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 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.

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.

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.

scholarly journals 5-Aminoimidazole-4-carboxamide riboside (AICAR) enhances GLUT2-dependent jejunal glucose transport: a possible role for AMPK

2005 ◽  
Vol 385 (2) ◽  
pp. 485-491 ◽  
Author(s):  
John WALKER ◽  
Humberto B. JIJON ◽  
Hugo DIAZ ◽  
Payam SALEHI ◽  
Thomas CHURCHILL ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Glucose Uptake ◽  
Glucose Transport ◽  
Western Blotting ◽  
Glucose Transporter ◽  
Mitogen Activated Protein Kinase ◽  
Mrna Levels ◽  
Energy Status ◽  
P38 Inhibitor ◽  
Mitogen Activated Protein

AMPK (AMP-activated protein kinase) is a key sensor of energy status within the cell. Activated by an increase in the AMP/ATP ratio, AMPK acts to limit cellular energy depletion by down-regulating selective ATP-dependent processes. The purpose of the present study was to determine the role of AMPK in regulating intestinal glucose transport. [3H]3-O-methyl glucose fluxes were measured in murine jejunum in the presence and absence of the AMPK activators AICAR (5-aminoimidazole-4-carboxamide riboside) and metformin and the p38 inhibitor, SB203580. To differentiate between a sodium-coupled (SGLT1) and diffusive (GLUT2) route of entry, fluxes were measured in the presence of the SGLT1 and GLUT2 inhibitors phloridzin and phloretin. Glucose transporter mRNA levels were measured by reverse transcriptase–PCR, and localization by Western blotting. Surface-expressed GLUT2 was assessed by luminal biotinylation. Activation of p38 mitogen-activated protein kinase was analysed by Western blotting. We found that treatment of jejunal tissue with AICAR resulted in enhanced net glucose uptake and was associated with phosphorylation of p38 mitogen-activated protein kinase. Inhibition of p38 abrogated the stimulation of AICAR-stimulated glucose uptake. Phloretin abolished the AICAR-mediated increase in glucose flux, whereas phloridzin had no effect, suggesting the involvement of GLUT2. In addition, AICAR decreased total protein levels of SGLT1, concurrently increasing levels of GLUT2 in the brush-border membrane. The anti-diabetic drug metformin, a known activator of AMPK, also induced the localization of GLUT2 to the luminal surface. We conclude that the activation of AMPK results in an up-regulation of non-energy requiring glucose uptake by GLUT2 and a concurrent down-regulation of sodium-dependent glucose transport.

scholarly journals A Novel 14-Kilodalton Protein Interacts with the Mitogen-Activated Protein Kinase Scaffold Mp1 on a Late Endosomal/Lysosomal Compartment

2001 ◽  
Vol 152 (4) ◽  
pp. 765-776 ◽  
Author(s):  
Winfried Wunderlich ◽  
Irene Fialka ◽  
David Teis ◽  
Arno Alpi ◽  
Andrea Pfeifer ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Complexes ◽  
Cell Types ◽  
Mapk Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Scaffolding Protein ◽  
Interacting Protein ◽  
Late Endosomes ◽  
Mitogen Activated Protein

We have identified a novel, highly conserved protein of 14 kD copurifying with late endosomes/lysosomes on density gradients. The protein, now termed p14, is peripherally associated with the cytoplasmic face of late endosomes/lysosomes in a variety of different cell types. In a two-hybrid screen with p14 as a bait, we identified the mitogen-activated protein kinase (MAPK) scaffolding protein MAPK/extracellular signal–regulated kinase (ERK) kinase (MEK) partner 1 (MP1) as an interacting protein. We confirmed the specificity of this interaction in vitro by glutathione S-transferase pull-down assays and by coimmunoprecipitation, cosedimentation on glycerol gradients, and colocalization. Moreover, expression of a plasma membrane–targeted p14 causes mislocalization of coexpressed MP1. In addition, we could reconstitute protein complexes containing the p14–MP1 complex associated with ERK and MEK in vitro. The interaction between p14 and MP1 suggests a MAPK scaffolding activity localized to the cytoplasmic surface of late endosomes/lysosomes, thereby combining catalytic scaffolding and subcellular compartmentalization as means to modulate MAPK signaling within a cell.

scholarly journals Adiponectin as Well as Compressive Forces Regulate in vitro β-Catenin Expression on Cementoblasts via Mitogen-Activated Protein Kinase Signaling Activation

2021 ◽  
Vol 9 ◽  
Author(s):  
Jiawen Yong ◽  
Julia von Bremen ◽  
Gisela Ruiz-Heiland ◽  
Sabine Ruf
Keyword(s):  
Protein Kinase ◽  
Signaling Pathways ◽  
Mapk Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Western Blots ◽  
Protein Levels ◽  
Mitogen Activated Protein ◽  
Gsk 3Β ◽  
Signaling Activation

