Taken together, our findings suggest that the observed temperature sensitivity of B1R-induced PLCβ activation is B1R-specific. In contrast, the enhanced stimulation of MAPK activity under hyperthermic conditions appears to be a common phenomenon for GPCRs.

scholarly journals p38 Mitogen-Activated Protein Kinase (MAPK) Is a Key Mediator in Glucocorticoid-Induced Apoptosis of Lymphoid Cells: Correlation between p38 MAPK Activation and Site-Specific Phosphorylation of the Human Glucocorticoid Receptor at Serine 211

2005 ◽  
Vol 19 (6) ◽  
pp. 1569-1583 ◽  
Author(s):  
Aaron L. Miller ◽  
M. Scott Webb ◽  
Alicja J. Copik ◽  
Yongxin Wang ◽  
Betty H. Johnson ◽  
...  
Keyword(s):  
P38 Mapk ◽  
Mapk Signaling ◽  
Lymphoid Cells ◽  
Mitogen Activated Protein Kinase ◽  
Enzymatic Assays ◽  
Mapk Activity ◽  
Mitogen Activated Protein ◽  
Elevated Activity ◽  
Induced Apoptosis

Abstract Glucocorticoids (GCs) induce apoptosis in lymphoid cells through activation of the GC receptor (GR). We have evaluated the role of p38, a MAPK, in lymphoid cell apoptosis upon treatment with the synthetic GCs dexamethasone (Dex) or deacylcortivazol (DAC). The highly conserved phosphoprotein p38 MAPK is activated by specific phosphorylation of its threonine180 and tyrosine182 residues. We show that Dex and DAC stimulate p38 MAPK phosphorylation and increase the mRNA of MAPK kinase 3, a specific immediate upstream activator of p38 MAPK. Enzymatic assays confirmed elevated activity of p38 MAPK. Pharmacological inhibition of p38 MAPK activity was protective against GC-driven apoptosis in human and mouse lymphoid cells. In contrast, inhibition of the MAPKs, ERK and cJun N-terminal kinase, enhanced apoptosis. Activated p38 MAPK phosphorylates specific downstream targets. Because phosphorylation of the GR is affected by MAPKs, we examined its phosphorylation state in our system. We found serine 211 of the human GR to be a substrate for p38 MAPK both in vitro and intracellularly. Mutation of this site to alanine greatly diminished GR-driven gene transcription and apoptosis. Our results clearly demonstrate a role for p38 MAPK signaling in the pathway of GC-induced apoptosis of lymphoid cells.

Role of the PI3K/PKB signaling pathway in cAMP-mediated translocation of rat liver Ntcp

1999 ◽  
Vol 277 (6) ◽  
pp. G1165-G1172 ◽  
Author(s):  
Cynthia R. L. Webster ◽  
M. Sawkat Anwer
Keyword(s):  
Plasma Membrane ◽  
Protein Kinase ◽  
Signaling Pathway ◽  
Mitogen Activated Protein Kinase ◽  
Phosphatidylinositol 3 Kinase ◽  
Pi3k Inhibitors ◽  
Mapk Activity ◽  
Mitogen Activated Protein ◽  
Pi3k Signaling Pathway ◽  
Stimulation Of

cAMP stimulates Na+-taurocholate (TC) cotransport by translocating the Na+-TC-cotransporting peptide (Ntcp) to the plasma membrane. The present study was undertaken to determine whether the phosphatidylinositol-3-kinase (PI3K)-signaling pathway is involved in cAMP-mediated translocation of Ntcp. The ability of cAMP to stimulate TC uptake declined significantly when hepatocytes were pretreated with PI3K inhibitors wortmannin or LY-294002. Wortmannin inhibited cAMP-mediated translocation of Ntcp to the plasma membrane. cAMP stimulated protein kinase B (PKB) activity by twofold within 5 min, an effect inhibited by wortmannin. Neither basal mitogen-activated protein kinase (MAPK) activity nor cAMP-mediated inhibition of MAPK activity was affected by wortmannin. cAMP also stimulated p70S6K activity. However, rapamycin, an inhibitor of p70S6K, failed to inhibit cAMP-mediated stimulation of TC uptake, indicating that the effect of cAMP is not mediated via p70S6K. Cytochalasin D, an inhibitor of actin filament formation, inhibited the ability of cAMP to stimulate TC uptake and Ntcp translocation. Together, these results suggest that the stimulation of TC uptake and Ntcp translocation by cAMP may be mediated via the PI3K/PKB signaling pathway and requires intact actin filaments.

