WorldWideScience

Sample records for two-site mapk dephosphorylation

  1. Brief Glutamine Pretreatment Increases Alveolar Macrophage CD163/Heme Oxygenase-1/p38-MAPK Dephosphorylation Pathway and Decreases Capillary Damage but Not Neutrophil Recruitment in IL-1/LPS-Insufflated Rats.

    Science.gov (United States)

    Fernandez-Bustamante, Ana; Agazio, Amanda; Wilson, Paul; Elkins, Nancy; Domaleski, Luke; He, Qianbin; Baer, Kaily A; Moss, Angela F D; Wischmeyer, Paul E; Repine, John E

    2015-01-01

    Glutamine (GLN) attenuates acute lung injury (ALI) but its effect on alveolar macrophages is unknown. We hypothesized that GLN pretreatment would induce the anti-inflammatory CD163/heme oxygenase (HO)-1/p38-MAPK dephosphorylation pathway in alveolar macrophages and reduce ALI in rats insufflated with interleukin-1 (IL-1) and lipopolysaccharide (LPS). Male Sprague-Dawley rats were randomized to the following groups: GLN-IL-1/LPS-, GLN+IL-1/LPS-, GLN-IL-1/LPS+, and GLN+IL-1/LPS+. GLN pretreatment was given via gavage (1 g/kg L-alanyl-L-glutamine) daily for 2 days. ALI was subsequently induced by insufflating 50 ng IL-1 followed by 5mg/kg E.coli LPS. After 24h, bronchoalveolar lavage (BAL) protein, lactate dehydrogenase (LDH) and neutrophil concentrations were analyzed. BAL alveolar macrophage CD163+ expression, HO-1 and p38-MAPK concentrations were measured, as well as alveolar macrophage tumor necrosis factor (TNF)-α and interleukin (IL)-10 concentrations. Histology and immunofluorescence studies were also performed. Following IL-1/LPS insufflation, GLN pretreated rats had significantly decreased BAL protein and LDH concentrations, but not BAL neutrophil counts, compared to non-GLN pretreated rats. The number of alveolar macrophages and the number of CD163+ macrophages were significantly increased in GLN pretreated IL-1/LPS-insufflated rats compared to non-GLN pretreated, IL-1/LPS-insufflated rats. GLN pretreatment before IL-1/LPS also significantly increased HO-1 concentrations and dephosphorylated p38-MAPK levels but not cytokine levels in alveolar macrophages. Immunofluorescence localized CD163 and HO-1 in alveolar macrophages. Short-term GLN pretreatment activates the anti-inflammatory CD163/HO-1/p38-MAPK dephosphorylation pathway of alveolar macrophages and decreases capillary damage but not neutrophil recruitment in IL-1/LPS-insufflated rats.

  2. Characterization of Entamoeba histolytica-induced dephosphorylation in Jurkat cells

    Indian Academy of Sciences (India)

    J E Teixeira; B J Mann

    2002-11-01

    Entamoeba histolytica killing of host cells is contact dependent and mediated by a Gal/GalNAc lectin. Upon contact with amoeba a rapid and extensive dephosphorylation of tyrosine phosphorylated host cell proteins is observed. This effect is mediated by the Gal/GalNAc lectin. However, it requires intact cells, as purified lectin failed to induce dephosphorylation in Jurkat cells. The nonpathogenic, but morphologically identical amoeba, Entamoeba moshkovskii also did not induce dephosphorylation in target cells. Treatment of Jurkat cells with phosphotyrosine phosphatase inhibitors has shown that a host phosphatase is responsible for dephosphorylation. However, it was found that the CD45 phosphotase was not necessary for dephosphorylation of host cell proteins.

  3. Expression of MaMAPK Gene in Seedlings of Malus L. under Water Stress

    Institute of Scientific and Technical Information of China (English)

    Li-Xin PENG; Ling-Kun GU; Cheng-Chao ZHENG; De-Quan LI; Huai-Rui SHU

    2006-01-01

    Seedlings of three species of Malus were used to study the expression of mitogen-activated protein kinase (MAPK) in response to water stress: Malus hupehensis, a drought-sensitive species; Malus sieversii, a drought-tolerant species; and Malus micromalus, a middle type. Results showed that Malus MAPK (MaMAPK, GenBank accession No. AF435805) was expressed in both roots and leaves of seedlings of the three Malus species treated with 20% polyethylene glycol for different time periods. Expression levels peaked at 1.5 h after treatment with polyethylene glycol, then decreased to their lowest levels. Liquid kinase assays indicated that the dynamic changes of MAPK activity were very similar to those of the relative expression of MaMAPK mRNA. However, the peak of the former occurred slightly behind the latter. It was noticed that, although the kinase activity decreased after the peak, it was still higher than that of the control during the whole time period. These results suggested that MaMAPK was regulated not only by water stress at the transcription level, but also by phosphorylation and dephosphorylation at the protein level. In addition,of these three apple species, the highest MAPK activity and MaMAPK expression level was found in M.sieversii, followed by M. micromalus and M. hupehensis, suggesting that MAPK might be correlated with drought tolerance in these three species. The different expression levels might be one of the molecular mechanisms of the different drought tolerances in Malus.

  4. Induced gelation in a two-site spatial coagulation model

    OpenAIRE

    Siegmund-Schultze, Rainer; Wagner, Wolfgang

    2006-01-01

    A two-site spatial coagulation model is considered. Particles of masses $m$ and $n$ at the same site form a new particle of mass $m+n$ at rate $mn$. Independently, particles jump to the other site at a constant rate. The limit (for increasing particle numbers) of this model is expected to be nondeterministic after the gelation time, namely, one or two giant particles randomly jump between the two sites. Moreover, a new effect of induced gelation is observed--the gelation happening at the site...

  5. Stereospecificity of inositol hexakisphosphate dephosphorylation by Paramecium phytase

    NARCIS (Netherlands)

    Kaay, Jeroen van der; Haastert, Peter J.M. van

    1995-01-01

    InsP(6) is an abundant compound in many micro-organisms, plants and animal cells. Its function and route of synthesis are still largely unknown. Degradation of InsP(6) is mediated by phytase, which in most organisms dephosphorylates InsP(6) in a relatively non-specific way. In the micro-organism Par

  6. A two-site bipolaron model for organic magnetoresistance

    Science.gov (United States)

    Wagemans, W.; Bloom, F. L.; Bobbert, P. A.; Wohlgenannt, M.; Koopmans, B.

    2008-04-01

    The recently proposed bipolaron model for large "organic magnetoresistance" (OMAR) at room temperature is extended to an analytically solvable two-site scheme. It is shown that even this extremely simplified approach reproduces some of the key features of OMAR, viz., the possibility to have both positive and negative magnetoresistance, as well as its universal line shapes. Specific behavior and limiting cases are discussed. Extensions of the model, to guide future experiments and numerical Monte Carlo studies, are suggested.

  7. Canonical and alternative MAPK signaling.

    Science.gov (United States)

    Pimienta, Genaro; Pascual, Jaime

    2007-11-01

    The archetype of MAPK cascade activation is somewhat challenged by the most recent discovery of unexpected phosphorylation patterns, alternative activation mechanisms and sub-cellular localization, in various members of this protein kinase family. In particular, activation by autophosphorylation pathways has now been described for the three best understood MAPK subgroups: ERK1/2; JNK1/2 and p38 alpha/beta. Also, a form of dosage compensation between homologs has been shown to occur in the case of ERK1/2 and JNK1/2. In this paper we summarize the MAPK activation pathway, with an emphasis on non-canonical examples. We use this information to propose a model for MAPK signal transduction that considers a cross-talk between MAPKs with different activation loop sequence motifs and unique C-terminal extensions. We highlight the occurrence of non-canonical substrate specificity during MAPK auto-activation, in strong connection with MAPK homo- and hetero-dimerization events.

  8. The Small C-terminal Domain Phosphatase 1 Inhibits Cancer Cell Migration and Invasion by Dephosphorylating Ser(P)68-Twist1 to Accelerate Twist1 Protein Degradation.

    Science.gov (United States)

    Sun, Tong; Fu, Junjiang; Shen, Tao; Lin, Xia; Liao, Lan; Feng, Xin-Hua; Xu, Jianming

    2016-05-27

    Twist1 is a basic helix-loop-helix transcription factor that strongly promotes epithelial-to-mesenchymal transition, migration, invasion, and metastasis of cancer cells. The MAPK-phosphorylated Twist1 on its serine 68 (Ser(P)(68)-Twist1) has a significantly enhanced stability and function to drive cancer cell invasion and metastasis. However, the phosphatase that dephosphorylates Ser(P)(68)-Twist1 and destabilizes Twist1 has not been identified and characterized. In this study, we screened a serine/threonine phosphatase cDNA expression library in HEK293T cells with ectopically coexpressed Twist1. We found that the small C-terminal domain phosphatase 1 (SCP1) specifically dephosphorylates Ser(P)(68)-Twist1 in both cell-free reactions and living cells. SCP1 uses its amino acid residues 43-63 to interact with the N terminus of Twist1. Increased SCP1 expression in cells decreased Ser(P)(68)-Twist1 and total Twist1 proteins, whereas knockdown of SCP1 increased Ser(P)(68)-Twist1 and total Twist1 proteins. Furthermore, the levels of SCP1 are negatively correlated with Twist1 protein levels in several cancer cell lines. SCP1-dephosphorylated Twist1 undergoes fast degradation via the ubiquitin-proteasome pathway. Importantly, an increase in SCP1 expression in breast cancer cells with either endogenous or ectopically expressed Twist1 largely inhibits the Twist1-induced epithelial-to-mesenchymal transition phenotype and the migration and invasion capabilities of these cells. These results indicate that SCP1 is the phosphatase that counterregulates the MAPK-mediated phosphorylation of Ser(68)-Twist1. Thus, an increase in SCP1 expression and activity may be a useful strategy for eliminating the detrimental roles of Twist1 in cancer cells.

  9. Effects of enzymatic dephosphorylation on infant in vitro gastrointestinal digestibility of milk protein concentrate.

    Science.gov (United States)

    Liu, Dasong; Wang, Yuanyuan; Yu, Yun; Hu, Jinhua; Lu, Naiyan; Regenstein, Joe M; Wang, Miao; Zhou, Peng

    2016-04-15

    This study investigated the effects of dephosphorylation extent on infant in vitro gastric clotting property and gastrointestinal digestibility of milk protein concentrate. Dephosphorylation was affected by phosphatase type and incubation pH. A series of milk protein concentrate with 0-69% dephosphorylation were obtained by incubation with calf intestinal alkaline phosphatase at pH 6.5 for 0-420 min. Both β- and αs1-caseins in the modified milk protein concentrate showed multiply dephosphorylated isoforms with different numbers of phosphate groups depending on the extent of dephosphorylation. With increased dephosphorylation of milk protein concentrate, the gastric clotting extent decreased and the gastrointestinal digestibility increased under infant in vitro conditions. These results suggested the potential of developing a dephosphorylated milk protein concentrate, with improved gastric clotting property and gastrointestinal digestibility, to simulate the multiply phosphorylated patterns of human casein and hence to further the humanization of infant formula on a molecular level.

  10. 4EBP1 Is Dephosphorylated by Respiratory Syncytial Virus Infection.

    Science.gov (United States)

    Pérez-Gil, Gustavo; Landa-Cardeña, Adriana; Coutiño, Rocío; García-Román, Rebeca; Sampieri, Clara L; Mora, Silvia I; Montero, Hilda

    2015-01-01

    Respiratory syncytial virus (RSV) requires protein biosynthesis machinery to generate progeny. There is evidence that RSV might alter some translation components since stress granules are formed in their host cells. Consistent with these observations, we found that RSV induces dephosphorylation of 4EBP1 (eIF4E-binding protein), an important cellular translation factor. Our results show no correlation between the 4EBP1 dephosphorylation time and the decrease in the global rate of protein synthesis. Interestingly, treatment with rapamycin stimulates virus generation. The results suggest that RSV is a virus that still contains unknown mechanisms involved in the translation of their mRNAs through the alteration or modification of some translation factors, such as 4EBP1, possibly to favor its replicative cycle. © 2015 S. Karger AG, Basel.

  11. Dephosphorylation of Centrins by Protein Phosphatase 2C and

    Directory of Open Access Journals (Sweden)

    Marie-Christin Thissen

    2009-01-01

    Full Text Available In the present study, we identified protein phosphatases dephosphorylating centrins previously phosphorylated by protein kinase CK2. The following phosphatases known to be present in the retina were tested: PP1, PP2A, PP2B, PP2C, PP5, and alkaline phosphatase. PP2C and were capable of dephosphorylating P-Thr138-centrin1 most efficiently. PP2C was inactive and the other retinal phosphatases also had much less or no effect. Similar results were observed for centrins 2 and 4. Centrin3 was not a substrate for CK2. The results suggest PP2C and to play a significant role in regulating the phosphorylation status of centrins in vivo.

  12. Structural basis of response regulator dephosphorylation by Rap phosphatases.

    Science.gov (United States)

    Parashar, Vijay; Mirouze, Nicolas; Dubnau, David A; Neiditch, Matthew B

    2011-02-08

    Bacterial Rap family proteins have been most extensively studied in Bacillus subtilis, where they regulate activities including sporulation, genetic competence, antibiotic expression, and the movement of the ICEBs1 transposon. One subset of Rap proteins consists of phosphatases that control B. subtilis and B. anthracis sporulation by dephosphorylating the response regulator Spo0F. The mechanistic basis of Rap phosphatase activity was unknown. Here we present the RapH-Spo0F X-ray crystal structure, which shows that Rap proteins consist of a 3-helix bundle and a tetratricopeptide repeat domain. Extensive biochemical and genetic functional studies reveal the importance of the observed RapH-Spo0F interactions, including the catalytic role of a glutamine in the RapH 3-helix bundle that inserts into the Spo0F active site. We show that in addition to dephosphorylating Spo0F, RapH can antagonize sporulation by sterically blocking phosphoryl transfer to and from Spo0F. Our structure-function analysis of the RapH-Spo0F interaction identified Rap protein residues critical for Spo0F phosphatase activity. This information enabled us to assign Spo0F phosphatase activity to a Rap protein based on sequence alone, which was not previously possible. Finally, as the ultimate test of our newfound understanding of the structural requirements for Rap phosphatase function, a non-phosphatase Rap protein that inhibits the binding of the response regulator ComA to DNA was rationally engineered to dephosphorylate Spo0F. In addition to revealing the mechanistic basis of response regulator dephosphorylation by Rap proteins, our studies support the previously proposed T-loop-Y allostery model of receiver domain regulation that restricts the aromatic "switch" residue to an internal position when the β4-α4 loop adopts an active-site proximal conformation.

  13. Two-Site Comparison of Transpiration by Larrea Tridentata

    Science.gov (United States)

    Cavanaugh, M. L.; Kurc, S. A.; Scott, R. L.; Bryant, R. B.

    2008-12-01

    As a result of landscape changes within the desert southwestern U.S. such as increased grazing, reduced wildfire frequency, and changes in atmospheric conditions, the native creosotebush (Larrea tridentata) has encroached upon historically grass-dominated ecosystems, expanding in range and land cover density. To understand how creosotebush influences the water budget of ecosystems, heat balance sap flow sensors were employed on creosotebush stems at both the Santa Rita Experimental Range (SRER) and Walnut Gulch Experimental Watershed (WGEW). Additionally, both sites are equipped with eddy covariance towers, associated micrometeorological measurements, and profiles of water content reflectometers for soil moisture. The differences found between the two sites, including soil type and precipitation regime, are the basis of the following hypotheses. Firstly, we hypothesize that we will not see transpiration (T) responses following storms less than 5 mm at both sites. Secondly, we hypothesize that at both sites we will see a lagged response of T to large precipitation events, with evaporation being the dominate component in the partitioning of evapotranspiration (ET) for the first two days. Thirdly, we hypothesize that the ratio of plant transpiration to total evapotranspiration (T/ET) will be less at SRER due to the larger amount of bare soil exposed at this site. In this study, we show data from one summer at both sites and show how these relate to different precipitation events and soil moisture reservoirs.

  14. MAPK Cascades in Guard Cell Signal Transduction

    Science.gov (United States)

    Lee, Yuree; Kim, Yun Ju; Kim, Myung-Hee; Kwak, June M.

    2016-01-01

    Guard cells form stomata on the epidermis and continuously respond to endogenous and environmental stimuli to fine-tune the gas exchange and transpirational water loss, processes which involve mitogen-activated protein kinase (MAPK) cascades. MAPKs form three-tiered kinase cascades with MAPK kinases and MAPK kinase kinases, by which signals are transduced to the target proteins. MAPK cascade genes are highly conserved in all eukaryotes, and they play crucial roles in myriad developmental and physiological processes. MAPK cascades function during biotic and abiotic stress responses by linking extracellular signals received by receptors to cytosolic events and gene expression. In this review, we highlight recent findings and insights into MAPK-mediated guard cell signaling, including the specificity of MAPK cascades and the remaining questions. PMID:26904052

  15. MAPK cascades in guard cell signal transduction

    Directory of Open Access Journals (Sweden)

    Yuree eLee

    2016-02-01

    Full Text Available Guard cells form stomata on the epidermis and continuously respond to endogenous and environmental stimuli to fine-tune the gas exchange and transpirational water loss, processes which involve mitogen-activated protein kinase (MAPK cascades. MAPKs form three-tiered kinase cascades with MAPK kinases and MAPK kinase kinases, by which signals are transduced to the target proteins. MAPK cascade genes are highly conserved in all eukaryotes, and they play crucial roles in myriad developmental and physiological processes. MAPK cascades function during biotic and abiotic stress responses by linking extracellular signals received by receptors to cytosolic events and gene expression. In this review, we highlight recent findings and insights into MAPK-mediated guard cell signaling, including the specificity of MAPK cascades and the remaining questions.

  16. Diesel and silica monitoring at two sites following hurricane sandy.

    Science.gov (United States)

    Freund, Alice; Zuckerman, Norman; Luo, Honghong; Hsu, Hsiao-Hsien; Lucchini, Roberto

    2014-01-01

    Following Hurricane Sandy, which hit New York City and New Jersey in October 2012, industrial hygienists from the Mount Sinai and Belleview/New York University occupational medicine clinics conducted monitoring for diesel exhaust and silica in lower Manhattan and Rockaway Peninsula. Average daytime elemental carbon levels at three stations in lower Manhattan on December 4, 2012, ranged from 9 to18 μg/m(3). Sub-micron particle counts at various times on the same day were over 200,000 particles per cubic centimeter on many streets in lower Manhattan. In Rockaway Peninsula on December 12, 2012, all average daytime elemental carbon levels were below a detection limit of approximately 7 μg/m(3). The average daytime crystalline silica dust concentration was below detection at two sites on Rockaway Peninsula, and was 0.015 mg/m(3) quartz where sand was being replaced on the beach. The daily average levels of elemental carbon and airborne particulates that we measured are in the range of levels that have been found to cause respiratory effects in sensitive subpopulations like asthmatic patients after 2 hr of exposure. Control of exposure to diesel exhaust must be considered following natural disasters where diesel-powered equipment is used in cleanup and recovery. Although peak silica exposures were not likely captured in this study, but were reported by a government agency to have exceeded recommended guidelines for at least one cleanup worker, we recommend further study of silica exposures when debris removal operations or traffic create visible levels of suspended dust from soil or sand.

  17. Stress activated MAPKs in plants

    NARCIS (Netherlands)

    Ligterink, W.

    2000-01-01

    Plants are exposed to a wide variety of extracellular stimuli and employ a broad set of signaling pathways to give the appropriate response. M itogen a ctivated p rotein k inases (MAPKs) play an important role in the signal transduction of yeast and animals and increasing

  18. Stress activated MAPKs in plants

    NARCIS (Netherlands)

    Ligterink, J.W.

    2000-01-01

    Plants are exposed to a wide variety of extracellular stimuli and employ a broad set of signaling pathways to give the appropriate response. M itogen a ctivated p rotein k inases (MAPKs) play an important role in

  19. Plant MAPK cascades: Just rapid signaling modules?

    KAUST Repository

    Boudsocq, Marie

    2015-08-27

    © 2015 Taylor & Francis Group, LLC. Abscisic acid (ABA) is a major phytohormone mediating important stress-related processes. We recently unveiled an ABA-activated MAPK signaling module constituted of MAP3K17/18-MKK3-MPK1/2/7/14. Unlike classical rapid MAPK activation, we showed that the activation of the new MAPK module is delayed and relies on the MAP3K protein synthesis. In this addendum, we discuss the role of this original and unexpected activation mechanism of MAPK cascades which suggests that MAPKs can regulate both early and longterm plant stress responses.