We aimed to investigate the molecular effect that adiponectin exerts on cementoblasts especially in the presence of compressive forces. OCCM-30 cells (M. Somerman, NIH, NIDCR, United States) were used. Real-time reverse transcriptase–polymerase chain reaction (RT-PCR) and western blots were employed to verify if the mRNA and protein levels of adiponectin receptors (AdipoRs), mitogen-activated protein kinase (MAPK), and β-catenin signaling were influenced by compressive forces or adiponectin. Moreover, siRNAs targeting P38α, JNK1, ERK1, ERK2, and AdipoRs as well as pharmacological MAPK inhibition were performed. We found that compressive forces increase the expression of AdipoRs. Adiponectin and compression up-regulate P38α,JNK1, ERK1, and ERK2 as well as β-catenin gene expression. Western blots showed that co-stimuli activate the MAPK and β-catenin signaling pathways. MAPK inhibition alters the compression-induced β-catenin activation and the siRNAs targeting AdipoRs, P38α, and JNK1, showing the interaction of single MAPK molecules and β-catenin signaling in response to compression or adiponectin. Silencing by a dominantly negative version of P38α and JNK1 attenuates adiponectin-induced TCF/LEF reporter activation. Together, we found that light compressive forces activate β-catenin and MAPK signaling pathways. Adiponectin regulates β-catenin signaling principally by inactivating the GSK-3β kinase activity. β-Catenin expression was partially inhibited by MAPK blockade, indicating that MAPK plays a crucial role regulating β-catenin during cementogenesis. Moreover, adiponectin modulates GSK-3β and β-catenin mostly through AdipoR1. P38α is a key connector between β-catenin, TCF/LEF transcription, and MAPK signaling pathway.

scholarly journals Ras/Mitogen-Activated Protein Kinase Signaling Activates Ets-1 and Ets-2 by CBP/p300 Recruitment

2004 ◽  
Vol 24 (24) ◽  
pp. 10954-10964 ◽  
Author(s):  
Charles E. Foulds ◽  
Mary L. Nelson ◽  
Adam G. Blaszczak ◽  
Barbara J. Graves
Keyword(s):  
Transcription Factors ◽  
Protein Kinase ◽  
Direct Interaction ◽  
Mapk Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Creb Binding Protein ◽  
Mitogen Activated Protein ◽  
P300 Binding

ABSTRACT Cell signaling affects gene expression by regulating the activity of transcription factors. Here, we report that mitogen-activated protein kinase (MAPK) phosphorylation of Ets-1 and Ets-2, at a conserved site N terminal to their Pointed (PNT) domains, resulted in enhanced transactivation by preferential recruitment of the coactivators CREB binding protein (CBP) and p300. We discovered this phosphorylation-augmented interaction in an unbiased affinity chromatography screen of HeLa nuclear extracts by using either mock-treated or ERK2-phosphorylated ETS proteins as ligands. Binding between purified proteins demonstrated a direct interaction. Both the phosphoacceptor site, which lies in an unstructured region, and the PNT domain were required for the interaction. Minimal regions that were competent for induced CBP/p300 binding in vitro also supported MAPK-enhanced transcription in vivo. CBP coexpression potentiated MEK1-stimulated Ets-2 transactivation of promoters with Ras-responsive elements. Furthermore, CBP and Ets-2 interacted in a phosphorylation-enhanced manner in vivo. This study describes a distinctive interface for a transcription factor-coactivator complex and demonstrates a functional role for inducible CBP/p300 binding. In addition, our findings decipher the mechanistic link between Ras/MAPK signaling and two specific transcription factors that are relevant to both normal development and tumorigenesis.

scholarly journals Dietary Lysine Levels Improved Antioxidant Capacity and Immunity via the TOR and p38 MAPK Signaling Pathways in Grass Carp, Ctenopharyngodon idellus Fry

2021 ◽  
Vol 12 ◽  
Author(s):  
Dongyu Huang ◽  
Sahya Maulu ◽  
Mingchun Ren ◽  
Hualiang Liang ◽  
Xianping Ge ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Grass Carp ◽  
Control Diet ◽  
Mapk Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Target Of Rapamycin ◽  
Interleukin 4 ◽  
Mrna Levels ◽  
Nuclear Factor ◽  
Mitogen Activated Protein

An 8-week rearing trial was designed to appraise the dietary lysine levels on intestinal antioxidant capacity and immunity of grass carp fry. Six practical diets were prepared with graded levels of lysine (1.44, 1.79, 1.97, 2.44, 2.56 and 2.87% dry matter), and these diets were fed to grass carp fry. The results showed that the activities of intestinal antioxidant factors including catalase and glutathione peroxidase were markedly improved by the 2.44% dietary lysine compared with the control diet (1.44% dietary lysine) (P &lt; 0.05). In terms of antioxidants, compared with the control diet, the 2.44% diet markedly upregulated the mRNA expression levels of target of rapamycin, S6 kinase1 and nuclear factor erythroid 2-related factor 2 pathway-related antioxidant genes, containing catalase and glutathione peroxidase 1α (P &lt; 0.05) and downregulated the mRNA levels of Kelch-like ECH-associated protein 1 (P &gt; 0.05). The mRNA levels of 4E-binding protein 2 showed the opposite trend compared with those of target of rapamycin, and the minimum value was observed in the group of 1.97% dietary lysine (P &lt; 0.05). In terms of immunity, compared with the 1.44% diet, the 2.44% diet markedly suppressed the intestinal p38 mitogen-activated protein kinase and interferon γ2 mRNA levels (P &lt; 0.05). Moreover, nuclear factor-kappa B p65, tumor necrosis factor α, interleukin 6, interleukin 8, and interleukin 15 mRNA levels all exhibited the same trend as p38 mitogen-activated protein kinase and interferon γ2; however, the difference among all the lysine treatments groups was not significant (P &gt; 0.05). The anti-inflammatory cytokines transforming growth factor β2 and interleukin 4/13B mRNA levels in the intestine were remarkably upregulated by high dietary lysine levels (2.56 and 2.87%) (P &lt; 0.05), and when the dietary lysine level reached 2.44%, the interleukin 4/13A mRNA levels were strikingly increased (P &lt; 0.05). Overall, the data suggested that 2.44% dietary lysine could strengthen the immune and antioxidant capacities of grass carp fry via activating the target of rapamycin (TOR) signaling pathway, and suppressing the p38 mitogen-activated protein kinase (p38 MAPK) signaling pathway, which then improve the survival rate.

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