scholarly journals Neuroprotective potential of isothiocyanates in an in vitro model of neuroinflammation

2020 ◽  
Author(s):  
Tiziana Latronico ◽  
Marilena Larocca ◽  
Serafina Milella ◽  
Anna Fasano ◽  
Rocco Rossano ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Neurological Diseases ◽  
Primary Cultures ◽  
Allyl Isothiocyanate ◽  
Mapk Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Pcr Analysis ◽  
Ros Production ◽  
Mitogen Activated Protein

AbstractIsothiocyanates (ITCs), present as glucosinolate precursors in cruciferous vegetables, have shown anti-inflammatory, antioxidant and anticarcinogenic activities. Here, we compared the effects of three different ITCs on ROS production and on the expression of matrix metalloproteinase (MMP)-2 and -9, which represent important pathogenetic factors of various neurological diseases. Primary cultures of rat astrocytes were activated by LPS and simultaneously treated with different doses of Allyl isothiocyanate (AITC), 2-Phenethyl isothiocyanate (PEITC) and 2-Sulforaphane (SFN). Results showed that SFN and PEITC were able to counteract ROS production induced by H2O2. The zymographic analysis of cell culture supernatants evidenced that PEITC and SFN were the most effective inhibitors of MMP-9, whereas, only SFN significantly inhibited MMP-2 activity. PCR analysis showed that all the ITCs used significantly inhibited both MMP-2 and MMP-9 expression. The investigation on the mitogen-activated protein kinase (MAPK) signaling pathway demonstrated that ITCs modulate MMP transcription by inhibition of extracellular-regulated protein kinase (ERK) activity. Results of this study suggest that ITCs could be promising nutraceutical agents for the prevention and complementary treatment of neurological diseases associated with MMP involvement.

Insulin induces Ca2+ influx into isolated rat hepatocyte couplets

1997 ◽  
Vol 272 (6) ◽  
pp. G1425-G1432 ◽  
Author(s):  
K. Benzeroual ◽  
G. van de Werve ◽  
S. Meloche ◽  
L. Mathe ◽  
A. Romanelli ◽  
...  
Keyword(s):  
Peptide Hormone ◽  
Insulin Binding ◽  
Mitogen Activated Protein Kinase ◽  
Rat Hepatocyte ◽  
Mapk Activity ◽  
Mitogen Activated Protein ◽  
Insulin Dependent ◽  
Dose Dependent Increase ◽  
Isolated Rat ◽  
Stimulation Of

Isolated rat hepatocyte couplets were used to study the direct effect of insulin on intracellular Ca2+ homeostasis. Insulin induced a dose-dependent increase in hepatocellular Ca2+ that was gradual, generally monophasic, and reversible. Chelation of extracellular Ca2+ abolished the insulin-induced Ca2+ response, and this suppression was not related to an effect on insulin binding, as indicated by displacement studies. We thus tested the effect of several Ca2+ channel inhibitors on insulin-induced Ca2+ influx. Verapamil at 20 or 200 microM was without effect, whereas 500 microM nickel and 50 microM gadolinium strongly inhibited insulin-induced Ca2+ entry. Finally, we tested whether insulin-induced Ca2+ movements were implicated in the stimulation of mitogen-activated protein kinase (MAPK) activity, which we measured with the use of an immune-complex assay. Verapamil was without effect on the insulin-dependent stimulation of p44mapk activity, whereas addition of ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid, nickel, or gadolinium strongly inhibited the effect of the peptide hormone. Our results indicate that insulin triggers Ca2+ influx into hepatocytes, possibly through the opening of channels on the plasma membrane, and that this effect is important for insulin activation of MAPK.

scholarly journals Phosphorylation of the ARE-binding protein DAZAP1 by ERK2 induces its dissociation from DAZ