  20. Spatial distribution and dose-response relationship for different operation modes in a reaction-diffusion model of the MAPK cascade

    Science.gov (United States)

    Zhao, Qi; Yi, Ming; Liu, Yan

    2011-10-01

    The mitogen-activated protein kinase (MAPK) cascade plays a critical role in the control of cell growth. Deregulation of this pathway contributes to the development of many cancers. To better understand its signal transduction, we constructed a reaction-diffusion model for the MAPK pathway. We modeled the three layers of phosphorylation-dephosphorylation reactions and diffusion processes from the cell membrane to the nucleus. Based on different types of feedback in the MAPK cascade, four operation modes are introduced. For each of the four modes, spatial distributions and dose-response curves of active kinases (i.e. ppMAPK) are explored by numerical simulation. The effects of propagation length, diffusion coefficient and feedback strength on the pathway dynamics are investigated. We found that intrinsic bistability in the MAPK cascade can generate a traveling wave of ppMAPK with constant amplitude when the propagation length is short. ppMAPK in this mode of intrinsic bistability decays more slowly than it does in all other modes as the propagation length increases. Moreover, we examined the global and local responses to Ras-GTP of these four modes, and demonstrated how the shapes of these dose-response curves change as the propagation length increases. Also, we found that larger diffusion constant gives a higher response level on the zero-order regime and makes the ppMAPK profiles flatter under strong Ras-GTP stimulus. Furthermore, we observed that spatial responses of ppMAPK are more sensitive to negative feedback than to positive feedback in the broader signal range. Finally, we showed how oscillatory signals pass through the kinase cascade, and found that high frequency signals are damped faster than low frequency ones.

  1. EhMAPK, the mitogen-activated protein kinase from Entamoeba histolytica is associated with cell survival.

    Science.gov (United States)

    Ghosh, Anupama Sardar; Ray, Doel; Dutta, Suman; Raha, Sanghamitra

    2010-10-08

    Mitogen Activated Protein Kinases (MAPKs) are a class of serine/threonine kinases that regulate a number of different cellular activities including cell proliferation, differentiation, survival and even death. The pathogen Entamoeba histolytica possess a single homologue of a typical MAPK gene (EhMAPK) whose identification was previously reported by us but its functional implications remained unexplored. EhMAPK, the only mitogen-activated protein kinase from the parasitic protist Entamoeba histolytica with Threonine-X-Tyrosine (TXY) phosphorylation motif was cloned, expressed in E. coli and functionally characterized under different stress conditions. The expression profile of EhMAPK at the protein and mRNA level remained similar among untreated, heat shocked and hydrogen peroxide-treated samples in all cases of dose and time. But a significant difference was obtained in the phosphorylation status of the protein in response to different stresses. Heat shock at 43°C or 0.5 mM H(2)O(2) treatment enhanced the phosphorylation status of EhMAPK and augmented the kinase activity of the protein whereas 2.0 mM H(2)O(2) treatment induced dephosphorylation of EhMAPK and loss of kinase activity. 2.0 mM H(2)O(2) treatment reduced parasite viability significantly but heat shock and 0.5 mM H(2)O(2) treatment failed to adversely affect E. histolytica viability. Therefore, a distinct possibility that activation of EhMAPK is associated with stress survival in E. histolytica is seen. Our study also gives a glimpse of the regulatory mechanism of the protein under in vivo conditions. Since the parasite genome lacks any typical homologue of mammalian MEK, the dual specificity kinases which are the upstream activators of MAPK, indications of the existence of some alternate regulatory mechanisms of the EhMAPK activity is perceived. These may include the autophosphorylation activity of the protein itself in combination with some upstream phosphatases which are not yet identified.

  2. EhMAPK, the mitogen-activated protein kinase from Entamoeba histolytica is associated with cell survival.

    Directory of Open Access Journals (Sweden)

    Anupama Sardar Ghosh

    Full Text Available Mitogen Activated Protein Kinases (MAPKs are a class of serine/threonine kinases that regulate a number of different cellular activities including cell proliferation, differentiation, survival and even death. The pathogen Entamoeba histolytica possess a single homologue of a typical MAPK gene (EhMAPK whose identification was previously reported by us but its functional implications remained unexplored. EhMAPK, the only mitogen-activated protein kinase from the parasitic protist Entamoeba histolytica with Threonine-X-Tyrosine (TXY phosphorylation motif was cloned, expressed in E. coli and functionally characterized under different stress conditions. The expression profile of EhMAPK at the protein and mRNA level remained similar among untreated, heat shocked and hydrogen peroxide-treated samples in all cases of dose and time. But a significant difference was obtained in the phosphorylation status of the protein in response to different stresses. Heat shock at 43°C or 0.5 mM H(2O(2 treatment enhanced the phosphorylation status of EhMAPK and augmented the kinase activity of the protein whereas 2.0 mM H(2O(2 treatment induced dephosphorylation of EhMAPK and loss of kinase activity. 2.0 mM H(2O(2 treatment reduced parasite viability significantly but heat shock and 0.5 mM H(2O(2 treatment failed to adversely affect E. histolytica viability. Therefore, a distinct possibility that activation of EhMAPK is associated with stress survival in E. histolytica is seen. Our study also gives a glimpse of the regulatory mechanism of the protein under in vivo conditions. Since the parasite genome lacks any typical homologue of mammalian MEK, the dual specificity kinases which are the upstream activators of MAPK, indications of the existence of some alternate regulatory mechanisms of the EhMAPK activity is perceived. These may include the autophosphorylation activity of the protein itself in combination with some upstream phosphatases which are not yet identified.

  3. TRAIL apoptosis is enhanced by quercetin through Akt dephosphorylation.

    Science.gov (United States)

    Kim, Young-Ho; Lee, Yong J

    2007-03-01

    TNF-related apoptosis-inducing ligand (TRAIL) is a promising cancer therapy that preferentially induces apoptosis in cancer cells. However, many neoplasms are resistant to TRAIL by mechanisms that are poorly understood. Here we demonstrated that human prostate cancer cells, but not normal prostate cells, are dramatically sensitized to TRAIL-induced apoptosis and caspase activation by quercetin. Quercetin, a ubiquitous bioactive plant flavonoid, has been shown to inhibit the proliferation of cancer cells. We have shown that quercetin can potentiate TRAIL-induced apoptotic death. Human prostate adenocarcinoma DU-145 and LNCaP cells were treated with various concentrations of TRAIL (10-200 ng/ml) and/or quercetin (10-200 microM) for 4 h. Quercetin, which caused no cytotoxicity by itself, promoted TRAIL-induced apoptosis. The TRAIL-mediated activation of caspase, and PARP (poly(ADP-ribose) polymerase) cleavage were both enhanced by quercetin. Western blot analysis showed that combined treatment with TRAIL and quercetin did not change the levels of TRAIL receptors (death receptors DR4 and DR5, and DcR2 (decoy receptor 2)) or anti-apoptotic proteins (FLICE-inhibitory protein (FLIP), inhibitor of apoptosis (IAP), and Bcl-2). However, quercetin promoted the dephosphorylation of Akt. Quercetin-induced potent inhibition of Akt phosphorylation. Taken together, the present studies suggest that quercetin enhances TRAIL-induced cytotoxicity by activating caspases and inhibiting phosphorylation of Akt.

  4. Signaling mechanisms and functional roles of cofilin phosphorylation and dephosphorylation.

    Science.gov (United States)

    Mizuno, Kensaku

    2013-02-01

    Cofilin and actin-depolymerizing factor (ADF) are actin-binding proteins that play an essential role in regulating actin filament dynamics and reorganization by stimulating the severance and depolymerization of actin filaments. Cofilin/ADF are inactivated by phosphorylation at the serine residue at position 3 by LIM-kinases (LIMKs) and testicular protein kinases (TESKs) and are reactivated by dephosphorylation by the slingshot (SSH) family of protein phosphatases and chronophin. This review describes recent advances in our understanding of the signaling mechanisms regulating LIMKs and SSHs and the functional roles of cofilin phospho-regulation in cell migration, tumor invasion, mitosis, neuronal development, and synaptic plasticity. Accumulating evidence demonstrates that the phospho-regulation of cofilin/ADF is a key convergence point of cell signaling networks that link extracellular stimuli to actin cytoskeletal dynamics and that spatiotemporal control of cofilin/ADF activity by LIMKs and SSHs plays a crucial role in a diverse array of cellular and physiological processes. Perturbations in the normal control of cofilin/ADF activity underlie many pathological conditions, including cancer metastasis and neurological and cardiovascular disorders.

  5. Dephosphorylation specificities of protein phosphatase for cardiac troponin I, troponin T, and sites within troponin T

    Directory of Open Access Journals (Sweden)

    2006-03-01

    Full Text Available Protein dephosphorylation by protein phosphatase 1 (PP1, acting in concert with protein kinase C (PKC and protein kinase A (PKA, is a pivotal regulatory mechanism of protein phosphorylation. Isolated rat cardiac myofibrils phosphorylated by PKC/PKA and dephosphorylated by PP1 were used in determining dephosphorylation specificities, Ca2+-stimulated Mg2+ATPase activities, and Ca2+ sensitivities. In reconstituted troponin (Tn complex, PP1 displayed distinct substrate specificity in dephosphorylation of TnT preferentially to TnI, in vitro. In situ phosphorylation of cardiomyocytes with calyculin A, a protein phosphatase inhibitor, resulted in an increase in the phosphorylation stiochiometry of TnT (0.3 to 0.5 (67%, TnI (2.6 to 3.6 (38%, and MLC2 (0.4 to 1.7 (325%. These results further confirmed that though MLC2 is the preferred target substrate for protein phosphatase in the thick filament, the Tn complex (TnI and TnT from thin filament and C-protein in the thick filament are also protein phosphatase substrates. Our in vitro dephosphorylation experiments revealed that while PP1 differentially dephosphorylated within TnT at multiple sites, TnI was uniformly dephosphorylated. Phosphopeptide maps from the in vitro experiments show that TnT phosphopeptides at spots 4A and 4B are much more resistant to PP1 dephosphorylation than other TnT phosphopeptides. Mg2+ATPase assays of myofibrils phosphorylated by PKC/PKA and dephosphorylated by PP1 delineated that while PKC and PKA phosphorylation decreased the Ca2+-stimulated Mg2+ATPase activities, dephosphorylation antagonistically restored it. PKC and PKA phosphorylation decreased Ca2+ sensitivity to 3.6 µM and 5.0 µM respectively. However, dephosphorylation restored the Mg2+ATPase activity of PKC (99% and PKA (95%, along with the Ca2+ sensitivities (3.3 µM and 3.0 µM, respectively.

  6. Simvastatin-induced breast cancer cell death and deactivation of PI3K/Akt and MAPK/ERK signalling are reversed by metabolic products of the mevalonate pathway.

    Science.gov (United States)

    Wang, Tingting; Seah, Serena; Loh, Xinyi; Chan, Ching-Wan; Hartman, Mikael; Goh, Boon-Cher; Lee, Soo-Chin

    2016-01-19

    Statins purportedly exert anti-tumoral effects on breast cancer. However, the biologic mechanisms for these actions are not fully elucidated. The aims of this study were 1) to explore the effects of simvastatin on apoptosis, proliferation as well as PI3K/Akt/mTOR and MAPK/ERK pathway in a window-of-opportunity breast cancer trial; 2) to further confirm findings from the clinical trial by functional studies; 3) to explore the regulatory role of mevalonate pathway on the anti-tumoral effects of simvastatin. In clinical samples, simvastatin led to increase in cleaved caspase-3 (p = 0.002) and decreased trend for Ki67 (p = 0.245). Simvastatin markedly suppressed PI3K/Akt/mTOR signalling by activating PTEN (p = 0.005) and by dephosphorylating Akt (p = 0.002) and S6RP (p = 0.033); it also inhibited MAPK/ERK pathway by dephosphorylating c-Raf (p = 0.018) and ERK1/2 (p = 0.002). In ER-positive (MCF-7, T47D) and ER-negative (MDA-MB-231, BT-549) breast cancer cells, simvastatin treatment consistently induced apoptosis and inhibited proliferation by deregulating caspase cascades and cell cycle proteins in a dose dependent manner. Concordantly, simvastatin strongly suppressed PI3K/Akt/mTOR pathway by enhancing PTEN expression and by further sequentially dephosphorylating downstream cascades including Akt, mTOR, p70S6K, S6RP and 4E-BP1. Furthermore, simvastatin significantly inhibited MAPK/ERK pathway by dephosphorylating sequential cascades such as c-Raf, MEK1/2 and ERK1/2. These simvastatin anti-tumoral effects were reversed by metabolic products of the mevalonate pathway, including mevalonate, farnesyl pyrophosphate and geranylgeranyl pyrophosphate. These findings shed light on the biological and potential anti-tumoral effects of simvastatin in breast cancer.

  7. A comparison of forest dynamics at two sites in the Southeastern Ozark Mountains of Missouri

    Science.gov (United States)

    Michael A. Jenkins; Stephen G. Pallardy

    1993-01-01

    Changes in tree species composition and regeneration patterns were studied in 53 permanent vegetation plots located at two sites (Pioneer Forest and University State Forest) in oak-hickory forests of southeastern Missouri where mortality and decline of red oak species have been identified. The two sites also exhibited differing levels of decline and mortality. Between...

  8. Ultrasensitivity in phosphorylation-dephosphorylation cycles with little substrate.

    Directory of Open Access Journals (Sweden)

    Bruno M C Martins

    Full Text Available Cellular decision-making is driven by dynamic behaviours, such as the preparations for sunrise enabled by circadian rhythms and the choice of cell fates enabled by positive feedback. Such behaviours are often built upon ultrasensitive responses where a linear change in input generates a sigmoidal change in output. Phosphorylation-dephosphorylation cycles are one means to generate ultrasensitivity. Using bioinformatics, we show that in vivo levels of kinases and phosphatases frequently exceed the levels of their corresponding substrates in budding yeast. This result is in contrast to the conditions often required by zero-order ultrasensitivity, perhaps the most well known means for how such cycles become ultrasensitive. We therefore introduce a mechanism to generate ultrasensitivity when numbers of enzymes are higher than numbers of substrates. Our model combines distributive and non-distributive actions of the enzymes with two-stage binding and concerted allosteric transitions of the substrate. We use analytical and numerical methods to calculate the Hill number of the response. For a substrate with [Formula: see text] phosphosites, we find an upper bound of the Hill number of [Formula: see text], and so even systems with a single phosphosite can be ultrasensitive. Two-stage binding, where an enzyme must first bind to a binding site on the substrate before it can access the substrate's phosphosites, allows the enzymes to sequester the substrate. Such sequestration combined with competition for each phosphosite provides an intuitive explanation for the sigmoidal shifts in levels of phosphorylated substrate. Additionally, we find cases for which the response is not monotonic, but shows instead a peak at intermediate levels of input. Given its generality, we expect the mechanism described by our model to often underlay decision-making circuits in eukaryotic cells.

  9. Regulation of Hippo pathway transcription factor TEAD by p38 MAPK-induced cytoplasmic translocation.

    Science.gov (United States)

    Lin, Kimberly C; Moroishi, Toshiro; Meng, Zhipeng; Jeong, Han-Sol; Plouffe, Steven W; Sekido, Yoshitaka; Han, Jiahuai; Park, Hyun Woo; Guan, Kun-Liang

    2017-07-28

    The Hippo pathway controls organ size and tissue homeostasis, with deregulation leading to cancer. The core Hippo components in mammals are composed of the upstream serine/threonine kinases Mst1/2, MAPK4Ks and Lats1/2. Inactivation of these upstream kinases leads to dephosphorylation, stabilization, nuclear translocation and thus activation of the major functional transducers of the Hippo pathway, YAP and its paralogue TAZ. YAP/TAZ are transcription co-activators that regulate gene expression primarily through interaction with the TEA domain DNA-binding family of transcription factors (TEAD). The current paradigm for regulation of this pathway centres on phosphorylation-dependent nucleocytoplasmic shuttling of YAP/TAZ through a complex network of upstream components. However, unlike other transcription factors, such as SMAD, NF-κB, NFAT and STAT, the regulation of TEAD nucleocytoplasmic shuttling has been largely overlooked. In the present study, we show that environmental stress promotes TEAD cytoplasmic translocation via p38 MAPK in a Hippo-independent manner. Importantly, stress-induced TEAD inhibition predominates YAP-activating signals and selectively suppresses YAP-driven cancer cell growth. Our data reveal a mechanism governing TEAD nucleocytoplasmic shuttling and show that TEAD localization is a critical determinant of Hippo signalling output.

  10. Protein phosphatase 2A subunit PR70 interacts with pRb and mediates its dephosphorylation.

    Science.gov (United States)

    Magenta, Alessandra; Fasanaro, Pasquale; Romani, Sveva; Di Stefano, Valeria; Capogrossi, Maurizio C; Martelli, Fabio

    2008-01-01

    The retinoblastoma tumor suppressor protein (pRb) regulates cell proliferation and differentiation via phosphorylation-sensitive interactions with specific targets. While the role of cyclin/cyclin-dependent kinase complexes in the modulation of pRb phosphorylation has been extensively studied, relatively little is known about the molecular mechanisms regulating phosphate removal by phosphatases. Protein phosphatase 2A (PP2A) is constituted by a core dimer bearing catalytic activity and one variable B regulatory subunit conferring target specificity and subcellular localization. We previously demonstrated that PP2A core dimer binds pRb and dephosphorylates pRb upon oxidative stress. In the present study, we identified a specific PP2A-B subunit, PR70, that was associated with pRb both in vitro and in vivo. PR70 overexpression caused pRb dephosphorylation; conversely, PR70 knockdown prevented both pRb dephosphorylation and DNA synthesis inhibition induced by oxidative stress. Moreover, we found that intracellular Ca(2+) mobilization was necessary and sufficient to trigger pRb dephosphorylation and PP2A phosphatase activity of PR70 was Ca(2+) induced. These data underline the importance of PR70-Ca(2+) interaction in the signal transduction mechanisms triggered by redox imbalance and leading to pRb dephosphorylation.

  11. mTORC2 modulates feedback regulation of p38 MAPK activity via DUSP10/MKP5 to confer differential responses to PP242 in glioblastoma

    Science.gov (United States)

    Benavides-Serrato, Angelica; Anderson, Lauren; Holmes, Brent; Cloninger, Cheri; Artinian, Nicholas; Bashir, Tariq; Gera, Joseph

    2014-01-01

    Dual-specificity phosphatases (DUSPs) dephosphorylate MAP kinases (MAPKs) resulting in their inactivation. Activation of MAPK signaling leads to enhanced DUSP expression, thus establishing feedback regulation of the MAPK pathway. The DUSPs are subject to regulation at the post-translational level via phosphorylation resulting in alterations of protein stability. Here we report that mTORC2 function leads to stabilization of the p38 MAPK phosphatase, DUSP10, thereby inhibiting p38 activity. We demonstrate that mTORC2 binds DUSP10 and phosphorylates DUSP10 on serine residues 224 and 230. These phosphorylation events block DUSP10 turnover resulting in inactivation of p38 signaling. We further show that insulin-stimulated PI3K/mTORC2 signaling regulates DUSP10 stability and p38 activity. Importantly, knockdown of DUSP10 or ectopic overexpression of nonphosphorylatable or phosphomimetic DUSP10 mutants was sufficient to confer differential mTOR kinase inhibitor responses to GBM cells in vitro and in murine xenografts. Finally, DUSP10 was shown to be overexpressed in a significant number of GBM patients. These data demonstrate the ability of the mTORC2 pathway to exert regulatory effects on the DUSP10/p38 feedback loop to control the cellular effects of mTOR kinase inhibitors in GBM and support the use of DUSP10 expression as a surrogate biomarker to predict responsiveness. PMID:25568665

  12. Dephosphorylation Pathway of D-myo-Inositol 1,4,5-trisphosphate in the Unicellular Green Alga Chlamydomonas eugametos

    NARCIS (Netherlands)

    Klerk, Hans; Himbergen, John A.J. van; Musgrave, Alan; Haastert, Peter J.M. van; Ende, Herman van den

    1994-01-01

    In vitro dephosphorylation of D-myo-inositol 1,4,5-trisphosphate [Ins(l,4,5)P-3] by vegetative cells, gametes and zygotes of the green alga Chlamydomonas eugametos was studied using a soluble cell fraction as enzyme source and labelled Ins(1,4,5)P-3 as substrate. This compound was dephosphorylated y

  13. Two dephosphorylation pathways of inositol 1,4,5-trisphosphate in homogenates of the cellular slime mould Dictyostelium discoideum

    NARCIS (Netherlands)

    Lookeren Campagne, Michiel M. van; Erneux, Cristophe; Eijk, Ronald van; Haastert, Peter J.M. van

    1988-01-01

    Dictyostelium discoideum homogenates contain phosphatase activity which rapidly dephosphorylates Ins(1,4,5)P3 (D-myo-inositol 1,4,5-trisphosphate) to Ins (myo-inositol). When assayed in Mg2+, Ins(1,4,5)P3 is dephosphorylated by the soluble Dictyostelium cell fraction to 20% Ins(1,4)P2 (D-myo-inosito

  14. Analysis list: Mapk8 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Mapk8 Neural,Pluripotent stem cell + mm9 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Mapk...8.1.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Mapk8.5.tsv http://dbarchive.b...iosciencedbc.jp/kyushu-u/mm9/target/Mapk8.10.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Mapk8.N...eural.tsv,http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Mapk8.Pluripotent_s

  15. On the role and fate of LPS-dephosphorylating activity in the rat liver

    NARCIS (Netherlands)

    Tuin, A; Huizinga -van der Vlag, Ali; van Loenen - Weemaes, Anne-miek; Meijer, DKF; Poelstra, K

    2006-01-01

    Gut-derived lipopolysaccharide (LPS) plays a role in the pathogenesis of liver diseases like fibrosis. The enzyme alkaline phosphatase (AP) is present in, among others, the intestinal wall and liver and has been previously shown to dephosphorylate LPS. Therefore, we investigated the effect of LPS on

  16. Dynamin and PTP-PEST cooperatively regulate Pyk2 dephosphorylation in osteoclasts.

    Science.gov (United States)

    Eleniste, Pierre P; Du, Liping; Shivanna, Mahesh; Bruzzaniti, Angela

    2012-05-01

    Bone loss is caused by the dysregulated activity of osteoclasts which degrade the extracellular bone matrix. The tyrosine kinase Pyk2 is highly expressed in osteoclasts, and mice lacking Pyk2 exhibit an increase in bone mass, in part due to impairment of osteoclast function. Pyk2 is activated by phosphorylation at Y402 following integrin activation, but the mechanisms leading to Pyk2 dephosphorylation are poorly understood. In the current study, we examined the mechanism of action of the dynamin GTPase on Pyk2 dephosphorylation. Our studies reveal a novel mechanism for the interaction of Pyk2 with dynamin, which involves the binding of Pyk2's FERM domain with dynamin's plextrin homology domain. In addition, we demonstrate that the dephosphorylation of Pyk2 requires dynamin's GTPase activity and is mediated by the tyrosine phosphatase PTP-PEST. The dephosphorylation of Pyk2 by dynamin and PTP-PEST may be critical for terminating outside-in integrin signaling, and for stabilizing cytoskeletal reorganization during osteoclast bone resorption.

  17. Involvement of GPIIb-IIIa on human platelets in phosphotyrosine-specific dephosphorylation.

    Science.gov (United States)

    Takayama, H; Ezumi, Y; Ichinohe, T; Okuma, M

    1993-07-15

    Washed platelets from either normal donors or patients with thrombasthenia lacking in integrin GPIIb-IIIa were stimulated by thrombin or STA2 with stirring and their kinetics of protein-tyrosine phosphorylation were compared. The early increase in protein-tyrosine phosphorylation on 115 and 75 kDa protein bands was observed within 10 s after stimulation in both normal and thrombasthenic platelets. While both 115 and 75 kDa tyrosine-phosphorylated protein bands were quickly dephosphorylated in normal platelets, thrombin-induced 115 kDa or STA2-induced 115 and 75 kDa protein bands were not dephosphorylated in thrombasthenic platelets. The delay of phosphotyrosine-specific dephosphorylation on those protein bands was observed when thrombin- or STA2-induced aggregation of normal platelets was inhibited by RGDS, an inhibitor of fibrinogen binding to GPIIb-IIIa. These data indicate that fibrinogen binding to GPIIb-IIIa is involved in the regulation of phosphotyrosine-specific dephosphorylation on certain protein bands.

  18. One-site versus two-site phacotrabeculectomy: a prospective randomized study

    Directory of Open Access Journals (Sweden)

    Moschos MM

    2015-08-01

    Full Text Available Marilita M Moschos,1 Irini P Chatziralli,2 Michael Tsatsos3 1First Department of Ophthalmology, University of Athens, 2Second Department of Ophthalmology, Ophthalmiatrion Athinon, Athens, Greece; 3Department of Ophthalmology, Cambridge University Hospital NHS, Cambridge, UK Purpose: The purpose of this study is to compare the efficacy and safety of one-site and two-site combined phacotrabeculectomy with foldable posterior chamber intraocular lens implantation.Methods: Thirty-four patients (41 eyes with glaucoma and cataract were randomly assigned to undergo either a one-site (22 eyes or a two-site (19 eyes combined procedure. One-site approach consisted of a standard superior phacotrabeculectomy with a limbus-based conjunctival flap, while two-site approach consisted of a clear cornea phacoemulsification and a separate superior trabeculectomy with a limbus-based conjunctival flap.Results: Mean follow-up period was 54 months (standard deviation [SD] 2.3. Mean preoperative intraocular pressure (IOP in the one-site group was 21.3 mmHg (SD 2.8 and in the two-site group was 21.8 mmHg (SD 3.0 (P>0.1. Mean postoperative IOP significantly decreased in both groups compared to the preoperative level and was 15.6 mmHg (SD 3.5 in the one-site group and 14.9 mmHg (SD 2.7 in the two-site group. Three months later, the difference between the two groups was not statistically significant (P=0.058. The one-site group required significantly more medications than the two-site group (P=0.03. Best-corrected visual acuity (BCVA improved similarly in both groups, but there was less postoperative (induced astigmatism in the two-site group in a marginal statistical level (P=0.058. Intra- and postoperative complications were comparable in the two groups.Conclusion: Both techniques yielded similar results concerning final BCVA and IOP reduction. However, the two-site group had less induced astigmatism and a better postoperative IOP control with less required

  19. The crystal structure of phosphorylated MAPK13 reveals common structural features and differences in p38 MAPK family activation.

    Science.gov (United States)

    Yurtsever, Zeynep; Scheaffer, Suzanne M; Romero, Arthur G; Holtzman, Michael J; Brett, Tom J

    2015-04-01

    The p38 MAP kinases (p38 MAPKs) represent an important family centrally involved in mediating extracellular signaling. Recent studies indicate that family members such as MAPK13 (p38δ) display a selective cellular and tissue expression and are therefore involved in specific diseases. Detailed structural studies of all p38 MAPK family members are crucial for the design of specific inhibitors. In order to facilitate such ventures, the structure of MAPK13 was determined in both the inactive (unphosphorylated; MAPK13) and active (dual phosphorylated; MAPK13/pTpY) forms. Here, the first preparation, crystallization and structure determination of MAPK13/pTpY are presented and the structure is compared with the previously reported structure of MAPK13 in order to facilitate studies for structure-based drug design. A comprehensive analysis of inactive versus active structures for the p38 MAPK family is also presented. It is found that MAPK13 undergoes a larger interlobe configurational rearrangement upon activation compared with MAPK14. Surprisingly, the analysis of activated p38 MAPK structures (MAP12/pTpY, MAPK13/pTpY and MAPK14/pTpY) reveals that, despite a high degree of sequence similarity, different side chains are used to coordinate the phosphorylated residues. There are also differences in the rearrangement of the hinge region that occur in MAPK14 compared with MAPK13 which would affect inhibitor binding. A thorough examination of all of the active (phosphorylated) and inactive (unphosphorylated) p38 MAPK family member structures was performed to reveal a common structural basis of activation for the p38 MAP kinase family and to identify structural differences that may be exploited for developing family member-specific inhibitors.

  20. Signatures of many-body localization in the dynamics of two-site entanglement

    Science.gov (United States)

    Iemini, Fernando; Russomanno, Angelo; Rossini, Davide; Scardicchio, Antonello; Fazio, Rosario

    2016-12-01

    We are able to detect clear signatures of dephasing—a distinct trait of many-body localization (MBL)—via the dynamics of two-site entanglement, quantified through the concurrence. Using the protocol implemented by M. Schreiber et al. [Science 349, 842 (2015), 10.1126/science.aaa7432], we show that in the MBL phase the average two-site entanglement decays in time as a power law, while in the Anderson localized phase it tends to a plateau. The power-law exponent is not universal and displays a clear dependence on the interaction strength. This behavior is also qualitatively different from the ergodic phase, where the two-site entanglement decays exponentially. All the results are obtained by means of time-dependent density matrix renormalization-group simulations and further corroborated by analytical calculations on an effective model. Two-site entanglement has been measured in cold atoms: our analysis paves the way for the first direct experimental test of many-body dephasing in the MBL phase.

  1. Exact solution of a generalized two-sites Bose-Hubbard model

    CERN Document Server

    Filho, Gilberto N Santos

    2016-01-01

    I introduce a new parametrization of a bosonic Lax operator for the algebraic Bethe ansatz method with the $gl(2)$-invariant $R$-matrix and use it to present the exact solution of a generalized two-sites Bose-Hubbard model with asymmetric tunnelling. In the no interaction limit I show that the Bethe ansatz equations can be written as a $S^{N-1}$ sphere, where $N$ is the total number of atoms in the condensate.

  2. Bethe states for the two-site Bose–Hubbard model: A binomial approach

    Directory of Open Access Journals (Sweden)

    Gilberto Santos

    2015-06-01

    Full Text Available We calculate explicitly the Bethe vectors states by the algebraic Bethe ansatz method with the gl(2-invariant R-matrix for the two-site Bose–Hubbard model. Using a binomial expansion of the n-th power of a sum of two operators we get and solve a recursion equation. We calculate the scalar product and the norm of the Bethe vectors states. The form factors of the imbalance current operator are also computed.

  3. Two-site Hubbard molecule with a spinless electron-positron pair

    KAUST Repository

    Cossu, Fabrizio

    2012-12-19

    We determine the eigenvalues of the two-site Hubbard molecule with one electron and one positron to describe the characteristics of electron-positron interactions in solids. While the effect of hopping is, in general, opposite to the effect of on-site interaction, we find a complex scenario for the electron-positron pair with a non-vanishing potential drop. We give analytical solutions and discuss the combined effects of the model parameters.

  4. Dynamin and PTP-PEST cooperatively regulate Pyk2 dephosphorylation in osteoclasts

    OpenAIRE

    2012-01-01

    Bone loss is caused by the dysregulated activity of osteoclasts which degrade the extracellular bone matrix. The tyrosine kinase Pyk2 is highly expressed in osteoclasts, and mice lacking Pyk2 exhibit an increase in bone mass, in part due to impairment of osteoclast function. Pyk2 is activated by phosphorylation at Y402 following integrin activation, but the mechanisms leading to Pyk2 dephosphorylation are poorly understood. In the current study, we examined the mechanism of action of the dyna...

  5. Nur1 dephosphorylation confers positive feedback to mitotic exit phosphatase activation in budding yeast.

    Directory of Open Access Journals (Sweden)

    Molly Godfrey

    2015-01-01

    Full Text Available Substrate dephosphorylation by the cyclin-dependent kinase (Cdk-opposing phosphatase, Cdc14, is vital for many events during budding yeast mitotic exit. Cdc14 is sequestered in the nucleolus through inhibitory binding to Net1, from which it is released in anaphase following Net1 phosphorylation. Initial Net1 phosphorylation depends on Cdk itself, in conjunction with proteins of the Cdc14 Early Anaphase Release (FEAR network. Later on, the Mitotic Exit Network (MEN signaling cascade maintains Cdc14 release. An important unresolved question is how Cdc14 activity can increase in early anaphase, while Cdk activity, that is required for Net1 phosphorylation, decreases and the MEN is not yet active. Here we show that the nuclear rim protein Nur1 interacts with Net1 and, in its Cdk phosphorylated form, inhibits Cdc14 release. Nur1 is dephosphorylated by Cdc14 in early anaphase, relieving the inhibition and promoting further Cdc14 release. Nur1 dephosphorylation thus describes a positive feedback loop in Cdc14 phosphatase activation during mitotic exit, required for faithful chromosome segregation and completion of the cell division cycle.

  6. Parathyroid hormone receptor recycling: role of receptor dephosphorylation and beta-arrestin.

    Science.gov (United States)

    Chauvin, Stephanie; Bencsik, Margaret; Bambino, Tom; Nissenson, Robert A

    2002-12-01

    The recovery of PTH receptor (PTHR) function after acute homologous receptor desensitization and down-regulation in bone and kidney cells has been attributed to receptor recycling. To determine the role of receptor dephosphorylation in PTHR recycling, we performed morphological and functional assays on human embryonic kidney 293 cells stably expressing wild-type (wt) or mutant PTHRs. Confocal microscopy and ligand binding assays revealed that the wt PTHR is rapidly recycled back to the plasma membrane after removal of the agonist. Receptors that were engineered to either lack the sites of phosphorylation or to resemble constitutively phosphorylated receptors were able to recycle back to the plasma membrane with the same kinetics as the wt PTHR. The PTHR was found to be dephosphorylated by an enzyme apparently distinct from protein phosphatases 1 or 2A. The PTHR and beta-arrestin-2-green fluorescent protein (GFP) were found to stably colocalize during PTHR internalization, whereas after agonist removal and during receptor recycling, the colocalization slowly disappeared. Experiments using phosphorylation-deficient PTHRs and a dominant-negative form of beta-arrestin showed that beta-arrestin does not regulate the efficiency of PTHR recycling. These studies indicate that, unlike many G protein-coupled receptors, PTHR recycling does not require receptor dephosphorylation or its dissociation from beta-arrestin.

  7. Cyclic nucleotide dependent dephosphorylation of regulator of G-protein signaling 18 in human platelets.

    LENUS (Irish Health Repository)

    Gegenbauer, Kristina

    2013-11-01

    Regulator of G-protein signaling 18 (RGS18) is a GTPase-activating protein that turns off Gq signaling in platelets. RGS18 is regulated by binding to the adaptor protein 14-3-3 via phosphorylated serine residues S49 and S218 on RGS18. In this study we confirm that thrombin, thromboxane A2, or ADP stimulate the interaction of RGS18 and 14-3-3 by increasing the phosphorylation of S49. Cyclic AMP- and cyclic GMP-dependent kinases (PKA, PKG) inhibit the interaction of RGS18 and 14-3-3 by phosphorylating S216. To understand the effect of S216 phosphorylation we studied the phosphorylation kinetics of S49, S216, and S218 using Phos-tag gels and phosphorylation site-specific antibodies in transfected cells and in platelets. Cyclic nucleotide-induced detachment of 14-3-3 from RGS18 coincides initially with double phosphorylation of S216 and S218. This is followed by dephosphorylation of S49 and S218. Dephosphorylation of S49 and S218 might be mediated by protein phosphatase 1 (PP1) which is linked to RGS18 by the regulatory subunit PPP1R9B (spinophilin). We conclude that PKA and PKG induced S216 phosphorylation triggers the dephosphorylation of the 14-3-3 binding sites of RGS18 in platelets.

  8. PP2A-mediated dephosphorylation of p107 plays a critical role in chondrocyte cell cycle arrest by FGF.

    Directory of Open Access Journals (Sweden)

    Victoria Kolupaeva

    Full Text Available FGF signaling inhibits chondrocyte proliferation, a cell type-specific response that is the basis for several genetic skeletal disorders caused by activating FGFR mutations. This phenomenon requires the function of the p107 and p130 members of the Rb protein family, and p107 dephosphorylation is one of the earliest distinguishing events in FGF-induced growth arrest. To determine whether p107 dephoshorylation played a critical role in the chondrocyte response to FGF, we sought to counteract this process by overexpressing in RCS chondrocytes the cyclin D1/cdk4 kinase complex. CyclinD/cdk4-expressing RCS cells became resistant to FGF-induced p107 dephosphorylation and growth arrest, and maintained significantly high levels of cyclin E/cdk2 activity and of phosphorylated p130 at later times of FGF treatment. We explored the involvement of a phosphatase in p107 dephosphorylation. Expression of the SV40 small T-Ag, which inhibits the activity of the PP2A phosphatase, or knockdown of the expression of the PP2A catalytic subunit by RNA interference prevented p107 dephosphorylation and FGF-induced growth arrest of RCS cells. Furthermore, an association between p107 and PP2A was induced by FGF treatment. Our data show that p107 dephosphorylation is a key event in FGF-induced cell cycle arrest and indicate that in chondrocytes FGF activates the PP2A phosphatase to promote p107 dephosphorylation.

  9. Successful recruitment methods in the community for a two-site clinical trial.

    Science.gov (United States)

    Fairbanks, Eileen; Shah, Shivani; Wilde, Mary H; McDonald, Margaret V; Brasch, Judith; McMahon, James M

    2014-11-01

    Effective screening and recruitment are essential to the success of randomized clinical trials. This report is to describe key screening and recruitment strategies in a two site randomized clinical trial (RCT) conducted in community settings with a vulnerable chronically ill population and to suggest valuable approaches when planning trials. Differences between sites in a complex study with two considerably different environments (academic versus home care) and their participant pools presented challenges which required different screening and recruitment methods. A high level of communication between sites, creative problem solving and the ability to be flexible when problems were encountered were needed for successful screening and recruitment.

  10. Two-site adsolubilization model of incorporation of fluoromonomers into fluorosurfactants formed on cotton fabric.

    Science.gov (United States)

    Hanumansetty, Srinivas; O'Rear, Edgar

    2014-04-01

    The adsorption of surfactants and adsolubilization of organic compounds on knit cotton fabric are fundamentally important in admicellar polymerization to impart characteristics like water repellency, stain resistance, and flame retardancy. The main objective of this research is to study adsorption and adsolubilization of fluororsurfactants and fluoromonomers used to obtain water repellency characteristics. Adsorption of nonionic (fluoroaliphatic amine oxide) and cationic (fluoroaliphatic quaternary ammonium surfactant) fluororsurfactants at the interface of cotton is investigated with and without fluoroacrylate monomers. A two-site adsolubilization model was used to predict the aggregation number of fluorosurfactant.

  11. MAPK Assays in Arabidopsis MAMP-PRR Signal Transduction.

    Science.gov (United States)

    Chung, Hoo Sun; Sheen, Jen

    2017-01-01

    Activation of MAPK (Mitogen-Activated Protein Kinase) cascades after MAMP (Microbe-Associated Molecular Pattern) perception through PRR (Pattern Recognition Receptor) is one of the first conserved responses when plants encounter microbial organisms. Phosphorylation of various cellular factors in the MAMP-PRR pathway by MAPK cascades is critical for broad-spectrum plant innate immunity. Measurement of MAPK activation and identification of MAPK phosphorylation targets in the MAMP-PRR signal transduction pathway are essential to understand how plants reprogram their cellular processes to cope with unfavorable microbial attack. Here, we describe detailed protocols of three assays measuring MAPK activity after MAMP perception: (1) immune-blotting analysis with anti-phospho ERK1/2 antibody; (2) in-gel kinase assay using a general substrate myelin basic protein (MBP); (3) an in vitro kinase assay to evaluate phosphorylation of MAPK substrate candidates during MAMP-PRR signaling based on a protoplast expression system.

  12. MAPK Phosphatase 5 Expression Induced by Influenza and Other RNA Virus Infection Negatively Regulates IRF3 Activation and Type I Interferon Response

    Directory of Open Access Journals (Sweden)

    Sharmy J. James

    2015-03-01

    Full Text Available The type I interferon system is essential for antiviral immune response and is a primary target of viral immune evasion strategies. Here, we show that virus infection induces the expression of MAPK phosphatase 5 (MKP5, a dual-specificity phosphatase (DUSP, in host cells. Mice deficient in MKP5 were resistant to H1N1 influenza infection, which is associated with increased IRF3 activation and type I interferon expression in comparison with WT mice. Increased type I interferon responses were also observed in MKP5-deficient cells and animals upon other RNA virus infection, including vesicular stomatitis virus and sendai virus. These observations were attributed to the ability of MKP5 to interact with and dephosphorylate IRF3. Our study reveals a critical function of a DUSP in negative regulation of IRF3 activity and demonstrates a mechanism by which influenza and other RNA viruses inhibit type I interferon response in the host through MKP5.

  13. MFHAS1 suppresses TLR4 signaling pathway via induction of PP2A C subunit cytoplasm translocation and inhibition of c-Jun dephosphorylation at Thr239.

    Science.gov (United States)

    Shi, Qiqing; Xiong, Bo; Zhong, Jing; Wang, Huihui; Ma, Duan; Miao, Changhong

    2017-08-01

    TLR4, an important Toll-like receptor in innate immunity, can be activated by LPS and induce proinflammatory cytokines to resist invasion of pathogenic microorganism, but excessive inflammation can trigger tissue injury. Many genes negatively regulate TLR4 signaling pathway. Recent studies found that malignant fibrous histiocytoma amplified sequence 1 (MFHAS1) suppressed the expression of cytokine IL6 in Raw264.7 cells stimulated by LPS, but the mechanisms remained unclear. This study investigated the role of MFHAS1 in TLR4 signaling pathway and the possible mechanisms implicated. The results indicated that the expression of MFHAS1 was significantly increased in cells stimulated with LPS. Up-regulation of MFHAS1 effectively suppressed inflammatory cytokine expression in cells exposed to LPS, whereas down-regulation of MFHAS1 markedly increased inflammatory cytokines expression. Co-immunoprecipitation, pull-down and immunofluorescence tests demonstrated that MFHAS1 combined with the B and C subunits of PP2A and induced cytoplasm translocation of the C subunit, leading to decrease dephosphorylation of c-Jun at Thr239 and increase degradation of c-Jun. Reduction of c-Jun protein results in decreased AP-1 activity, which is independent of inhibition of JNK or p38MAPK phosphorylation. Taken together, these results indicate that MFHAS1 suppresses TLR4 signaling pathway through induction of PP2A C subunit cytoplasm translocation and subsequent c-Jun degradation, leading finally to decrease AP-1 activity and cytokines expression. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Cloning and Preliminary Function Analysis of DcMAPK1 and DcMAPK2 in Carnation%香石竹DcMAPK1和DcMAPK2的克隆及功能初步分析

    Institute of Scientific and Technical Information of China (English)

    马悦悦; 肖志娜; 汤娜; 刘娟旭; 余义勋

    2015-01-01

    分离了香石竹(Dianthus caryophyllus)两个促分裂原活化蛋白质激酶(mitogen-ac-tivated protein kinase,MAPK)基因cDNA全长,分别命名为DcMAPK1和DcMAPK2,其中DcMAPK1与拟南芥AtMAPK6高度同源.DcMAPK1在香石竹花瓣衰老过程表达增加,乙烯处理显著促进其表达,瞬时抑制DcMAPK1后切花瓶插寿命显著延长.而DcMAPK2在花瓣衰老过程中组成型表达,乙烯处理对其表达无显著影响,瞬时抑制DcMAPK2对切花瓶插寿命无显著影响.