2006 ◽  
Vol 399 (2) ◽  
pp. 265-273 ◽  
Author(s):  
Simon Morton ◽  
Huei-Ting Yang ◽  
Ntsane Moleleki ◽  
David G. Campbell ◽  
Philip Cohen ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Rna Binding ◽  
Binding Protein ◽  
Mitogen Activated Protein Kinase ◽  
Raw 264.7 ◽  
Raw 264.7 Macrophages ◽  
Hek 293 ◽  
Extracellular Signal ◽  
Mitogen Activated Protein

A protein in RAW 264.7 macrophages, which became phosphorylated in response to LPS (lipopolysaccharide), was identified as the RNA-binding protein called DAZAP1 [DAZ (deleted in azoospermia)-associated protein 1]. The phosphorylation of this protein was prevented by specific inhibition of MKK1 [MAPK (mitogen-activated protein kinase) kinase 1], indicating that it was phosphorylated via the classical MAPK cascade. Further experiments showed that DAZAP1 was phosphorylated stoichiometrically in vitro by ERK2 (extracellular-signal-regulated protein kinase 2) at two Thr-Pro sequences (Thr269 and Thr315), and that both sites became phosphorylated in HEK-293 (human embryonic kidney 293) cells in response to PMA or EGF (epidermal growth factor), or RAW 264.7 macrophages in response to LPS. Phosphorylation induced by each stimulus was prevented by two structurally distinct inhibitors of MKK1 (PD184352 and U0126), demonstrating that DAZAP1 is a physiological substrate for ERK1/ERK2. The mutation of Thr269 and Thr315 to aspartate or the phosphorylation of these residues caused DAZAP1 to dissociate from its binding partner DAZ. DAZ interacts with PABP [poly(A)-binding protein] and thereby stimulates the translation of mRNAs containing short poly(A) tails [Collier, Gorgoni, Loveridge, Cooke and Gray (2005) EMBO J. 24, 2656–2666]. In the present study we have shown that DAZ cannot bind simultaneously to DAZAP1 and PABP, and suggest that the phosphorylation-induced dissociation of DAZ and DAZAP1 may allow the former to stimulate translation by interacting with PABP.

scholarly journals Insulin acts via mitogen-activated protein kinase phosphorylation in rabbit blastocysts

Reproduction ◽  
2004 ◽  
Vol 128 (5) ◽  
pp. 517-526 ◽  
Author(s):  
Anne Navarrete Santos ◽  
Sarah Tonack ◽  
Michaela Kirstein ◽  
Marie Pantaleon ◽  
Peter Kaye ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Glucose Transport ◽  
Mapk Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Preimplantation Embryos ◽  
Mrna Levels ◽  
Glut4 Translocation ◽  
Mitogen Activated Protein ◽  
Rabbit Embryos

The addition of insulin during in vitro culture has beneficial effects on rabbit preimplantation embryos leading to increased cell proliferation and reduced apoptosis. We have previously described the expression of the insulin receptor (IR) and the insulin-responsive glucose transporters (GLUT) 4 and 8 in rabbit preimplantation embryos. However, the effects of insulin on IR signaling and glucose metabolism have not been investigated in rabbit embryos. In the present study, the effects of 170 nM insulin on IR, GLUT4 and GLUT8 mRNA levels, Akt and Erk phosphorylation, GLUT4 translocation and methyl glucose transport were studied in cultured day 3 to day 6 rabbit embryos. Insulin stimulated phosphorylation of the mitogen-activated protein kinase (MAPK) Erk1/2 and levels of IR and GLUT4 mRNA, but not phosphorylation of the phosphatidylinositol 3-kinase-dependent protein kinase, Akt, GLUT8 mRNA levels, glucose uptake or GLUT4 translocation. Activation of the MAPK signaling pathway in the absence of GLUT4 translocation and of a glucose transport response suggest that in the rabbit preimplantation embryo insulin is acting as a growth factor rather than a component of glucose homeostatic control.

scholarly journals Identification of RAS-Mitogen-Activated Protein Kinase Signaling Pathway Modulators in an ERF1 Redistribution® Screen

2006 ◽  
Vol 11 (4) ◽  
pp. 423-434 ◽  
Author(s):  
Charlotta Grånäs ◽  
Betina Kerstin Lundholt ◽  
Frosty Loechel ◽  
Hans-Christian Pedersen ◽  
Sara Petersen Bjørn ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Signaling Pathway ◽  
Kinase Inhibitors ◽  
Mapk Pathway ◽  
Mapk Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Mitogen Activated Protein ◽  
A Cell ◽  
Protein Kinase Signaling