  15. Phytochemical differences between Calia secundiflora (Leguminosae) growing at two sites in Mexico.

    Science.gov (United States)

    Zavala-Chávez, Fernando; García-Mateos, Rosario; Soto-Hernández, Marcos; Kite, Geofrey

    2006-01-01

    The ecology and quinolizidine alkaloid chemistry of Calia secundiflora (Ortega) Yakovlev growing at two sites in Mexico were compared. At one site (Hidalgo) the vegetation was dominated by Flourensia resinosa and C. secundiflora, at the other site (Queretaro) C. secundiflora and Dodanaea viscosa were dominant. The Hidalgo site had shallower soils with less organic matter, N, P, and CaCO3. Seeds of C. secundiflora from each site accumulated a similar range of quinolizidine alkaloids, but the profile of alkaloids in the leaves and roots were different. The leaves and roots of plants at Hidalgo accumulated a similar range of alkaloids to the seeds with cytisine and/or N-methylcytisine being most abundant, whereas at Queretaro the leaves and roots accumulated lupinine, with other alkaloids being relatively minor constituents. The latter profile has not been reported previously for C. secundiflora.

  16. Analytical diagonalization study of a two-orbital Hubbard model on a two-site molecule

    Energy Technology Data Exchange (ETDEWEB)

    Amendola, Maria Emilia, E-mail: canio@sa.infn.it [Dipartimento di Matematica, Università di Salerno, I-84084 Fisciano (Italy); Romano, Alfonso [CNR-SPIN, I-84084 Fisciano (Italy); Dipartimento di Fisica “E. R. Caianiello”, Università di Salerno, I-84084 Fisciano (Italy); Noce, Canio, E-mail: canio@sa.infn.it [CNR-SPIN, I-84084 Fisciano (Italy); Dipartimento di Fisica “E. R. Caianiello”, Università di Salerno, I-84084 Fisciano (Italy)

    2015-12-15

    We present the exact solution of a two-orbital Hubbard model on a two-site molecule for arbitrary electron filling and arbitrary interaction couplings. The knowledge of the many-particle spectrum, determined via a diagonalization procedure performed by fully taking into account the symmetry properties of the model, has been used to investigate the temperature dependence of charge, spin and orbital response functions as well as of the intra- and inter-orbital on-site occupations. We point out that this study may allow easy access to many interesting features of the model and may serve as a reference tool for various numerical or perturbation methods dealing with complex correlated electron models defined on a lattice, in particular in the case in which strong local interactions dominate over kinetic effects.

  17. Establishment of a Two-site ELISA for Detection of Chlamydia Trachomatis

    Institute of Scientific and Technical Information of China (English)

    WANG Huj(王慧); XIONG Ljkuan(熊礼宽); LI Ying(李影)

    2002-01-01

    Objective:To establish a rapid and simple assay for the diagnosis of Chlamydia trachonatis (CT) infection.Methods BALB/c mice were immunized with SDS-PAGE purified major outer membrane protein (MOMP) from CT and the monoclonal antibodies were obtained subsequently.Two-site ELISA was developed to detect CT infection.Results: The established assay was able to detect as low as 1.248ug/ml MOMP with interrun and inrun CV 6.9% and 3.1% respectively. 94% (34/36) of culture-positive samples were found to be positive in the current examination,indicating the high sensitivity of this assay.Conclusion: The assay is applicable for clinical diagnosis of CT infection.

  18. Two-site kinetic modeling of bacteriophages transport through columns of saturated dune sand.

    Science.gov (United States)

    Schijven, Jack F; Hassanizadeh, S Majid; de Bruin, Ria H A M

    2002-08-01

    Breakthrough curves, on a semi-log scale, from tests in porous media with block-input of viruses, bacteria, protozoa and colloidal particles often exhibit a typical skewness: a rather slowly rising limb and a smooth transition of a declining limb to a very long tail. One-site kinetic models fail to fit the rising and declining limbs together with the tail satisfactorily. Inclusion of an equilibrium adsorption site does not seem to improve simulation results. This was encountered in the simulation of breakthrough curves from a recent field study on the removal of bacteriophages MS2 and PRD1 by passage through dune sand. In the present study, results of laboratory experiments for the study of this issue are presented. Breakthrough curves of salt and bacteriophages MS2, PRDI, and phiX174 in 1 D column experiments have been measured. One- and two-site kinetic models have been applied to fit and predict breakthrough curves from column experiments. The two-site model fitted all breakthrough curves very satisfactorily, accounting for the skewness of the rising limb as well as for the smooth transition of the declining limb to the tail of the breakthrough curve. The one-site model does not follow the curvature of the breakthrough tail, leading to an overestimation of the inactivation rate coefficient for attached viruses. Interaction with kinetic site 1 is characterized by relatively fast attachment and slow detachment, whereas attachment to and detachment from kinetic site 2 is fast. Inactivation of viruses and interaction with kinetic site 2 provide only a minor contribution to removal. Virus removal is mainly determined by the attachment to site 1. Bacteriophage phiX174 attached more than MS2 and PRD1, which can be explained by the greater electrostatic repulsion that MS2 and PRD1 experience compared to the less negatively charged phiX174.

  19. The roles of MAPKs in disease

    Institute of Scientific and Technical Information of China (English)

    Michael C Lawrence; Svetlana Earnest; Wei Chen; Melanie H Cobb; Arif Jivan; Chunli Shao; Lingling Duan; Daryl Goad; Elma Zaganjor; Jihan Osbome; Kathleen McGlynn; Steve Stippec

    2008-01-01

    MAP kinases transduce signals that are involved in a multitude of cellular pathways and functions in response to a variety of ligands and cell stimuli. Aberrant or inappropriate functions of MAPKs have now been identified in diseases ranging from cancer to inflammatory disease to obesity and diabetes. In many cell types, the MAPKs ERK1/2 are linked to cell proliferation. ERK1/2 are thought to play a role in some cancers, because mutations in Ras and B-Raf, which can activate the ERK1/2 cascade, are found in many human tumors. Abnormal ERK1/2 signaling has also been found in polycystic kidney disease, and serious developmental disorders such as cardio-facio-cutaneous syndrome arise from mutations in components of the ERK1/2 cascade. ERK1/2 are essential in well-differentiated cells and have been linked to long-term potentiation in neurons and in maintenance of epithelial polarity. Additionally, ERK1/2 are important for insulin gene transcription in pancreatic beta cells, which produce insulin in response to increases in circulating glucose to permit efficient glucose utilization and storage in the organism. Nutrients and hormones that induce or repress insulin secretion activate and/or inhibit ERK1/2 in a manner that reflects the secretory demand on beta cells. Disturbances in this and other regulatory pathways may result in the contribution of ERK1/2 to the etiology of certain human disorders.

  20. Obesity does not Lead to Imbalance Between Myocardial Phospholamban Phosphorylation and Dephosphorylation

    Directory of Open Access Journals (Sweden)

    Paula Paccielli Freire

    2014-07-01

    Full Text Available Background: The activation of the beta-adrenergic system promotes G protein stimulation that, via cyclic adenosine monophosphate (cAMP, alters the structure of protein kinase A (PKA and leads to phospholamban (PLB phosphorylation. This protein participates in the system that controls intracellular calcium in muscle cells, and it is the primary regulator of sarcoplasmic reticulum calcium pump activity. In obesity, the beta-adrenergic system is activated by the influence of increased leptin, therefore, resulting in higher myocardial phospholamban phosphorylation via cAMP-PKA. Objective: To investigate the involvement of proteins which regulate the degree of PLB phosphorylation due to beta-adrenergic activation in obesity. In the present study, we hypothesized that there is an imbalance between phospholamban phosphorylation and dephosphorylation, with prevalence of protein phosphorylation. Methods: Male Wistar rats were randomly distributed into two groups: control (n = 14, fed with normocaloric diet; and obese (n = 13, fed with a cycle of four unsaturated high-fat diets. Obesity was determined by the adiposity index, and protein expressions of phosphatase 1 (PP-1, PKA, PLB, phosphorylated phospholamban at serine16 (PPLB-Ser16 were assessed by Western blot. Results: Obesity caused glucose intolerance, hyperinsulinemia, hypertriglyceridemia, hyperleptinemia and did not alter the protein expression of PKA, PP-1, PLB, PPLB-Ser16. Conclusion: Obesity does not promote an imbalance between myocardial PLB phosphorylation and dephosphorylation via beta-adrenergic system.

  1. Dephosphorylated parafibromin is a transcriptional coactivator of the Wnt/Hedgehog/Notch pathways.

    Science.gov (United States)

    Kikuchi, Ippei; Takahashi-Kanemitsu, Atsushi; Sakiyama, Natsuki; Tang, Chao; Tang, Pei-Jung; Noda, Saori; Nakao, Kazuki; Kassai, Hidetoshi; Sato, Toshiro; Aiba, Atsu; Hatakeyama, Masanori

    2016-09-21

    Evolutionally conserved Wnt, Hedgehog (Hh) and Notch morphogen pathways play essential roles in the development, homeostasis and pathogenesis of multicellular organisms. Nevertheless, mechanisms that intracellularly coordinate these signal inputs remain poorly understood. Here we found that parafibromin, a component of the PAF complex, competitively interacts with β-catenin and Gli1, thereby potentiating transactivation of Wnt- and Hh-target genes in a mutually exclusive manner. Parafibromin also binds to the Notch intracellular domain (NICD), enabling concerted activation of Wnt- and Notch-target genes. The transcriptional platform function of parafibromin is potentiated by tyrosine dephosphorylation, mediated by SHP2 phosphatase, while it is attenuated by tyrosine phosphorylation, mediated by PTK6 kinase. Consequently, acute loss of parafibromin in mice disorganizes the normal epithelial architecture of the intestine, which requires coordinated activation/inactivation of Wnt, Hh and/or Notch signalling. Parafibromin integrates and converts signals conveyed by these morphogen pathways into appropriate transcriptional outputs in a tyrosine phosphorylation/dephosphorylation-regulated manner.

  2. Obesity does not Lead to Imbalance Between Myocardial Phospholamban Phosphorylation and Dephosphorylation

    Energy Technology Data Exchange (ETDEWEB)

    Freire, Paula Paccielli, E-mail: freirepp@hotmail.com; Alves, Carlos Augusto Barnabe; Deus, Adriana Fernandes de [Departamento de Clínica Médica - Faculdade de Medicina de Botucatu - Universidade Estadual Paulista, Botucatu, SP (Brazil); Leopoldo, Ana Paula Lima; Leopoldo, André Soares [Centro de Educação Física e Desportos - Universidade Federal do Espírito Santo, Vitória, ES (Brazil); Silva, Danielle Cristina Tomaz da; Tomasi, Loreta Casquel de; Campos, Dijon Henrique Salomé; Cicogna, Antonio Carlos [Departamento de Clínica Médica - Faculdade de Medicina de Botucatu - Universidade Estadual Paulista, Botucatu, SP (Brazil)

    2014-07-15

    The activation of the beta-adrenergic system promotes G protein stimulation that, via cyclic adenosine monophosphate (cAMP), alters the structure of protein kinase A (PKA) and leads to phospholamban (PLB) phosphorylation. This protein participates in the system that controls intracellular calcium in muscle cells, and it is the primary regulator of sarcoplasmic reticulum calcium pump activity. In obesity, the beta-adrenergic system is activated by the influence of increased leptin, therefore, resulting in higher myocardial phospholamban phosphorylation via cAMP-PKA. To investigate the involvement of proteins which regulate the degree of PLB phosphorylation due to beta-adrenergic activation in obesity. In the present study, we hypothesized that there is an imbalance between phospholamban phosphorylation and dephosphorylation, with prevalence of protein phosphorylation. Male Wistar rats were randomly distributed into two groups: control (n = 14), fed with normocaloric diet; and obese (n = 13), fed with a cycle of four unsaturated high-fat diets. Obesity was determined by the adiposity index, and protein expressions of phosphatase 1 (PP-1), PKA, PLB, phosphorylated phospholamban at serine16 (PPLB-Ser16) were assessed by Western blot. Obesity caused glucose intolerance, hyperinsulinemia, hypertriglyceridemia, hyperleptinemia and did not alter the protein expression of PKA, PP-1, PLB, PPLB-Ser16. Obesity does not promote an imbalance between myocardial PLB phosphorylation and dephosphorylation via beta-adrenergic system.

  3. PHLPP1 regulates contact inhibition by dephosphorylating Mst1 at the inhibitory site

    Energy Technology Data Exchange (ETDEWEB)

    Jung, Sujin; Kang, Jeong Gu [Targeted Gene Regulation Research Center, KRIBB, 125 Gwahak-ro, Yuseong-gu, Deajeon (Korea, Republic of); Lee, Ju Hee [Targeted Gene Regulation Research Center, KRIBB, 125 Gwahak-ro, Yuseong-gu, Deajeon (Korea, Republic of); Department of Biomolecular Science, University of Science and Technology, 217 Gajeong-ro, Yuseong-gu, Daejeon (Korea, Republic of); Song, Kyoung Jin [Targeted Gene Regulation Research Center, KRIBB, 125 Gwahak-ro, Yuseong-gu, Deajeon (Korea, Republic of); Ko, Jeong-Heon [Targeted Gene Regulation Research Center, KRIBB, 125 Gwahak-ro, Yuseong-gu, Deajeon (Korea, Republic of); Department of Biomolecular Science, University of Science and Technology, 217 Gajeong-ro, Yuseong-gu, Daejeon (Korea, Republic of); Kim, Yong-Sam, E-mail: omsys1@kribb.re.kr [Targeted Gene Regulation Research Center, KRIBB, 125 Gwahak-ro, Yuseong-gu, Deajeon (Korea, Republic of); Department of Biomolecular Science, University of Science and Technology, 217 Gajeong-ro, Yuseong-gu, Daejeon (Korea, Republic of)

    2014-01-24

    Highlights: • PHLPP1 regulates contact inhibition by dephosphorylating Mst1 at Thr{sup 387}. • Overexpression of PHLPP1 sensitizes contact inhibition. • Tumor cells with suppressed PHLPP1 expression are refractory to apoptosis and highly proliferative. • Loss or down-regulation of PHLPP1 may drive tumor development and progression. - Abstract: Contact inhibition has been largely elusive despite that a loss of contact inhibition is a critical event for cancer development and progression. Here, we report that PHLPP1 is a binding protein for Mst1 and it modulates the Hippo pathway by dephosphorylating Mst1 at the inhibitory Thr{sup 387} of Mst1. Yap1 was localized predominantly in the nucleus but marginally in the cytoplasm in HeLa cells under sparse conditions, whereas the functional protein was more directed to sequestration in the cytoplasm under dense environments. Furthermore, loss of PHLPP1 resulted in a failure of the apoptotic control. It is interesting that down-regulated expression of PHLPP1 appears to mimic the loss of contact inhibition, a hallmark of cancer.

  4. Yersinia virulence factor YopM induces sustained RSK activation by interfering with dephosphorylation.

    Directory of Open Access Journals (Sweden)

    Moritz Hentschke

    Full Text Available BACKGROUND: Pathogenic yersiniae inject several effector proteins (Yops into host cells, which subverts immune functions and enables the bacteria to survive within the host organism. YopM, whose deletion in enteropathogenic yersiniae results in a dramatic loss of virulence, has previously been shown to form a complex with and activate the multifunctional kinases PKN2 and RSK1 in transfected cells. METHODOLOGY/PRINCIPAL FINDINGS: In a near physiological approach with double-affinity-tagged YopM being translocated into the macrophage cell line J774A.1 via the natural type three secretion system of Yersinia we verified the interaction of YopM with PKN2 and RSK1 and detected association with additional PKN and RSK isoforms. In transfected and infected cells YopM induced sustained phosphorylation of RSK at its activation sites serine-380 and serine-221 even in the absence of signalling from its upstream kinase ERK1/2, suggesting inhibition of dephosphorylation. ATP-depletion and in vitro assays using purified components directly confirmed that YopM shields RSK isoforms from phosphatase activity towards serines 380 and 221. CONCLUSIONS/SIGNIFICANCE: Our study suggests that during Yersinia infection YopM induces sustained activation of RSK by blocking dephosphorylation of its activatory phosphorylation sites. This may represent a novel mode of action of a bacterial virulence factor.

  5. PTPRT regulates the interaction of Syntaxin-binding protein 1 with Syntaxin 1 through dephosphorylation of specific tyrosine residue

    Energy Technology Data Exchange (ETDEWEB)

    Lim, So-Hee; Moon, Jeonghee [Biomedical Proteomics Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon 305-806 (Korea, Republic of); Lee, Myungkyu [Bionanotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon 305-806 (Korea, Republic of); Lee, Jae-Ran, E-mail: leejr@kribb.re.kr [Biomedical Proteomics Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon 305-806 (Korea, Republic of)

    2013-09-13

    Highlights: •PTPRT is a brain-specific, expressed, protein tyrosine phosphatase. •PTPRT regulated the interaction of Syntaxin-binding protein 1 with Syntaxin 1. •PTPRT dephosphorylated the specific tyrosine residue of Syntaxin-binding protein 1. •Dephosphorylation of Syntaxin-binding protein 1 enhanced the interaction with Syntaxin 1. •PTPRT appears to regulate the fusion of synaptic vesicle through dephosphorylation. -- Abstract: PTPRT (protein tyrosine phosphatase receptor T), a brain-specific tyrosine phosphatase, has been found to regulate synaptic formation and development of hippocampal neurons, but its regulation mechanism is not yet fully understood. Here, Syntaxin-binding protein 1, a key component of synaptic vesicle fusion machinery, was identified as a possible interaction partner and an endogenous substrate of PTPRT. PTPRT interacted with Syntaxin-binding protein 1 in rat synaptosome, and co-localized with Syntaxin-binding protein 1 in cultured hippocampal neurons. PTPRT dephosphorylated tyrosine 145 located around the linker between domain 1 and 2 of Syntaxin-binding protein 1. Syntaxin-binding protein 1 directly binds to Syntaxin 1, a t-SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) protein, and plays a role as catalysts of SNARE complex formation. Syntaxin-binding protein 1 mutant mimicking non-phosphorylation (Y145F) enhanced the interaction with Syntaxin 1 compared to wild type, and therefore, dephosphorylation of Syntaxin-binding protein 1 appeared to be important for SNARE-complex formation. In conclusion, PTPRT could regulate the interaction of Syntaxin-binding protein 1 with Syntaxin 1, and as a result, the synaptic vesicle fusion appeared to be controlled through dephosphorylation of Syntaxin-binding protein 1.

  6. Protein phosphatase PphA from Synechocystis sp. PCC 6803: the physiological framework of PII-P dephosphorylation.

    Science.gov (United States)

    Kloft, Nicole; Rasch, Grit; Forchhammer, Karl

    2005-04-01

    The phosphorylated signal transduction protein P(II) (P(II)-P) in the cyanobacterium Synechocystis sp. strain PCC 6803 is dephosphorylated by PphA, a protein phosphatase of the 2C family (PP2C). In this study, the physiological conditions of P(II)-P dephosphorylation were investigated with respect to the in vivo specificity of P(II)-P towards PphA and the cellular abundance of PphA in cells growing under different nitrogen regimes. Furthermore, the consequences of impaired P(II)-P dephosphorylation with respect to short-term inhibition of glutamine synthetase (GS) were studied. With a contribution of approximately 15 % of total Mn(2+)-dependent p-nitrophenyl phosphate hydrolysis activity, PphA has only a minor impact on the total PP2C activity in Synechocystis extracts. Nevertheless, residual P(II)-P dephosphorylation in PphA-deficient cells could only be observed after prolonged incubation in the presence of ammonium. The abundance of PphA correlates with the phosphorylation state of P(II) under nitrogen-replete conditions and is specifically enhanced by nitrite. Regulation of pphA expression operates at the post-transcriptional level. In the presence of nitrate/nitrite, PphA is present in molar excess over P(II)-P, enabling the cells to rapidly dephosphorylate P(II)-P in response to changing environmental conditions. A PphA-deficient mutant is not impaired in short-term inhibition of GS activity following ammonium treatment. Down-regulation of GS occurs by induction of gif genes (encoding GS inactivating factors 7 and 17), which is controlled by NtcA-mediated gene repression. Thus, impaired P(II)-P dephosphorylation does not affect ammonium-prompted inactivation of NtcA.

  7. Blockade of p38 MAPK inhibits chronic allograft vasculopathy.

    Science.gov (United States)

    Ollinger, Robert; Thomas, Michael; Kogler, Pamela; Hermann, Martin; Weiss, Helmut; Mark, Walter; Bilban, Martin; Troppmair, Jakob; Bach, Fritz H; Margreiter, Raimund

    2008-01-27

    Long-term survival after solid-organ transplantation is hampered by chronic changes in the arteries of the grafts, called chronic allograft vasculopathy (CAV). The lesions consist mainly of proliferating vascular smooth muscle cells that cause narrowing of the vessels; these lesions can develop within a few months. There is no effective treatment to prevent CAV. We previously noted that the pharmacological inhibition of p38 mitogen-activated protein kinase (MAPK) suppresses the proliferation of vascular smooth muscle cells. We hypothesized that in vivo inhibition of p38 MAPK in mice bearing allogeneic aortic allografts would prevent CAV. We here report that blockade of p38 MAPK, a signaling molecule involved in cell division, apoptosis, and cell death, markedly suppresses CAV. Given recent data indicating that inhibition of p38 MAPK is a promising approach for the treatment of autoimmune diseases plus our present findings, p38 MAPK blockade for CAV seems a reasonable approach to consider for clinical application.

  8. Regulation of cross-talk in yeast MAPK signaling pathways.

    Science.gov (United States)

    Saito, Haruo

    2010-12-01

    MAP kinase (MAPK) modules are conserved three-kinase cascades that serve central roles in intracellular signal transduction in eukaryotic cells. MAPK pathways of different inputs and outputs use overlapping sets of signaling components. In yeast, for example, three MAPK pathways (pheromone response, filamentous growth response, and osmostress adaptation) all use the same Ste11 MAPK kinase kinase (MAPKKK). How undesirable leakage of signal, or cross-talk, is prevented between these pathways has been a subject of intensive study. This review discusses recent findings from yeast that indicate that there is no single mechanism, but that a combination of four general strategies (docking interactions, scaffold proteins, cross-pathway inhibition, and kinetic insulation) are utilized for the prevention of cross-talk between any two MAPK modules.

  9. Cytosolic purine 5'-nucleotidases of rat liver and human red blood cells: regulatory properties and role in AMP dephosphorylation.

    Science.gov (United States)

    Van den Berghe, G; Bontemps, F; Vincent, M F

    1988-01-01

    Of the various species of cellular 5'-nucleotidases, membranous, lysosomal and cytosolic, only the latter are likely to play a role in the physiologic dephosphorylation of the 5'-nucleoside monophosphates present in the cytoplasm. The necessity to preserve cellular ATP renders a strict control of the dephosphorylation as well as of the deamination of AMP mandatory, because both nucleotides are maintained in equilibrium by adenylate kinase. Our studies of cytosolic purine 5'-nucleotidases purified from rat liver and from human erythrocytes, reviewed in this presentation, have shown that both display complex kinetic properties. Both enzymes have markedly higher affinities for IMP and for GMP than for AMP. In addition, they are stimulated by nucleoside triphosphates, among them ATP and GTP, and inhibited by Pi. The erythrocytic purine 5'-nucleotidase is also stimulated by glycerate 2,3-bisphosphate. It could thus be expected that under conditions of ATP and GTP breakdown, particularly when accompanied by an increase in Pi, the dephosphorylation of AMP would be curtailed. To verify this hypothesis, experiments were performed with isolated rat hepatocytes and with human red blood cells. The rate of dephosphorylation of AMP was measured by following time-wise the production of adenosine in the presence of coformycin (or deoxycoformycin) and 5-iodotubercidin. The coformycins inhibit the deamination of adenosine into inosine by adenosine deaminase, and 5-iodotubercidin inhibits the recycling of adenosine into AMP by adenosine kinase. Upon induction of ATP catabolism by the addition of fructose to isolated rat hepatocytes, the dephosphorylation of AMP was nearly completely suppressed. In accordance with these results, the activity of the rat liver cytosolic 5'-nucleotidase, assayed in the presence of concentrations of substrate and effectors mimicking those measured in intact cells following the addition of fructose, was decreased as compared to control conditions. In

  10. IODP Expedition 322 Drills Two Sites to Document Inputs to The Nankai Trough Subduction Zone

    Directory of Open Access Journals (Sweden)

    Yu’suke Kubo

    2010-09-01

    Full Text Available Ocean Drilling Program were to sample and log the incoming sedimentary strata and uppermost igneous basement of the Shikoku Basin, seaward of the Nankai Trough (southwestern Japan. Characterization of these subduction inputs is one piece of the overall science plan for the Nankai Trough Seismogenic Zone Experiment. Before we can assess how various material properties evolve down the dip of the plate interface, and potentially change the fault’s behavior from stable sliding to seismogenic slip, we must determine the initial pre-subduction conditions. Two sites were drilled seaward of the trench to demonstrate how facies characterand sedimentation rates responded to bathymetric architecture. Site C0011 is located on the northwest flank of a prominent basement high (Kashinosaki Knoll, and Site C0012 is located near the crest of the seamount. Even though significant gaps remain in the coring record, and attempts to recover wireline logs at Site C0012 failed, correlations can be made between stratigraphic units at the two sites.Sedimentation rates slowed down throughout the condensed section above the basement high, but the seafloor relief was never high enough during the basin’s evolution to prevent the accumulation of sandy turbidites near the crest of the seamount. We discovered a new stratigraphic unit, the middle Shikoku Basin facies, which is typified by late Miocene volcaniclastic turbidites. The sediment-basalt contact was recovered intact at Site C0012, giving a minimumbasement age of 18.9 Ma. Samples of interstitial water show a familiar freshening trend with depth at Site C0011, but chlorinity values at Site C0012 increase above the values for seawater toward the basement contact. The geochemical trends at Site C0012 are probably a response to hydration reactions in the volcaniclastic sediment and diffusional exchange with seawater-like fluid in the upper igneous basement. These data are important because they finallyestablish an

  11. PM2.5 Indoor Air Quality at Two Sites in London Ontario - A Case Study

    Science.gov (United States)

    Mates, A. V.; Xu, X.; Gilliland, J.; Maltby, M. J.

    2010-12-01

    Studies have shown an association between ambient fine particulate matter (PM2.5) and health impacts, particularly for the elderly and children. As part of a larger study, PM2.5 concentrations were measured using the DustTrak (Model 8520, TSI, St. Paul, MN, USA) at two schools within the city of London, Ontario (Canada). Site A was in a suburban environment while site B was in an urban setting. Monitoring took place for 3 weeks during winter (Feb. 16 - Mar. 8) and 3 weeks during spring (May 05 - 25) of 2010. The winter campaign monitored indoor PM2.5 only, while the spring campaign added outdoor monitors (PM2.5 and CO2) after the first week. Ten min. concentrations were used for analysis. Indoor measurements were split into weekday and weekend. For the same time interval, the outdoor concentrations showed mean values of 18 and 21 μg/m3 for sites A & B, respectively, both under the Canada Wide Standard of 30 μg/m3. Measurements at the two sites showed good associations (R^2 = 0.44), during the spring campaign. This indicates that the outdoor PM2.5 had similar sources. For indoor concentrations, Site B showed a significantly different mean concentration 5 times higher compared to site A during the winter ( 8.1 vs. 1.5 μg/m3 ) and 3 times higher (11.9 vs. 3.7 μg/m3) during the spring campaign. Since the outdoor concentrations were similar the large difference in indoor concentrations could be attributed to the following factors: site B being an older building, and the different physical characteristics between the two sites. The spring measurements showed an increase of 50% from weekday to weekend for site A and 22% for site B. The higher level of PM2.5 during weekends is possibly due to the infiltration of outdoor air while the ventilation/filtration system is shut off. During the winter campaign, Site A showed a 14% higher concentration during weekdays compared to weekends while site B weekend concentrations were 17% higher compared to weekday, which will be

  12. Comprehensive analysis of phosphorylation sites in Tensin1 reveals regulation by p38MAPK.

    Science.gov (United States)

    Hall, Emily H; Balsbaugh, Jeremy L; Rose, Kristie L; Shabanowitz, Jeffrey; Hunt, Donald F; Brautigan, David L

    2010-12-01

    Tensin1 is the archetype of a family of focal adhesion proteins. Tensin1 has a phosphotyrosine binding domain that binds the cytoplasmic tail of β-integrin, a Src homology 2 domain that binds focal adhesion kinase, p130Cas, and the RhoGAP called deleted in liver cancer-1, a phosphatase and tensin homology domain that binds protein phosphatase-1α and other regions that bind F-actin. The association between tensin1 and these partners affects cell polarization, migration, and invasion. In this study we analyzed the phosphorylation of human S-tag-tensin1 expressed in HEK293 cells by mass spectrometry. Peptides covering >90% of the sequence initially revealed 50 phosphorylated serine/phosphorylated threonine (pSer/pThr) but no phosphorylated tyrosine (pTyr) sites. Addition of peroxyvanadate to cells to inhibit protein tyrosine phosphatases exposed 10 pTyr sites and addition of calyculin A to cells to inhibit protein phosphatases type 1 and 2A gave a total of 62 pSer/pThr sites. We also characterized two sites modified by O-linked N-acetylglucosamine. Tensin1 F302A, which does not bind protein phosphatase-1, showed > twofold enhanced phosphorylation of seven sites. The majority of pSer/pThr have adjacent proline (Pro) residues and we show endogenous p38 mitogen activated protein kinase (MAPK) associated with and phosphorylated tensin1 in an in vitro kinase assay. Recombinant p38α MAPK also phosphorylated S-tag-tensin1, resulting in decreased binding with deleted in liver cancer-1. Activation of p38 MAPK in cells by sorbitol-induced hyperosmotic stress increased phosphorylation of S-tag-tensin1, which reduced binding to deleted in liver cancer-1 and increased binding to endogenous pTyr proteins, including p130Cas and focal adhesion kinase. These data demonstrate that tensin1 is extensively phosphorylated on Ser/Thr residues in cells and phosphorylation by p38 MAPK regulates the specificity of the tensin1 Src homology 2 domain for binding to different proteins. Tensin1

  13. Structure of an epiphytic hydroid community on Cystoseira at two sites of different wave exposure

    Directory of Open Access Journals (Sweden)

    Anuschka Faucci

    2000-12-01

    Full Text Available Epiphytism is a strategy by which opportunistic species such as hydroids, escape the intense levels of competition in marine hard bottom communities. Species of the macroalgae Cystoseira have a seasonal turnover of the frond, and we hypothesise that epiphytic hydroids colonising such an unstable substrate might show some degree of specialisation. Here the first systematic study on hydroid-Cystoseira communities is presented. In particular, the seasonal and spatial distribution of epiphytic hydroids on three species of Cystoseira at two sites of different wave exposure at Porto Cesareo (Ionian Sea/Italy were investigated. Thirty-two hydroid species were recorded which are well known from other substrates and thus are not specific to Cystoseira; dominant species were all thecates. The influence of biological factors such as competition and the structure and abundance of the host, seem to have little influence on the hydroid community. The factors of greatest influence were mostly abiotic: sedimentation rate, nutrient levels, temperature and most especially water movement. The importance of water movement was evident in the higher colonisation of algal stems, higher hydroid frequency, larger colonies, reduced colony height, species composition, and distribution on the stems at the wave-exposed site.

  14. Pseudospin representation of the two-site Anderson-Hubbard model

    Science.gov (United States)

    Wortis, Rachel; Kennett, Malcolm

    The state of an Anderson localized system can be described in terms of the occupation of a set of single-particle wave functions which are localized in space. When interactions are added, single-particle wave functions are no longer well defined, so what is a useful description of the state of a many-body localized system and what about it is localized? Given that any system with Hilbert-space dimension 2N may be described by an Ising-type Hamiltonian, it has been proposed that in a fully many-body localized system the Ising pseudospins in this representation may be chosen to be local. Actually constructing these spins is non-trivial. While a number of approaches have been proposed, few explicit examples exist and almost all work has been on spin systems. Here we present the Hamiltonian of a two-site Hubbard model with disorder and nearest-neighbor interactions written in terms of pseudospins, and we explore the form of these pseudospins and their evolution as a function of hopping amplitude. Supported by NSERC of Canada.

  15. Differential arsenic binding in the sediments of two sites in Chile's lower Loa River basin.

    Science.gov (United States)

    Bugueño, Manuel P; Acevedo, Sara E; Bonilla, Carlos A; Pizarro, Gonzalo E; Pasten, Pablo A

    2014-01-01

    Fluvial sediments from two lower Loa River basin sites in northern Chile were compared in order to probe the effects of vegetation and organic matter (OM) on As accumulation in fluvial environments. The two sites were the Sloman dam, which lacks macrophytes and has a low OM content (2.4%) in sediments, and the Quillagua Oasis, which is 23 km downstream from the Sloman site and has a higher OM (6.2%) in sediments and abundant aquatic plant life. The Quillagua site had preferential As enrichment with a co-occurrence pattern that differed from that of the Sloman site, which had a lower As concentration (1528 vs. 262 mg/kg d.w., respectively). At the Quillagua site, As concentration was strongly correlated with Mn and OM (r = 0.91 and 0.85, respectively); while at the Sloman site, As concentration in sediments was significantly correlated with Ca and Sr (r = 0.63 and 0.54, respectively). Sequential extraction analyses showed that the Sloman site had higher percentage of easily exchangeable As within the surface sediment (12%, 45 mg/kg d.w.) compared with the Quillagua site (3%, 40 mg/kg d.w.). These contrasting results suggest that both vegetation and OM control the immobilization and accumulation of As in the arid Loa River basin.

  16. Flowering phenology of tree rhododendron along an elevation gradient in two sites in the Eastern Himalayas

    Science.gov (United States)

    Ranjitkar, Sailesh; Luedeling, Eike; Shrestha, Krishna Kumar; Guan, Kaiyun; Xu, Jianchu

    2013-03-01

    Flowering phenology of tree rhododendron ( Rhododendron arboreum Sm.) was monitored in situ along elevation gradients in two distinct ecological settings. Observations were carried out in Gaoligong Nature Reserve (GNR) in China and in the Kanchenjunga Conservation Area (KCA) in Nepal. Using the crown density method, flowering events of the selected species were recorded. Flowering duration and synchrony were determined within each site and along the elevation gradient in each study area. Our observations showed high synchrony throughout the elevation gradient, especially for peak flowering. Mean 15-day soil temperature, soil parameters (soil moisture, nitrogen, organic matter and pH), age of the observed trees, and site characteristics (litter cover, canopy cover, inclination) were related to mean initial and peak flowering dates using partial least squares regression (PLS). Results differed between the two sites, but winter temperature was the most important variable affecting the regression model for both initial flowering and peak flowering at both sites. After temperature, soil moisture was the most important variable for explaining initial flowering dates. The distribution of tree rhododendron indicates that it is able to grow in a wide range of habitats with different environmental conditions. The recent trend of rising winter-spring temperature and the detected bloom-advancing effect of high temperatures during this period suggest that tree rhododendron might expand its distributional range in response to global warming.

  17. Identification of phosphatase that dephosphorylates xylose in the glycosaminoglycan-protein linkage region of proteoglycans.

    Science.gov (United States)

    Koike, Toshiyasu; Izumikawa, Tomomi; Sato, Ban; Kitagawa, Hiroshi

    2014-03-07

    Recently, we demonstrated that FAM20B is a kinase that phosphorylates the xylose (Xyl) residue in the glycosaminoglycan-protein linkage region of proteoglycans. The phosphorylation of Xyl residues by FAM20B enhances the formation of the linkage region. Rapid dephosphorylation is probably induced just after synthesis of the linker and just before polymerization initiates. Indeed, in vitro chondroitin or heparan sulfate polymerization does not occur when the Xyl residue of the tetrasaccharide linkage region is phosphorylated. However, the enzyme responsible for the dephosphorylation of Xyl remains unknown. Here, we identified a novel protein that dephosphorylates the Xyl residue and designated it 2-phosphoxylose phosphatase. The phosphatase efficiently removed the phosphate from the phosphorylated trisaccharide, Galβ1-3Galβ1-4Xyl(2-O-phosphate), but not from phosphorylated tetrasaccharide, GlcUAβ1-3Galβ1-3Galβ1-4Xyl(2-O-phosphate). Additionally, RNA interference-mediated inhibition of 2-phosphoxylose phosphatase resulted in increased amounts of GlcNAcα1-4GlcUAβ1-3Galβ1-3Galβ1-4Xyl(2-O-phosphate), Galβ1-3Galβ1-4Xyl(2-O-phosphate), and Galβ1-4Xyl(2-O-phosphate) in the cells. Gel filtration analysis of the glycosaminoglycan chains synthesized in the knockdown cells revealed that these cells produced decreased amounts of glycosaminoglycan chains and that the chains had similar lengths to those in the mock-transfected cells. Transcripts encoding this phosphatase were ubiquitously, but differentially, expressed in human tissues. Moreover, the phosphatase localized to the Golgi and interacted with the glucuronyltransferase-I involved in the completion of the glycosaminoglycan-protein linkage region. Based on these findings, we conclude that transient phosphorylation of the Xyl residue in the glycosaminoglycan-protein linkage region controls the formation of glycosaminoglycan chains of proteoglycans.

  18. Specific dephosphorylation by phosphatases 1 and 2A of a nuclear protein structurally and immunologically related to nucleolin

    DEFF Research Database (Denmark)

    Schneider, H R; Mieskes, G; Issinger, O G

    1989-01-01

    A new nuclear substrate (N-60) for phosphatase 1 and 2Ac has been described. In contrast to nucleolin (C23), to which it is structurally and immunologically related, N-60 becomes dephosphorylated to 51% and 41% by phosphatases 1 and 2Ac, respectively, within 10 min. Incubation up to 20 min led...

  19. Determination of the Mixing Layer Height Over two Sites, Using Pilot Balloons During the MILAGRO Campaign

    Science.gov (United States)

    Wohrnschimmel, H.; Alonso, A. L.; Ángeles, F.; Sosa, G.; Varela, J.; Cárdenas, B.

    2007-12-01

    Among the mechanisms that affect air quality there is a variety of meteorological processes. An important process in this context are the changes in the mixing layer height during a day and over the year. The mixing layer height is the portion of the atmosphere close to the surface layer where air pollutants get diluted, without leaving this layer. Therefore, it is important to describe the variations in the height of the mixing layer, i.e. the vertical dilution of air pollution, since this is a process mitigating naturally the impact of emissions. There exist different methods to obtain information on the mixing layer height, among them radio soundings, the application of vertical wind profilers, and launching pilot balloons. In this study, pilot balloons have been used simultaneously over two sites of the Mexico City Metropolitan Area during the MILAGRO campaign in March 2006. The objective was to determine the vertical wind profiles and derive information on the mixing layer height. Daily, four pilot balloons were launched, at 9:00, 12:00, 15:00, and 18:00 hours, over Tenango del Aire (a rural area in the Southeast of Mexico City), and over Ciudad Universitaria, in the Southern metropolitan area. At some occasions, night time measurements have been carried out at 21:00 and 24:00. A variability of the diurnal evolution of the mixing layer was observed along March, which could be related to surface temperature. The diurnal evolution showed a sudden growth of the mixing layer between 9:00 and 12:00 hours. Data intercomparisons were carried out for pilot balloons versus radio soundings during a few days at a third site, Tula, in the North of Mexico City. Both intercomparisons showed that pilot balloons are an effective method to obtain information about the development of the mixing layer.

  20. Direct measurement of human plasma corticotropin-releasing hormone by two-site immunoradiometric assay

    Energy Technology Data Exchange (ETDEWEB)

    Linton, E.A.; McLean, C.; Nieuwenhuyzen Kruseman, A.C.; Tilders, F.J.; Van der Veen, E.A.; Lowry, P.J.

    1987-05-01

    A ''two-site'' immunoradiometric assay (IRMA) which allows the direct estimation of human CRH (hCRH) in plasma is described. Using this IRMA, basal levels of CRH in normal subjects ranged from 2-28 pg/mL (mean, 15 +/- 7 (+/- SD) pg/mL; n = 58). Values in men and women were similar. Plasma CRH values within this range were also found in patients with Cushing's syndrome, Addison's disease, and Nelson's syndrome, with no correlation between plasma CRH and ACTH levels in these patients. Elevated plasma CRH levels were found in pregnant women near term (1462 +/- 752 (+/- SD) pg/mL; n = 55), and the dilution curve of this CRH-like immunoreactivity paralleled the IRMA standard curve. After its immunoadsorption from maternal plasma, this CRH-like material eluted on reverse phase high performance liquid chromatography with a retention time identical to that of synthetic CRH and had equipotent bioactivity with the synthetic peptide in the perfused anterior pituitary cell bioassay. Circulating CRH was not detected in Wistar rats, even after adrenalectomy and subsequent ether stress. Synthetic hCRH was degraded by fresh human plasma relatively slowly; 65% of added CRH remained after 1 h of incubation at 37 C. Degradation was inhibited by heat treatment (54 C; 1 h), cold treatment (4 C; 4 h), or freezing and thawing. Loss of synthetic rat CRH occurred more rapidly when fresh rat plasma was used; only 20% of added CRH remained under the same conditions. The inability to measure CRH in peripheral rat plasma may be due to the presence of active CRH-degrading enzymes which fragment the CRH molecule into forms not recognized by the CRH IRMA.