The RAS-mitogen-activated protein kinase (MAPK) signaling pathway has a central role in regulating the proliferation and survival of both normal and tumor cells. This pathway has been 1 focus area for the development of anticancer drugs, resulting in several compounds, primarily kinase inhibitors, in clinical testing. The authors have undertaken a cell-based, high-throughput screen using a novel ERF1 Redistribution® assay to identify compounds that modulate the signaling pathway. The hit compounds were subsequently tested for activity in a functional cell proliferation assay designed to selectively detect compounds inhibiting the proliferation of MAPK pathway-dependent cancer cells. The authors report the identification of 2 cell membrane-permeable compounds that exhibit activity in the ERF1 Redistribution® assay and selectively inhibit proliferation of MAPK pathway-dependent malignant melanoma cells at similar potencies (IC50 =< 5 μM). These compounds have drug-like structures and are negative in RAF, MEK, and ERK in vitro kinase assays. Drugs belonging to these compound classes may prove useful for treating cancers caused by excessive MAPK pathway signaling. The results also show that cell-based, high-content Redistribution® screens can detect compounds with different modes of action and reveal novel targets in a pathway known to be disease relevant.

Aggravation of ADAMTS and Matrix Metalloproteinase Production and Role of ERK1/2 Pathway in the Interaction of Osteoarthritic Subchondral Bone Osteoblasts and Articular Cartilage Chondrocytes — Possible Pathogenic Role in Osteoarthritis

2012 ◽  
Vol 39 (3) ◽  
pp. 621-634 ◽  
Author(s):  
INDIRA PRASADAM ◽  
ROSS CRAWFORD ◽  
YIN XIAO
Keyword(s):  
Articular Cartilage ◽  
Signaling Pathway ◽  
Subchondral Bone ◽  
Mapk Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Mitogen Activated Protein ◽  
Coculture Model ◽  
Degradative Enzymes ◽  
A Disintegrin And Metalloproteinase

Objective.Degradative enzymes, such as A disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) and matrix metalloproteinases (MMP), play key roles in development of osteoarthritis (OA). We investigated if crosstalk between subchondral bone osteoblasts (SBO) and articular cartilage chondrocytes (ACC) in OA alters the expression and regulation of ADAMTS5, ADAMTS4, MMP-1, MMP-2, MMP-3, MMP-8, MMP-9, and MMP-13, and also tested the possible involvement of mitogen-activated protein kinase (MAPK) signaling pathway during this process.Methods.ACC and SBO were isolated from normal and OA patients. An in vitro coculture model was developed to study the regulation of ADAMTS and MMP under normal and OA joint crosstalk conditions. The MAPK-ERK inhibitor PD98059 was applied to delineate the involvement of specific pathways during this interaction process.Results.Indirect coculture of OA SBO with normal ACC resulted in significantly increased expression of ADAMTS5, ADAMTS4, MMP-2, MMP-3, and MMP-9 in ACC, whereas coculture of OA ACC led to increased MMP-1 and MMP-2 expression in normal SBO. Upregulation of ADAMTS and MMP under these conditions was correlated with activation of the MAPK-ERK1/2 signaling pathway, and addition of the MAPK-ERK inhibitor PD98059 reversed the overexpression of ADAMTS and MMP in cocultures.Conclusion.These results add to the evidence that in human OA, altered bidirectional signals between SBO and ACC significantly influence the critical features of both cartilage and bone by producing abnormal levels of ADAMTS and MMP. We have demonstrated for the first time that this altered crosstalk was mediated by the phosphorylation of MAPK-ERK1/2 signaling pathway.

scholarly journals Mating and Pathogenic Development of the Smut Fungus Ustilago maydis Are Regulated by One Mitogen-Activated Protein Kinase Cascade

2003 ◽  
Vol 2 (6) ◽  
pp. 1187-1199 ◽  
Author(s):  
Philip Müller ◽  
Gerhard Weinzierl ◽  
Andreas Brachmann ◽  
Michael Feldbrügge ◽  
Regine Kahmann
Keyword(s):  
Protein Kinase ◽  
Ustilago Maydis ◽  
Mapk Signaling ◽  
Tube Formation ◽  
Mitogen Activated Protein Kinase ◽  
Phytopathogenic Fungus ◽  
Mapk Kinase ◽  
Mitogen Activated Protein ◽  
Kinase Cascade