  1. Soil micromorphology, geochemistry and microbiology at two sites on James Ross Island, Maritime Antarctica

    Science.gov (United States)

    Meier, Lars A.; Krauze, Patryk; Prater, Isabel; Scholten, Thomas; Wagner, Dirk; Kühn, Peter; Mueller, Carsten W.

    2017-04-01

    Referring to the fundamental question in ecosystem research, how biotic and abiotic processes interact, only a few studies exist for polar regions that integrate microbiological and soil scientific studies . Soils comprise the complex structure and environment that fosters water storage and nutrient cycling determined by its unique chemical, physical and biological properties with respect to the specific climate and parent material. In the extreme environment of Antarctica, soil biological processes are primarily controlled by microbial communities (Bacteria, Archaea and Fungi), and thus microbiota may also determine soils chemical and physical properties in a landscape lacking higher plants at an average air temperature below 0°C. James Ross Island, Maritime Antarctica, offers a pristine laboratory and an exceptional opportunity to study pedogenesis without the influence of vascular plants and burrowing animals. We analysed micromorphological features, chemical and microbiological measures at two sites on James Ross Island (Brandy Bay and St. Martha Cove) with similar substrates (mostly fine-grained calcareous sandstones and siltstones of the Alpha Member of the Santa Martha Formation with varying amounts of conglomerates and mudstones) at similar topographic positions (small plateaus at similar elevation (80m a.s.l.)). The sites represent luv- and leeward conditions with respect to the main southwesterly winds. The climate on James Ross Island is to be described as semi-arid polar-continental, which is in clear contrast to the Southern Shetlands (e.g. King George Island) north of the Antarctic Peninsula. We will present first results of soil physical (bulk density, soil moisture and grains size distribution), pedochemical (SOC, total N and S, pH, CECeff, and pedogenic oxides) micromorphological and microbial analyses (Microbial DNA content, microbial abundances).

  2. Temporal variation of fine particle mass at two sites in Mexico City

    Energy Technology Data Exchange (ETDEWEB)

    Serrano, P.; Allen, G. [Harvard School of Public Health, Boston, MA (United States); Castillejos, M. [Univ. Autonoma Metropolitana, Xochimilco (Mexico); Gold, D.; Speizer, F. [Brigham and Women`s Hospital, Boston, MA (United States); Hernandez, M. [Inst. Nacional de Salud Publica, Cuernavaca (Mexico); Hayes, C.; McDonnell, W. [Environmental Protection Agency, Research Triangle Park, NC (United States)

    1994-12-31

    Simultaneous sampling of fine mass (PM{sub 2.5}, using an integrated 24 hour gravimetric method) and the particle scattering extinction coefficient (b{sub sp}, using a heated integrating nephelometer) were used to estimate continuous fine particle concentration at two sites in Mexico City. Linear regression analysis of the 24 h averages of b{sub sp} and the PM{sub 2.5} integrated samples was done on a seasonal basis. The coefficients of determination (R{sup 2}) between these methods ranged from 0.84 to 0.90 for the different sampling periods. These data are the first attempt to describe the diurnal variation of fine mass in Mexico City. Distinct and different diurnal patterns were observed for both sites. For the site located near an industrialized area, a sharp peak occurred between 0700 and 0900 hours and a second smaller but broader peak occurred late at night. This site is characterized by the presence of primary pollutants, with PM{sub 10} annual mean concentrations exceeding 150 {micro}g {center_dot} m{sup {minus}3}. The second bite is located in a residential area down wind of the industrialized area, and is characterized by the presence of secondary pollutants with much lower PM{sub 10} concentrations (annual mean of under 50 {micro}g {center_dot} m{sup {minus}3}). The diurnal fine mass pattern at this site had a broad peak between 0900 and 1200 hours. On individual days, fine mass was sometimes highly correlated with ozone.

  3. The pattern of coral reef development at two sites in the Seychelles

    Science.gov (United States)

    Collier, J. S.; Humber, S. R.

    2004-12-01

    Extensive drilling and dating of modern reef complexes worldwide have shown them to be just thin veneers of Holocene material deposited on older Pleistocene limestone. Where the Pleistocene substrate has been imaged seismically it is often characterised by rough karst topography that is thought to be the result of exposure to subaerial weathering during sea-level low stands. The importance of this antecedent karst topography relative to organic growth processes in controlling present day reef morphology has been much debated. We present high-resolution side-scan backscatter and 2D bathymetric survey results from two sites in the Seychelles (western Indian Ocean). These sites are relatively protected and reef morphology is not modified by tropical storms. The sonogram mosaics reveal areas of uniform backscatter intensity, together with regions of coarse image texture and isolated objects. Based on image texture and gross morphology, the mosaics have been classified into two categories of sediment (fine and coarse sand), seagrass beds, and four categories of reef (low relief reefs, patches, pinnacles, high relief continuous reef). The interpretation was ground truthed by independent diver observations and sediment grain size analysis. The pattern of reef development in these two leeward reefs is complex, with no shore-parallel zoning. The total spatial coverage of the reef classes is only 15% of the total surveyed area. Within these mapped reef areas, actual hard coral cover is estimated to be < 50%, so the actual hard-ground coverage is relatively small. Within the various reef types, no simple relationship between morphological parameters such as spatial area, perimeter water depth and height was found. We speculate that the dominant control on reef development is antecedent granite or karst limestone topography.

  4. PERFORMANCE OF 28-YEAR-OLD PROVENANCES OF LIQUIDAMBAR STYRACIFLUA AT TWO SITES IN WESTERN KENYA

    Directory of Open Access Journals (Sweden)

    Joram M. E. Mbinga

    2015-03-01

    Full Text Available The phenotypic variation in growth of ten 28-year-old Liquidambar styraciflua provenances was studied at two sites, Lugari and Kakamega in Western Kenya. The trial at each site was established in a Randomized Block Design with the ten provenances replicated in four blocks. Each block had ten square plots corresponding to the ten provenances and each plot had thirty six trees. Analysis of growth data gave results which showed a significant difference (P<0.01 in survival, and growth parameters of tree height, and diameter at breast height (Dbh among provenances at both sites. Ranking based on height growth of the provenances showed the best provenance at Lugari site was Finca Las Victoria-Guatemala, with mean height and Dbh of 35.8 m and 37.1 cm respectively, while the leading provenance at Kakamega site was Tactic Coban, Guatemala, with a mean height and Dbh of 28.7 m and 26.8 cm respectively. The provenance with highest mean survival at Lugari was Finca Las Victoria-Guatemala with a value of 29.2%, while at Kakamega the best provenance was Franklin, Virginia–USA, with a value of 72.2%. Provenance by site interaction was significant as shown by the difference in performance of provenances between the sites. A comparison of the result from the best performing provenances with the growth of the most commonly grown Cupressus lusitanica species in similar sites in Kenya indicates the high potential of L. styraciflua as a commercial plantation species in medium altitude sites in Kenya.

  5. Specificity determinants in phosphoinositide dephosphorylation: crystal structure of an archetypal inositol polyphosphate 5-phosphatase.

    Science.gov (United States)

    Tsujishita, Y; Guo, S; Stolz, L E; York, J D; Hurley, J H

    2001-05-04

    Inositol polyphosphate 5-phosphatases are central to intracellular processes ranging from membrane trafficking to Ca(2+) signaling, and defects in this activity result in the human disease Lowe syndrome. The 1.8 resolution structure of the inositol polyphosphate 5-phosphatase domain of SPsynaptojanin bound to Ca(2+) and inositol (1,4)-bisphosphate reveals a fold and an active site His and Asp pair resembling those of several Mg(2+)-dependent nucleases. Additional loops mediate specific inositol polyphosphate contacts. The 4-phosphate of inositol (1,4)-bisphosphate is misoriented by 4.6 compared to the reactive geometry observed in the apurinic/apyrimidinic endonuclease 1, explaining the dephosphorylation site selectivity of the 5-phosphatases. Based on the structure, a series of mutants are described that exhibit altered substrate specificity providing general determinants for substrate recognition.

  6. Interaction of thiamin diphosphate with phosphorylated and dephosphorylated mammalian pyruvate dehydrogenase complex.

    Science.gov (United States)

    Liu, Xiaoqing; Bisswanger, Hans

    2005-01-01

    Kinetic and binding studies were carried out on substrate and cofactor interaction with the pyruvate dehydrogenase complex from bovine heart. Fluoropyruvate and pyruvamide, previously described as irreversible and allosteric inhibitors, respectively, are strong competitive inhibitors with respect to pyruvate. Binding of thiamin diphosphate was used to study differences between the active dephosphorylated and inactive phosphorylated enzyme states by spectroscopic methods. The change in both the intrinsic tryptophan fluorescence and the fluorescence of the 6-bromoacetyl-2-dimethylaminonaphthalene-labelled enzyme complex produced on addition of the cofactor showed similar binding behaviour for both enzyme forms, with slightly higher affinity for the phosphorylated form. Changes in the CD spectrum, especially the negative Cotton effect at 330 nm as a function of cofactor concentration, both in the absence and presence of pyruvate, also revealed no drastic differences between the two enzyme forms. Thus, inactivation of the enzyme activity of the pyruvate dehydrogenase complex is not caused by impeding the binding of substrate or cofactor.

  7. Lanthanide-Mediated Dephosphorylation Used for Peptide Cleavage during Solid Phase Peptide Synthesis

    Directory of Open Access Journals (Sweden)

    Byunghee Yoo

    2013-04-01

    Full Text Available Lanthanide(III ions can accelerate the hydrolysis of phosphomonoesters and phosphodiesters in neutral aqueous solution. In this paper, lanthanide-mediated dephosphorylation has been applied in aqueous media as an orthogonal cleavage condition that can be employed in conventional solid phase peptide synthesis (SPPS. A phosphorylated polymeric support for SPPS was developed using Boc chemistry. The cleavage of resin-bound phosphates was investigated with the addition of Eu(III, Yb(III, acid or base, a mixture of solvents or different temperatures. To demonstrate the utility of this approach for SPPS, a peptide sequence was synthesized on a phosphorylated polymeric support and quantitatively cleaved with lanthanide ions in neutral aqueous media. The protecting groups for side chains were retained during peptide cleavage using lanthanide ions. This new methodology provides a mild orthogonal cleavage condition of phosphoester as a linker during SPPS.

  8. An ester extract of Cochinchina momordica seeds induces differentiation of melanoma B16 F1 cells via MAPKs signaling.

    Science.gov (United States)

    Zhao, Lian-Mei; Han, Li-Na; Ren, Feng-Zhi; Chen, Shu-Hong; Liu, Li-Hua; Wang, Ming-Xia; Sang, Mei-Xiang; Shan, Bao-En

    2012-01-01

    Cochinchina momordica seeds (CMS) have been widely used due to antitumor activity by Mongolian tribes of China. However, the details of the underlying mechanisms remain unknown. In the present study, we found that an EtOAc (ethyl ester) extract of CMS (CMSEE) induced differentiation and caused growth inhibition of melanoma B16 F1 cells. CMSEE at the concentration of 5-200 μg/ml exhibited strongest anti-proliferative effects on B16 F1 cells among other CMS fractions (water or petroleum ether). Moreover, CMSEE induced melanoma B16 F1 cell differentiation, characterized by dendrite-like outgrowth, increasing melanogenesis production, as well as enhancing tyrosinase activity. Western blot analysis showed that sustained phosphorylation of p38 MAP accompanied by decrease in ERK1/2 and JNK dephosphorylation were involved in CMSEE-induced B16 F1 cell differentiation. Notably, 6 compounds that were isolated and identified may be responsible for inducing differentiation of CMSEE. These results indicated that CMSEE contributes to the differentiation of B16 F1 cells through modulating MAPKs activity, which may throw some light on the development of potentially therapeutic strategies for melanoma treatment.

  9. A non-Mendelian MAPK-generated hereditary unit controlled by a second MAPK pathway in Podospora anserina.

    Science.gov (United States)

    Lalucque, Hervé; Malagnac, Fabienne; Brun, Sylvain; Kicka, Sébastien; Silar, Philippe

    2012-06-01

    The Podospora anserina PaMpk1 MAP kinase (MAPK) signaling pathway can generate a cytoplasmic and infectious element resembling prions. When present in the cells, this C element causes the crippled growth (CG) cell degeneration. CG results from the inappropriate autocatalytic activation of the PaMpk1 MAPK pathway during growth, whereas this cascade normally signals stationary phase. Little is known about the control of such prion-like hereditary units involved in regulatory inheritance. Here, we show that another MAPK pathway, PaMpk2, is crucial at every stage of the fungus life cycle, in particular those controlled by PaMpk1 during stationary phase, which includes the generation of C. Inactivation of the third P. anserina MAPK pathway, PaMpk3, has no effect on the development of the fungus. Mutants of MAPK, MAPK kinase, and MAPK kinase kinase of the PaMpk2 pathway are unable to present CG. This inability likely relies upon an incorrect activation of PaMpk1, although this MAPK is normally phosphorylated in the mutants. In PaMpk2 null mutants, hyphae are abnormal and PaMpk1 is mislocalized. Correspondingly, stationary phase differentiations controlled by PaMpk1 are defective in the mutants of the PaMpk2 cascade. Constitutive activation of the PaMpk2 pathway mimics in many ways its inactivation, including an effect on PaMpk1 localization. Analysis of double and triple mutants inactivated for two or all three MAPK genes undercover new growth and differentiation phenotypes, suggesting overlapping roles. Our data underscore the complex regulation of a prion-like element in a model organism.

  10. Chemical characterization and factor analysis of PM2.5 in two sites of Monterrey, Mexico.

    Science.gov (United States)

    Martinez, Marco A; Caballero, Porfirio; Carrillo, Olivia; Mendoza, Alberto; Mejia, Gerardo Manuel

    2012-07-01

    The Monterrey Metropolitan Area (MMA) has shown a high concentration of PM2.5 in its atmosphere since 2003. The contribution of possible sources of primary PM2.5 and its precursors is not known. In this paper we present the results of analyzing the chemical composition of sixty 24-hr samples of PM2.5 to determine possible sources of PM2.5 in the MMA. The samples were collected at the northeast and southeast of the MMA between November 22 and December 12, 2007, using low-volume devices. Teflon and quartz filters were used to collect the samples. The concentrations of 16 airborne trace elements were determined using x-ray fluorescence (XRF). Anions and cations were determined using ion chromatography. Organic carbon (OC) and elemental carbon (EC) were determined by thermal optical analysis. The results show that Ca had the maximum mean concentration of all elements studied, followed by S. Enrichment factors above 50 were calculated for S, Cl, Cu, Zn, Br and Pb. This indicates that these elements may come from anthropogenic sources. Overall, the major average components of PM2.5 were OC (41.7%), SO4(2-) (22.9%), EC (7.4%), crustal material (11.4%), and NO3- (12.6%), which altogether accounted for 96% of the mass. Statistically, we did not find any difference in SO4(2-) concentrations between the two sites. The fraction of secondary organic carbon was between 24% and 34%. The results of the factor analysis performed over 10 metals and OC and EC show that there are three main sources of PM2.5: crustal material and vehicle exhaust; industrial activity; and fuel oil burning. The results show that SO4(2-), OC, and crustal material are important components of PM2.5 in MMA. Further work is necessary to evaluate the proportion of secondary inorganic and organic aerosol in order to have a better understanding of the sources and precursors of aerosols in the MMA.

  11. MiR-26b inhibits melanoma cell proliferation and enhances apoptosis by suppressing TRAF5-mediated MAPK activation

    Energy Technology Data Exchange (ETDEWEB)

    Li, Meng; Long, Chaoqin; Yang, Guilan; Luo, Yang; Du, Hua, E-mail: lemonlives_dr@163.com

    2016-03-11

    Alterations in microRNA-26b (miR-26b) expression have been shown to participate in various malignant tumor developments. However, the possible function of miR-26b in human melanoma cells remains unclarified. In this study, quantitative polymerase chain reaction was used to explore the expression profiles of miR-26b in melanoma cells. The effect of miR-26b on cell viability was determined by using MTT assays and colony formation assay. The apoptosis levels were evaluated by using Annexin V/fluorescein isothiocyanate (FITC) apoptosis detection kit and the apoptosis cells were confirmed by Transmission Electron Microscopy (TEM). Luciferase reporter plasmids were constructed to confirm direct targeting. Our study found that the expression of miR-26b was downregulated in human melanoma specimens. Overexpression of miR-26b significantly increased the anti-proliferative effects and apoptosis in A375 and B16F10 melanoma cells. In addition, luciferase gene reporter assays confirmed that TRAF5 was a direct target gene of miR-26b and the anti-tumor effect of miR-26b in melanoma cells was significantly counteracted by treatment with TRAF5 overexpression. Furthermore, the molecular mechanisms underlying the tumor suppressor of miR-26b in malignant melanomas may be due to the dephosphorylation of MAPK pathway caused by the decrease in TRAF5 expression when miR-26b is up-regulated in melanoma cells. These findings indicate that miR-26b might influence TRAF5-MAPK signaling pathways to facilitate the malignant progression of melanoma cells. - Highlights: • miR-26b is downregulated in human melanomas. • miR-26b suppressed melanoma cell proliferation and enhanced cell apoptosis. • TRAF5 is a direct target of miR-26b and inversely correlates with miR-26b expression. • miR-26b modulated MAPK signaling pathway by targeting TRAF5.

  12. The crystal structure of phosphorylated MAPK13 reveals common structural features and differences in p38 MAPK family activation

    OpenAIRE

    Yurtsever, Zeynep; Scheaffer, Suzanne M.; Romero, Arthur G.; Holtzman, Michael J.; Brett, Tom J.

    2015-01-01

    The p38 MAP kinases are an important family of drug targets for a myriad of inflammatory, as well as other, diseases. Presented here is the crystal structure of the active form of MAPK13 (p38d) as well as a comprehensive analysis of all known apo inactive and active p38 MAPK structures, revealing a common mode of activation as well as some unique structural features.

  13. Protein phosphatase 1γ is responsible for dephosphorylation of histone H3 at Thr 11 after DNA damage.

    Science.gov (United States)

    Shimada, Midori; Haruta, Mayumi; Niida, Hiroyuki; Sawamoto, Kazunobu; Nakanishi, Makoto

    2010-11-01

    The DNA-damage-induced transcriptional suppression of cell cycle regulatory genes correlates with a reduction in histone H3-Thr 11 phosphorylation (H3-pThr 11) on their promoters that is partly mediated by the dissociation of Chk1 from chromatin. In this study, we identify protein phosphatase 1γ (PP1γ) as a phosphatase responsible for DNA-damage-induced H3-pThr 11 dephosphorylation. PP1γ is activated after DNA damage, which is mainly mediated by a reduction in Cdk-dependent phosphorylation of PP1γ at Thr 311. The depletion of PP1γ sensitizes HCT116 cells to DNA damage. Our results suggest that the ataxia telangiectasia, mutated and Rad3-related-Chk1 axis regulates H3-pThr 11 dephosphorylation on DNA damage, at least in part by the activation of PP1γ through Chk1-dependent inhibition of Cdks.

  14. Release of Ecdysteroid-Phosphates from Egg Yolk Granules and Their Dephosphorylation during Early Embryonic Development in Silkworm, Bombyx mori

    OpenAIRE

    Yamada, Ryouichi; Yamahama, Yumi; Sonobe, Haruyuki

    2005-01-01

    Newly laid eggs of many insect species store maternal ecdysteroids as physiologically inactive phosphoric esters. In the silkworm Bombyx mori, we previously reported the presence of a specific enzyme, called ecdysteroid-phosphate phosphatase (EPPase), which catalyzes the dephosphorylation of ecdysteroid-phosphates to increase the amount of free ecdysteroids during early embryonic development. In this study, we demonstrated that (1) EPPase is found in the cytosol of yolk cells, (2) ecdysteroid...

  15. Glutamate Counteracts Dopamine/PKA Signaling via Dephosphorylation of DARPP-32 Ser-97 and Alteration of Its Cytonuclear Distribution.

    Science.gov (United States)

    Nishi, Akinori; Matamales, Miriam; Musante, Veronica; Valjent, Emmanuel; Kuroiwa, Mahomi; Kitahara, Yosuke; Rebholz, Heike; Greengard, Paul; Girault, Jean-Antoine; Nairn, Angus C

    2017-01-27

    The interaction of glutamate and dopamine in the striatum is heavily dependent on signaling pathways that converge on the regulatory protein DARPP-32. The efficacy of dopamine/D1 receptor/PKA signaling is regulated by DARPP-32 phosphorylated at Thr-34 (the PKA site), a process that inhibits protein phosphatase 1 (PP1) and potentiates PKA action. Activation of dopamine/D1 receptor/PKA signaling also leads to dephosphorylation of DARPP-32 at Ser-97 (the CK2 site), leading to localization of phospho-Thr-34 DARPP-32 in the nucleus where it also inhibits PP1. In this study the role of glutamate in the regulation of DARPP-32 phosphorylation at four major sites was further investigated. Experiments using striatal slices revealed that glutamate decreased the phosphorylation states of DARPP-32 at Ser-97 as well as Thr-34, Thr-75, and Ser-130 by activating NMDA or AMPA receptors in both direct and indirect pathway striatal neurons. The effect of glutamate in decreasing Ser-97 phosphorylation was mediated by activation of PP2A. In vitro phosphatase assays indicated that the PP2A/PR72 heterotrimer complex was likely responsible for glutamate/Ca(2+)-regulated dephosphorylation of DARPP-32 at Ser-97. As a consequence of Ser-97 dephosphorylation, glutamate induced the nuclear localization in cultured striatal neurons of dephospho-Thr-34/dephospho-Ser-97 DARPP-32. It also reduced PKA-dependent DARPP-32 signaling in slices and in vivo Taken together, the results suggest that by inducing dephosphorylation of DARPP-32 at Ser-97 and altering its cytonuclear distribution, glutamate may counteract dopamine/D1 receptor/PKA signaling at multiple cellular levels. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  16. Slingshot-3 dephosphorylates ADF/cofilin but is dispensable for mouse development.

    Science.gov (United States)

    Kousaka, Kazuyoshi; Kiyonari, Hiroshi; Oshima, Naoko; Nagafuchi, Akira; Shima, Yasuyuki; Chisaka, Osamu; Uemura, Tadashi

    2008-05-01

    Actin-depolymerizing factor (ADF) and cofilin constitute a family of key regulators of actin filament dynamics. ADF/cofilin is inactivated by phosphorylation at Ser-3 by LIM-kinases and reactivated by dephosphorylation by Slingshot (SSH) family phosphatases. Defects in LIM kinases or ADF/cofilin have been implicated in morbidity in human or mice; however, the roles of mammalian SSH in vivo have not been addressed. In this study, we examined the endogenous expression of each mouse SSH member in various cell lines and tissues, and showed that SSH-3L protein was strongly expressed in epithelial cells. Our structure-function analysis of SSH-3L suggested the possibility that the C-tail unique to SSH-3L negatively regulates the catalytic activity of this phosphatase. Furthermore we made ssh-3 knockout mice to examine its potential in vivo roles. Unexpectedly, ssh-3 was not essential for viability, fertility, or development of epithelial tissues; and ssh-3 did not genetically modify the corneal disorder of the corn1/ADF/destrin mutant.

  17. Protein serine/threonine phosphatase PPEF-1 suppresses genotoxic stress response via dephosphorylation of PDCD5

    Science.gov (United States)

    Park, Soo-Yeon; Seo, Jaesung; Choi, Hyo-Kyoung; Oh, Hye-Jeong; Guk, Garam; Lee, Yoo-Hyun; Lee, Jeongmin; Jun, Woo Jin; Choi, Kyung-Chul; Yoon, Ho-Geun

    2017-01-01

    Programmed cell death 5 (PDCD5) is believed to play a crucial role in p53 activation; however, the underlying mechanism of how PDCD5 function is regulated during apoptosis remains obscure. Here, we report that the serine/threonine phosphatase PPEF-1 interacts with and dephosphorylates PDCD5 at Ser-119, which leads to PDCD5 destabilization. Overexpression of wild-type PPEF-1, but not inactive PPEF-1D172N, efficiently suppressed CK2α-mediated stabilization of PDCD5 and p53-mediated apoptosis in response to etoposide (ET). Conversely, PPEF-1 knockdown further enhanced genotoxic stress responses. Notably, PPEF-1 suppressed p53-mediated genotoxic stress response via negative regulation of PDCD5. We also determined that overexpression of wild-type PPEF-1, but not inactive PPEF-1D172N, significantly increased tumorigenic growth and chemoresistance of A549 human lung carcinoma cells. Collectively, these data demonstrate that PPEF-1 plays a pivotal role in tumorigenesis of lung cancer cells by reducing PDCD5-mediated genotoxic stress responses. PMID:28051100

  18. Dephosphorylation of cardiomyocyte Cx43 is associated with myocardial ischemia and reperfusion injury

    Institute of Scientific and Technical Information of China (English)

    Zhijuan Cao; Xuan Xu; Linli Que; Qi Chen; Yuehua Li

    2009-01-01

    Objective:Myocardial ischemia/reperfusion(I/R) injury is the leading cause of death in the world. However, the details of the mechanism of its pathophysioiogy are still unknown. The present study was designed to investigate the role of connexin 43(Cx63) in acute models of myocardial I/R injury. Methods: Male C57BL/6 mice were subjected to myocardial ischemia(45 rain) followed by reperfusion(4 hrs) in vivo. The whole operation was monitored using a two-lead ECG. Hearts were harvested and the level of protein was assessed by western blot analysis. Haematoxylin and Eosin(HE) staining was used to detect the extent of neutrophil infiltration. The expression level of IL-6 was detected by ELISA. Results: A murine myocardial I/R injury model was constructed successfully. Phosphorylated Cx43 decreased 83.45% while non-phosphorylated Cx43 increased 1.62- fold in the myocardium after I/R injury. Neutrophil infiltration and the expression of the inflammatory cytokine IL-6 increased in the myocardium following I/R. Conclusion: During myocardial I/R injury, cardiomyocyte Cx43 is dephosphorylated, and this may be associated with an inflammatory response.

  19. Site-specific dephosphorylation of tau of apolipoprotein E-deficient and control mice by M1 muscarinic agonist treatment.

    Science.gov (United States)

    Genis, I; Fisher, A; Michaelson, D M

    1999-01-01

    Apolipoprotein E (apoE)-deficient mice have memory deficits that are associated with synaptic loss of basal forebrain cholinergic projections and with hyperphosphorylation of distinct epitopes of the microtubule-associated protein tau. Furthermore, treatment of apoE-deficient mice with the M1 selective agonist 1-methylpiperidine-4-spiro-(2'-methylthiazoline) [AF150(S)] abolishes their memory deficits and results in recovery of their brain cholinergic markers. In the present study, we used a panel of anti-tau monoclonal antibodies to further map the tau epitopes that are hyperphosphorylated in apoE-deficient mice and examined the effects of prolonged treatment with AF150(S). This revealed that tau of apoE-deficient mice contains a distinct, hyperphosphorylated "hot spot" domain which is localized N-terminally to the microtubule binding domain of tau, and that AF150(S) has an epitope-specific tau dephosphorylating effect whose magnitude is affected by apoE deficiency. Accordingly, epitopes which reside in the hyperphosphorylated "hot spot" are dephosphorylated by AF150(S) in apoE-deficient mice but are almost unaffected in the controls, whereas epitopes which flank this tau domain are dephosphorylated by AF150(S) in both mice groups. In contrast, epitopes located at the N and C terminals of tau are unaffected by AF150(S) in both groups of mice. These findings suggest that apoE deficiency results in hyperphosphorylation of a distinct tau domain whose excess phosphorylation can be reduced by muscarinic treatment.

  20. PTEN dephosphorylates AKT to prevent the expression of GLUT1 on plasmamembrane and to limit glucose consumption in cancer cells.

    Science.gov (United States)

    Phadngam, Suratchanee; Castiglioni, Andrea; Ferraresi, Alessandra; Morani, Federica; Follo, Carlo; Isidoro, Ciro

    2016-12-20

    GLUT1 is the facilitative transporter playing the major role in the internalization of glucose. Basally, GLUT1 resides on vesicles located in a para-golgian area, and is translocated onto the plasmamembrane upon activation of the PI3KC1-AKT pathway. In proliferating cancer cells, which demand a high quantity of glucose for their metabolism, GLUT1 is permanently expressed on the plasmamembrane. This is associated with the abnormal activation of the PI3KC1-AKT pathway, consequent to the mutational activation of PI3KC1 and/or the loss of PTEN. The latter, in fact, could antagonize the phosphorylation of AKT by limiting the availability of Phosphatidylinositol (3,4,5)-trisphosphate. Here, we asked whether PTEN could control the plasmamembrane expression of GLUT1 also through its protein-phosphatase activity on AKT. Experiments of co-immunoprecipitation and in vitro de-phosphorylation assay with homogenates of cells transgenically expressing the wild type or knocked-down mutants (lipid-phosphatase, protein-phosphatase, or both) isoforms demonstrated that indeed PTEN physically interacts with AKT and drives its dephosphorylation, and so limiting the expression of GLUT1 at the plasmamembrane. We also show that growth factors limit the ability of PTEN to dephosphorylate AKT. Our data emphasize the fact that PTEN acts in two distinct steps of the PI3k/AKT pathway to control the expression of GLUT1 at the plasmamembrane and, further, add AKT to the list of the protein substrates of PTEN.

  1. Sit4p/PP6 regulates ER-to-Golgi traffic by controlling the dephosphorylation of COPII coat subunits.

    Science.gov (United States)

    Bhandari, Deepali; Zhang, Jinzhong; Menon, Shekar; Lord, Christopher; Chen, Shuliang; Helm, Jared R; Thorsen, Kevin; Corbett, Kevin D; Hay, Jesse C; Ferro-Novick, Susan

    2013-09-01

    Traffic from the endoplasmic reticulum (ER) to the Golgi complex is initiated when the activated form of the GTPase Sar1p recruits the Sec23p-Sec24p complex to ER membranes. The Sec23p-Sec24p complex, which forms the inner shell of the COPII coat, sorts cargo into ER-derived vesicles. The coat inner shell recruits the Sec13p-Sec31p complex, leading to coat polymerization and vesicle budding. Recent studies revealed that the Sec23p subunit sequentially interacts with three different binding partners to direct a COPII vesicle to the Golgi. One of these binding partners is the serine/threonine kinase Hrr25p. Hrr25p phosphorylates the COPII coat, driving the membrane-bound pool into the cytosol. The phosphorylated coat cannot rebind to the ER to initiate a new round of vesicle budding unless it is dephosphorylated. Here we screen all known protein phosphatases in yeast to identify one whose loss of function alters the cellular distribution of COPII coat subunits. This screen identifies the PP2A-like phosphatase Sit4p as a regulator of COPII coat dephosphorylation. Hyperphosphorylated coat subunits accumulate in the sit4Δ mutant in vivo. In vitro, Sit4p dephosphorylates COPII coat subunits. Consistent with a role in coat recycling, Sit4p and its mammalian orthologue, PP6, regulate traffic from the ER to the Golgi complex.

  2. Impaired insulin activation and dephosphorylation of glycogen synthase in skeletal muscle of women with polycystic ovary syndrome is reversed by pioglitazone treatment

    DEFF Research Database (Denmark)

    Glintborg, Dorte; Højlund, Kurt; Andersen, Nicoline Resen

    2008-01-01

    . No significant abnormalities in GSK-3alpha or -3beta were found in PCOS subjects. Pioglitazone treatment improved insulin-stimulated glucose metabolism and GS activity in PCOS (all P ... of GS including absent dephosphorylation at sites 2+2a contributes to insulin resistance in skeletal muscle in PCOS. The ability of pioglitazone to enhance insulin sensitivity, in part, involves improved insulin action on GS activity and dephosphorylation at NH2-terminal sites....

  3. Functional analysis of the MAPK pathways in fungi.

    Science.gov (United States)

    Martínez-Soto, Domingo; Ruiz-Herrera, José

    2017-07-18

    The Mitogen-Activated Protein Kinase (MAPK) signaling pathways constitute one of the most important and evolutionarily conserved mechanisms for the perception of extracellular information in all the eukaryotic organisms. The MAPK pathways are involved in the transfer to the cell of the information perceived from extracellular stimuli, with the final outcome of activation of different transcription factors that regulate gene expression in response to them. In all species of fungi, the MAPK pathways have important roles in their physiology and development; e.g. cell cycle control, mating, morphogenesis, response to different stresses, resistance to UV radiation and to temperature changes, cell wall assembly and integrity, degradation of cellular organelles, virulence, cell-cell signaling, fungus-plant interaction, and response to damage-associated molecular patterns (DAMPs). Considering the importance of the phylogenetically conserved MAPK pathways in fungi, an updated review of the knowledge on them is discussed in this article. This information reveals their importance, their distribution in fungal species evolutionarily distant and with different lifestyles, their organization and function, and the interactions occurring between different MAPK pathways, and with other signaling pathways, for the regulation of the most complex cellular processes. Copyright © 2017 Asociación Española de Micología. Publicado por Elsevier España, S.L.U. All rights reserved.

  4. Mitophagy is primarily due to alternative autophagy and requires the MAPK1 and MAPK14 signaling pathways.

    Science.gov (United States)

    Hirota, Yuko; Yamashita, Shun-ichi; Kurihara, Yusuke; Jin, Xiulian; Aihara, Masamune; Saigusa, Tetsu; Kang, Dongchon; Kanki, Tomotake

    2015-01-01

    In cultured cells, not many mitochondria are degraded by mitophagy induced by physiological cellular stress. We observed mitophagy in HeLa cells using a method that relies on the pH-sensitive fluorescent protein Keima. With this approach, we found that mitophagy was barely induced by carbonyl cyanide m-chlorophenyl hydrazone treatment, which is widely used as an inducer of PARK2/Parkin-related mitophagy, whereas a small but modest amount of mitochondria were degraded by mitophagy under conditions of starvation or hypoxia. Mitophagy induced by starvation or hypoxia was marginally suppressed by knockdown of ATG7 and ATG12, or MAP1LC3B, which are essential for conventional macroautophagy. In addition, mitophagy was efficiently induced in Atg5 knockout mouse embryonic fibroblasts. However, knockdown of RAB9A and RAB9B, which are essential for alternative autophagy, but not conventional macroautophagy, severely suppressed mitophagy. Finally, we found that the MAPKs MAPK1/ERK2 and MAPK14/p38 were required for mitophagy. Based on these findings, we conclude that mitophagy in mammalian cells predominantly occurs through an alternative autophagy pathway, requiring the MAPK1 and MAPK14 signaling pathways.

  5. Sequence alignment reveals possible MAPK docking motifs on HIV proteins.

    Directory of Open Access Journals (Sweden)

    Perry Evans

    Full Text Available Over the course of HIV infection, virus replication is facilitated by the phosphorylation of HIV proteins by human ERK1 and ERK2 mitogen-activated protein kinases (MAPKs. MAPKs are known to phosphorylate their substrates by first binding with them at a docking site. Docking site interactions could be viable drug targets because the sequences guiding them are more specific than phosphorylation consensus sites. In this study we use multiple bioinformatics tools to discover candidate MAPK docking site motifs on HIV proteins known to be phosphorylated by MAPKs, and we discuss the possibility of targeting docking sites with drugs. Using sequence alignments of HIV proteins of different subtypes, we show that MAPK docking patterns previously described for human proteins appear on the HIV matrix, Tat, and Vif proteins in a strain dependent manner, but are absent from HIV Rev and appear on all HIV Nef strains. We revise the regular expressions of previously annotated MAPK docking patterns in order to provide a subtype independent motif that annotates all HIV proteins. One revision is based on a documented human variant of one of the substrate docking motifs, and the other reduces the number of required basic amino acids in the standard docking motifs from two to one. The proposed patterns are shown to be consistent with in silico docking between ERK1 and the HIV matrix protein. The motif usage on HIV proteins is sufficiently different from human proteins in amino acid sequence similarity to allow for HIV specific targeting using small-molecule drugs.

  6. Acidosis-induced p38 MAPK activation and its implication in regulation of cardiac contractility

    Institute of Scientific and Technical Information of China (English)

    Ming ZHENG; Rong HOU; Rui-ping XIAO

    2004-01-01

    AIM: To determine the possible role of pH in mediating activation of p38 mitogen-activated protein kinase (MAPK) and the consequent function of activated p38 MAPK in regulating cardiac contractility. METHODS: Adult rat cardiomyocytes were isolated and cultured. Low pH media was used to induce intracellular acidosis and contraction of single cardiomyocyte was measured. RESULTS: Phosphorylation of p38 MAPK was increased during ischemia, and pHi was decreased. Intracellular acidosis activated p38 MAPK to a similar level as ischemia. Inhibition of p38 MAPK activation by SB203580, a specific inhibitor of p38 MAPK, reversed acidosis-mediated reduction of myocyte contractility. CONCLUSION: In adult rat cardiomyocytes, intracellular acidification activated p38 MAPK and decreased cardiac contractility. Pretreatment of cardiomyocytes with SB203580 completely blocked p38 MAPK activation and partially reversed acidosis-mediated decline of cardiac contractility.

  7. cAMP-Induced Histones H3 Dephosphorylation Is Independent of PKA and MAP Kinase Activations and Correlates With mTOR Inactivation.

    Science.gov (United States)

    Rodriguez, Pedro; Rojas, Juan

    2016-03-01

    cAMP is a second messenger well documented to be involved in the phosphorylation of PKA, MAP kinase, and histone H3 (H3). Early, we reported that cAMP also induced H3 dephosphorylation in a variety of proliferating cell lines. Herein, it is shown that cAMP elicits a biphasic H3 dephosphorylation independent of PKA activation in cycling cells. H89, a potent inhibitor of PKA catalytic sub-unite, could not abolish this effect. Additionally, H89 induces a rapid and biphasic H3 serine 10 dephosphorylation, while a decline in the basal phosphorylation of CREB/ATF-1 is observed. Rp-cAMPS, an analog of cAMP and specific inhibitor of PKA, is unable to suppress cAMP-mediated H3 dephosphorylation, whereas Rp-cAMPS effectively blocks CREB/ATF-1 hyper-phosphorylation by cAMP and its inducers. Interestingly, cAMP exerts a rapid and profound H3 dephosphorylation at much lower concentration (50-fold lower, 0.125 mM) than the concentration required for maximal CREB/ATF-1 phosphorylation (5 mM). Much higher cAMP concentration is required to fully induce CREB/ATF-1 gain in phosphate (5 mM), which correlates with the inhibition of H3 dephosphorylation. Also, the dephosphorylation of H3 does not overlap at onset of MAP kinase phosphorylation pathways, p38 and ERK. Surprisingly, rapamycin (an mTOR inhibitor), cAMP, and its natural inducer isoproterenol, elicit identical dephosphorylation kinetics on both S6K1 ribosomal kinase (a downstream mTOR target) and H3. Finally, cAMP-induced H3 dephosphorylation is PP1/2-dependent. The results suggest that a pathway, requiring much lower cAMP concentration to that required for CREB/ATF-1 hyper-phosphorylation, is responsible for histone H3 dephosphorylation and may be linked to mTOR down regulation.

  8. Synthesis and dephosphorylation of MARCKS in the late stages of megakaryocyte maturation drive proplatelet formation.

    Science.gov (United States)

    Machlus, Kellie R; Wu, Stephen K; Stumpo, Deborah J; Soussou, Thomas S; Paul, David S; Campbell, Robert A; Kalwa, Hermann; Michel, Thomas; Bergmeier, Wolfgang; Weyrich, Andrew S; Blackshear, Perry J; Hartwig, John H; Italiano, Joseph E

    2016-03-17

    Platelets are essential for hemostasis, and thrombocytopenia is a major clinical problem. Megakaryocytes (MKs) generate platelets by extending long processes, proplatelets, into sinusoidal blood vessels. However, very little is known about what regulates proplatelet formation. To uncover which proteins were dynamically changing during this process, we compared the proteome and transcriptome of round vs proplatelet-producing MKs by 2D difference gel electrophoresis (DIGE) and polysome profiling, respectively. Our data revealed a significant increase in a poorly-characterized MK protein, myristoylated alanine-rich C-kinase substrate (MARCKS), which was upregulated 3.4- and 5.7-fold in proplatelet-producing MKs in 2D DIGE and polysome profiling analyses, respectively. MARCKS is a protein kinase C (PKC) substrate that binds PIP2. In MKs, it localized to both the plasma and demarcation membranes. MARCKS inhibition by peptide significantly decreased proplatelet formation 53%. To examine the role of MARCKS in the PKC pathway, we treated MKs with polymethacrylate (PMA), which markedly increased MARCKS phosphorylation while significantly inhibiting proplatelet formation 84%, suggesting that MARCKS phosphorylation reduces proplatelet formation. We hypothesized that MARCKS phosphorylation promotes Arp2/3 phosphorylation, which subsequently downregulates proplatelet formation; both MARCKS and Arp2 were dephosphorylated in MKs making proplatelets, and Arp2 inhibition enhanced proplatelet formation. Finally, we used MARCKS knockout (KO) mice to probe the direct role of MARCKS in proplatelet formation; MARCKS KO MKs displayed significantly decreased proplatelet levels. MARCKS expression and signaling in primary MKs is a novel finding. We propose that MARCKS acts as a "molecular switch," binding to and regulating PIP2 signaling to regulate processes like proplatelet extension (microtubule-driven) vs proplatelet branching (Arp2/3 and actin polymerization-driven).

  9. File list: Oth.PSC.10.Mapk8.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.PSC.10.Mapk8.AllCell mm9 TFs and others Mapk8 Pluripotent stem cell SRX032471,S...RX032481,SRX032470,SRX032480 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.PSC.10.Mapk8.AllCell.bed ...