ABSTRACT In the phytopathogenic fungus Ustilago maydis, pheromone-mediated cell fusion is a prerequisite for the generation of the infectious dikaryon. The pheromone signal elevates transcription of the pheromone genes and elicits formation of conjugation hyphae. Cyclic AMP and mitogen-activated protein kinase (MAPK) signaling are involved in this process. The MAPK cascade is presumed to be composed of Ubc4 (MAPK kinase kinase), Fuz7 (MAPK kinase), and Ubc3/Kpp2 (MAPK). We isolated the kpp4 gene and found it to be allelic to ubc4. Epistasis analyses with constitutively active alleles of kpp4 and fuz7 substantiate that Kpp4, Fuz7, and Kpp2/Ubc3 are components of the same module. Moreover, we demonstrate that Fuz7 activates Kpp2 and shows interactions in vitro. Signaling via this cascade regulates expression of pheromone-responsive genes, presumably through acting on the transcription factor Prf1. Interestingly, the same cascade is needed for conjugation tube formation, and this process does not involve Prf1. In addition, fuz7 as well as kpp4 deletion strains are nonpathogenic, while kpp2 deletion mutants are only attenuated in pathogenesis. Here we show that strains expressing the unphosphorylatable allele kpp2T182A/Y184F are severely affected in tumor induction and display defects in early infection-related differentiation.

GLUT4 translocation precedes the stimulation of glucose uptake by insulin in muscle cells: potential activation of GLUT4 via p38 mitogen-activated protein kinase

2001 ◽  
Vol 359 (3) ◽  
pp. 639-649 ◽  
Author(s):  
Romel SOMWAR ◽  
David Y. KIM ◽  
Gary SWEENEY ◽  
Carol HUANG ◽  
Wenyan NIU ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Glucose Uptake ◽  
P38 Mapk ◽  
Mitogen Activated Protein Kinase ◽  
Kinase Assay ◽  
Glut4 Translocation ◽  
L6 Myotubes ◽  
Mitogen Activated Protein ◽  
Stimulation Of

We previously reported that SB203580, an inhibitor of p38 mitogen-activated protein kinase (p38 MAPK), attenuates insulin-stimulated glucose uptake without altering GLUT4 translocation. These results suggested that insulin might activate GLUT4 via a p38 MAPK-dependent pathway. Here we explore this hypothesis by temporal and kinetic analyses of the stimulation of GLUT4 translocation, glucose uptake and activation of p38 MAPK isoforms by insulin. In L6 myotubes stably expressing GLUT4 with an exofacial Myc epitope, we found that GLUT4 translocation (t1/2 = 2.5min) preceded the stimulation of 2-deoxyglucose uptake (t1/2 = 6min). This segregation of glucose uptake from GLUT4 translocation became more apparent when the two parameters were measured at 22°C. Preincubation with the p38 MAPK inhibitors SB202190 and SB203580 reduced insulin-stimulated transport of either 2-deoxyglucose or 3-O-methylglucose by 40–60%. Pretreatment with SB203580 lowered the apparent transport Vmax of insulin-mediated 2-deoxyglucose and 3-O-methylglucose without any significant change in the apparent Km for either hexose. The IC50 values for the partial inhibition of 2-deoxyglucose uptake by SB202190 and SB203580 were 1 and 2μM respectively, and correlated with the IC50 for full inhibition of p38 MAPK by the two inhibitors in myotubes (2 and 1.4μM, respectively). Insulin caused a dose- (EC50 = 15nM) and time- (t1/2 = 3min) dependent increase in p38 MAPK phosphorylation, which peaked at 10min (2.3±0.3-fold). In vitro kinase assay of immunoprecipitates from insulin-stimulated myotubes showed activation of p38α (2.6±0.3-fold) and p38β (2.3±0.2-fold) MAPK. These results suggest that activation of GLUT4 follows GLUT4 translocation and that both mechanisms contribute to the full stimulation of glucose uptake by insulin. Furthermore, activation of GLUT4 may occur via an SB203580-sensitive pathway, possibly involving p38 MAPK.

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