  10. File list: Oth.PSC.05.Mapk8.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.PSC.05.Mapk8.AllCell mm9 TFs and others Mapk8 Pluripotent stem cell SRX032471,S...RX032480,SRX032481,SRX032470 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.PSC.05.Mapk8.AllCell.bed ...

  11. File list: Oth.ALL.20.Mapk8.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.ALL.20.Mapk8.AllCell mm9 TFs and others Mapk8 All cell types SRX085427,SRX03248...9,SRX032481,SRX032470,SRX085426,SRX032471,SRX032480,SRX032490 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.ALL.20.Mapk8.AllCell.bed ...

  12. File list: Oth.ALL.05.Mapk8.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.ALL.05.Mapk8.AllCell mm9 TFs and others Mapk8 All cell types SRX085427,SRX08542...6,SRX032490,SRX032471,SRX032480,SRX032489,SRX032481,SRX032470 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.ALL.05.Mapk8.AllCell.bed ...

  13. File list: Oth.Neu.50.Mapk8.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Neu.50.Mapk8.AllCell mm9 TFs and others Mapk8 Neural SRX032489,SRX085426,SRX085...427,SRX032490 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.Neu.50.Mapk8.AllCell.bed ...

  14. File list: Oth.ALL.50.Mapk8.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.ALL.50.Mapk8.AllCell mm9 TFs and others Mapk8 All cell types SRX032489,SRX03248...1,SRX032470,SRX085426,SRX085427,SRX032480,SRX032471,SRX032490 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.ALL.50.Mapk8.AllCell.bed ...

  15. File list: Oth.PSC.20.Mapk8.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.PSC.20.Mapk8.AllCell mm9 TFs and others Mapk8 Pluripotent stem cell SRX032481,S...RX032470,SRX032471,SRX032480 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.PSC.20.Mapk8.AllCell.bed ...

  16. File list: Oth.PSC.50.Mapk8.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.PSC.50.Mapk8.AllCell mm9 TFs and others Mapk8 Pluripotent stem cell SRX032481,S...RX032470,SRX032480,SRX032471 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.PSC.50.Mapk8.AllCell.bed ...

  17. File list: Oth.Neu.20.Mapk8.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Neu.20.Mapk8.AllCell mm9 TFs and others Mapk8 Neural SRX085427,SRX032489,SRX085...426,SRX032490 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.Neu.20.Mapk8.AllCell.bed ...

  18. File list: Oth.Neu.05.Mapk8.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Neu.05.Mapk8.AllCell mm9 TFs and others Mapk8 Neural SRX085427,SRX085426,SRX032...490,SRX032489 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.Neu.05.Mapk8.AllCell.bed ...

  19. File list: Oth.ALL.10.Mapk8.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.ALL.10.Mapk8.AllCell mm9 TFs and others Mapk8 All cell types SRX085427,SRX03249...0,SRX085426,SRX032489,SRX032471,SRX032481,SRX032470,SRX032480 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.ALL.10.Mapk8.AllCell.bed ...

  20. File list: Oth.Neu.10.Mapk8.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Neu.10.Mapk8.AllCell mm9 TFs and others Mapk8 Neural SRX085427,SRX032490,SRX085...426,SRX032489 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.Neu.10.Mapk8.AllCell.bed ...

  1. Protein Tyrosine Phosphatases, TC-PTP, SHP1, and SHP2, Cooperate in Rapid Dephosphorylation of Stat3 in Keratinocytes Following UVB Irradiation

    Science.gov (United States)

    Kim, Dae Joon; Tremblay, Michel L.; DiGiovanni, John

    2010-01-01

    Stat3 is initially dephosphorylated in murine keratinocytes in response to UVB irradiation. Treatment with Na3VO4 desensitized keratinocytes to UVB-induced apoptosis with the recovery of phosphorylated Stat3 protein levels, implying that a protein tyrosine phosphatase (PTP) is involved in this mechanism. In the current work, we report that three PTPs including TC45 (the nuclear form of TC-PTP), SHP1, and SHP2 are involved in this rapid dephosphorylation of Stat3 in keratinocytes induced by UVB irradiation. Dephosphorylation of Stat3 was increased rapidly after UVB irradiation of cultured keratinocytes. Knockdown of TC-PTP, SHP1, or SHP2 using RNAi showed that these PTPs are likely responsible for most of the rapid Stat3 dephosphorylation observed following UVB irradiation. The level of phosphorylated Stat3 was significantly higher in keratinocytes transfected with TC-PTP, SHP1, or SHP2 siRNA in the presence or absence of UVB compared with keratinocytes transfected with control siRNA. TC45 was mainly localized in the cytoplasm of keratinocytes and translocated from cytoplasm to nucleus upon UVB irradiation. Stat3 dephosphorylation was associated with nuclear translocation of TC45. Further studies revealed that knockdown of all three phosphatases, using RNAi, prevented the rapid dephosphorylation of Stat3 following UVB irradiation. In mouse epidermis, the level of phosphorylated Stat3 was initially decreased, followed by a significant increase at later time points after UVB exposure. The levels of Stat3 target genes, such as cyclin D1 and c-Myc, followed the changes in activated Stat3 in response to UVB irradiation. Collectively, these results suggest that three phosphatases, TC45, SHP1, and SHP2, are primarily responsible for UVB-mediated Stat3 dephosphorylation and may serve as part of an initial protective mechanism against UV skin carcinogenesis. PMID:20421975

  2. Noise propagation in two-step series MAPK cascade.

    Directory of Open Access Journals (Sweden)

    Venkata Dhananjaneyulu

    Full Text Available Series MAPK enzymatic cascades, ubiquitously found in signaling networks, act as signal amplifiers and play a key role in processing information during signal transduction in cells. In activated cascades, cell-to-cell variability or noise is bound to occur and thereby strongly affects the cellular response. Commonly used linearization method (LM applied to Langevin type stochastic model of the MAPK cascade fails to accurately predict intrinsic noise propagation in the cascade. We prove this by using extensive stochastic simulations for various ranges of biochemical parameters. This failure is due to the fact that the LM ignores the nonlinear effects on the noise. However, LM provides a good estimate of the extrinsic noise propagation. We show that the correct estimate of intrinsic noise propagation in signaling networks that contain at least one enzymatic step can be obtained only through stochastic simulations. Noise propagation in the cascade depends on the underlying biochemical parameters which are often unavailable. Based on a combination of global sensitivity analysis (GSA and stochastic simulations, we developed a systematic methodology to characterize noise propagation in the cascade. GSA predicts that noise propagation in MAPK cascade is sensitive to the total number of upstream enzyme molecules and the total number of molecules of the two substrates involved in the cascade. We argue that the general systematic approach proposed and demonstrated on MAPK cascade must accompany noise propagation studies in biological networks.

  3. Tamarix gallica phenolics protect IEC-6 cells against H2O2 induced stress by restricting oxidative injuries and MAPKs signaling pathways.

    Science.gov (United States)

    Bettaib, Jamila; Talarmin, Hélène; Droguet, Mickaël; Magné, Christian; Boulaaba, Mondher; Giroux-Metges, Marie-Agnès; Ksouri, Riadh

    2017-05-01

    Polyphenolic compounds gained interest in the pharmaceutical research area due to their beneficial properties. Herein, antioxidant and cytoprotective capacities of T. gallica extract on H2O2-challenged rat small intestine epithelial cells were investigated. To set stress conditions, IEC-6 cultures were challenged with numerous H2O2 doses and durations. Then, 40μM H2O2 during 4h were selected to assess the cytoprotective effect of different T. gallica extract concentrations. Oxidative parameters, measured through CAT and SOD activities as well as MDA quantification were assessed. In addition, the expression of possibly involved MAPKs was also valued. Main results reported that T. gallica was rich in polyphenols and exhibited an important antioxidant activity (DPPH Assay, IC50=6μgmL(-1); ABTS(+) test, IC50=50μgmL(-1); Fe-reducing power, EC50=100μgmL(-1)). The exposure of IEC-6 cultures to 40μM H2O2 during 4h caused oxidative stress manifested by (i) over 70% cell mortality, (ii) over-activity of CAT (246%), (iii) excess in MDA content (18.4nmolmg(-1)) and (iiii) a trigger of JNK phosphorylation. Pretreatment with T. gallica extract, especially when used at 0.25μgmL(-1), restored cell viability to 122%, and normal cell morphology in H2O2-chalenged cells. In addition, this extract normalized CAT activity and MDA content (100% and 14.7nmolmg(-1), respectively) to their basal levels as compared to control cells. Furthermore, stopping cell death seems to be due to dephosphorylated JNK MAPK exerted by T. gallica bioactive compounds. In all, T. gallica components provided a cross-talk between regulatory pathways leading to an efficient cytoprotection against harmful oxidative stimulus. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  4. Nuclear localization domains of GATA activator Gln3 are required for transcription of target genes through dephosphorylation in Saccharomyces cerevisiae.

    Science.gov (United States)

    Numamoto, Minori; Tagami, Shota; Ueda, Yusuke; Imabeppu, Yusuke; Sasano, Yu; Sugiyama, Minetaka; Maekawa, Hiromi; Harashima, Satoshi

    2015-08-01

    The GATA transcription activator Gln3 in the budding yeast (Saccharomyces cerevisiae) activates transcription of nitrogen catabolite repression (NCR)-sensitive genes. In cells grown in the presence of preferred nitrogen sources, Gln3 is phosphorylated in a TOR-dependent manner and localizes in the cytoplasm. In cells grown in non-preferred nitrogen medium or treated with rapamycin, Gln3 is dephosphorylated and is transported from the cytoplasm to the nucleus, thereby activating the transcription of NCR-sensitive genes. Caffeine treatment also induces dephosphorylation of Gln3 and its translocation to the nucleus and transcription of NCR-sensitive genes. However, the details of the mechanism by which phosphorylation controls Gln3 localization and transcriptional activity are unknown. Here, we focused on two regions of Gln3 with nuclear localization signal properties (NLS-K, and NLS-C) and one with nuclear export signal (NES). We constructed various mutants for our analyses: gln3 containing point mutations in all potential phosphoacceptor sites (Thr-339, Ser-344, Ser-347, Ser-355, Ser-391) in the NLS and NES regions to produce non-phosphorylatable (alanine) or mimic-phosphorylatable (aspartic acid) residues; and deletion mutants. We found that phosphorylation of Gln3 was impaired in all of these mutations and that the aspartic acid substitution mutants showed drastic reduction of Gln3-mediated transcriptional activity despite the fact that the mutations had no effect on nuclear localization of Gln3. Our observations suggest that these regions are required for transcription of target genes presumably through dephosphorylation.

  5. Magnetite/Ceria-Codecorated Titanoniobate Nanosheet: A 2D Catalytic Nanoprobe for Efficient Enrichment and Programmed Dephosphorylation of Phosphopeptides.

    Science.gov (United States)

    Min, Qianhao; Li, Siyuan; Chen, Xueqin; Abdel-Halim, E S; Jiang, Li-Ping; Zhu, Jun-Jie

    2015-05-13

    Global characterization and in-depth understanding of phosphoproteome based on mass spectrometry (MS) desperately needs a highly efficient affinity probe during sample preparation. In this work, a ternary nanocomposite of magnetite/ceria-codecorated titanoniobate nanosheet (MC-TiNbNS) was synthesized by the electrostatic assembly of Fe3O4 nanospheres and in situ growth of CeO 2 nanoparticles on pre-exfoliated titanoniobate and eventually utilized as the probe and catalyst for the enrichment and dephosphorylation of phosphopeptides. The two-dimensional (2D) structured titanoniobate nanosheet not only promoted the efficacy of capturing phosphopeptides with enlarged surface area, but also functioned as a substrate for embracing the magnetic anchor Fe3O4 to enable magnetic separation and mimic phosphatase CeO2 to produce identifying signatures of phosphopeptides. Compared to single-component TiNbNS or CeO2 nanoparticles, the ternary nanocomposite provided direct evidence of the number of phosphorylation sites while maintaining the enrichment efficiency. Moreover, by altering the on-sheet CeO2 coverage, the dephosphorylation activity could be fine-tuned, generating continuously adjustable signal intensities of both phosphopeptides and their dephosphorylated tags. Exhaustive detection of both mono- and multiphosphorylated peptides with precise counting of their phosphorylation sites was achieved in the primary mass spectra in the cases of digests of standard phosphoprotein and skim milk, as well as a more complex biological sample, human serum. With the resulting highly informative mass spectra, this multifunctional probe can be used as a promising tool for the fast and comprehensive characterization of phosphopeptides in MS-based phosphoproteomics.

  6. p38 MAPK Signaling in Postnatal Tendon Growth and Remodeling

    Science.gov (United States)

    Schwartz, Andrew J.; Sarver, Dylan C.; Sugg, Kristoffer B.; Dzierzawski, Justin T.; Gumucio, Jonathan P.; Mendias, Christopher L.

    2015-01-01

    Tendon is a dynamic tissue whose structure and function is influenced by mechanical loading, but little is known about the fundamental mechanisms that regulate tendon growth and remodeling in vivo. Data from cultured tendon fibroblasts indicated that the p38 MAPK pathway plays an important role in tendon fibroblast proliferation and collagen synthesis in vitro. To gain greater insight into the mechanisms of tendon growth, and explore the role of p38 MAPK signaling in this process, we tested the hypotheses that inducing plantaris tendon growth through the ablation of the synergist Achilles tendon would result in rapid expansion of a neotendon matrix surrounding the original tendon, and that treatment with the p38 MAPK inhibitor SB203580 would prevent this growth. Rats were treated with vehicle or SB203580, and subjected to synergist ablation by bilateral tenectomy of the Achilles tendon. Changes in histological and biochemical properties of plantaris tendons were analyzed 3, 7, or 28 days after overload, and comparisons were made to non-overloaded animals. By 28 days after overload, tendon mass had increased by 30% compared to non-overloaded samples, and cross-sectional area (CSA) increased by around 50%, with most of the change occurring in the neotendon. The expansion in CSA initially occurred through the synthesis of a hyaluronic acid rich matrix that was progressively replaced with mature collagen. Pericytes were present in areas of active tendon growth, but never in the original tendon ECM. Inhibition of p38 MAPK resulted in a profound decrease in IL6 expression, and had a modest effect on the expression of other ECM and cell proliferation genes, but had a negligible impact on overall tendon growth. The combined results from this study provided novel insights into tendon mechanobiology, and suggest that p38 MAPK signaling does not appear to be necessary for tendon growth in vivo. PMID:25768932

  7. p38 MAPK signaling in postnatal tendon growth and remodeling.

    Directory of Open Access Journals (Sweden)

    Andrew J Schwartz

    Full Text Available Tendon is a dynamic tissue whose structure and function is influenced by mechanical loading, but little is known about the fundamental mechanisms that regulate tendon growth and remodeling in vivo. Data from cultured tendon fibroblasts indicated that the p38 MAPK pathway plays an important role in tendon fibroblast proliferation and collagen synthesis in vitro. To gain greater insight into the mechanisms of tendon growth, and explore the role of p38 MAPK signaling in this process, we tested the hypotheses that inducing plantaris tendon growth through the ablation of the synergist Achilles tendon would result in rapid expansion of a neotendon matrix surrounding the original tendon, and that treatment with the p38 MAPK inhibitor SB203580 would prevent this growth. Rats were treated with vehicle or SB203580, and subjected to synergist ablation by bilateral tenectomy of the Achilles tendon. Changes in histological and biochemical properties of plantaris tendons were analyzed 3, 7, or 28 days after overload, and comparisons were made to non-overloaded animals. By 28 days after overload, tendon mass had increased by 30% compared to non-overloaded samples, and cross-sectional area (CSA increased by around 50%, with most of the change occurring in the neotendon. The expansion in CSA initially occurred through the synthesis of a hyaluronic acid rich matrix that was progressively replaced with mature collagen. Pericytes were present in areas of active tendon growth, but never in the original tendon ECM. Inhibition of p38 MAPK resulted in a profound decrease in IL6 expression, and had a modest effect on the expression of other ECM and cell proliferation genes, but had a negligible impact on overall tendon growth. The combined results from this study provided novel insights into tendon mechanobiology, and suggest that p38 MAPK signaling does not appear to be necessary for tendon growth in vivo.

  8. Dephosphorylated cofilin expression is associated with poor prognosis in cases of human breast cancer: a tissue microarray analysis

    Directory of Open Access Journals (Sweden)

    Maimaiti Y

    2016-10-01

    Full Text Available Yusufu Maimaiti,1,2,* Zeming Liu,1,* Jie Tan,1 Kelimu Abudureyimu,2 Bangxing Huang,3 Chunping Liu,1 Yawen Guo,1 Changwen Wang,1 Xiu Nie,3 Jing Zhou,1 Tao Huang1 1Department of Breast and Thyroid Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 2Department of General Surgery, Research Institute of Minimally Invasive, People’s Hospital of Xinjiang Uygur Autonomous Region, Urumqi, 3Department of Pathology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People’s Republic of China *These authors contributed equally to this work Background: Proteins in the cofilin pathway regulate actin dynamics and may be involved in cancer cell migration and invasion. However, there are no direct data that suggest that dephosphorylated cofilin can affect breast cancer prognosis.Methods: We assessed the expressions of cofilin and phosphorylated cofilin (P-cofilin in breast cancer tissue microarrays (290 patients, mean follow-up: 95.7±2.49 months to evaluate dephosphorylated cofilin and its relationship with breast cancer prognosis. The associations of pathological characteristics with cumulative survival were evaluated using Kaplan–Meier analysis.Results: Univariate analyses revealed that overall survival was associated with cofilin levels, N category, TNM stage, estrogen receptor status, progesterone receptor status, and molecular subtypes. Cofilin status and TNM stage independently affected overall survival, although P-cofilin expression was not associated with patient survival. In the P-cofilin-negative subgroup, cofilin expression was significantly associated with patient survival, although cofilin expression was not significantly associated with patient survival in the P-cofilin-positive subgroup. We further analyzed the P-cofilin-negative cases and found that Ki-67 expression was significantly elevated in the subgroup that was strongly positive for

  9. Connecting two-component regulatory systems by a protein that protects a response regulator from dephosphorylation by its cognate sensor

    OpenAIRE

    Kato, Akinori; Groisman, Eduardo A.

    2004-01-01

    A fundamental question in signal transduction is how an organism integrates multiple signals into a cellular response. Here we report the mechanism by which the Salmonella PmrA/PmrB two-component system responds to the signal controlling the PhoP/PhoQ two-component system. We establish that the PhoP-activated PmrD protein binds to the phosphorylated form of the response regulator PmrA, preventing both its intrinsic dephosphorylation and that promoted by its cognate sensor kinase PmrB. This re...

  10. p38MAPK gene transfection induced the apoptosis of rat glioma cells C6

    Institute of Scientific and Technical Information of China (English)

    ZHANG Bi-cheng; LI Qing; YE Jing; WANG Ying-mei; LIN Sheng-cai

    2001-01-01

    To study the effect ofp38MAPK transfecfion on the biological characteristics of rat glioma cells C6. Methods: p38MAPK was transfected into C6 cells by lipofectin. Expression ofp38MAPK in C6 cells before and after transfection was detected by immunocytochemistry and Western-blot analysis. HE staining,transmission electron microscopy and flow cytometry were used to observe the cell morphology, adhesion and study the cell cycle. Results: p38MAPK expressed in C6 cells after transfection. Cell biological characteristics changed,and apoptotic cells emerged. Conclusion: Exogenous p38MAPK could induce the apoptosis of C6 cells.

  11. Mangrove sediment core analysis of foraminiferal assemblages - a study at two sites along the western coast of India

    Directory of Open Access Journals (Sweden)

    P. Vidya

    2014-02-01

    Full Text Available Mangroves are an unique habitat and are largely influenced by sea level changes and wave energy. Foraminifera (Protista preserved in mangrove sediments provide an excellent proxy for deducing past conditions. One meter deep mangrove core samples at two sites on the western coast of India were collected. The foraminiferal assemblages at various depths showed significant changes in the abundance and diversity down the cores. A total of 59 species belonging to 32 genera, 24 families and five suborders were identified from the cores of these two sites. The cores showed an abundance of genus Rotalidium particularly the species Rotalidium annectans. Other species identified include Ammonia, Elphidium, Nonion, Spiroloculina, Quinqueloculina, Globigerinoides, etc. The pH, organic matter and CaCO3 also showed variations down the cores. There was a lack of correlation between sediment characteristics and the abundance of foraminifera in the cores. The low diversity and differences in distribution of foraminifera compared to surface intertidal samples may be due to intense post depositional changes or anthropogenic disturbances. The mangrove ecology thus appears disturbed by various factors.

  12. LeMAPK1, LeMAPK2, and LeMAPK3 are associated with nitric oxide-induced defense response against Botrytis cinerea in the Lycopersicon esculentum fruit.

    Science.gov (United States)

    Zheng, Yanyan; Hong, Hui; Chen, Lin; Li, Jingyuan; Sheng, Jiping; Shen, Lin

    2014-02-12

    Nitric oxide (NO) and mitogen-activated protein kinases (MAPKs) are signal molecules involved in the disease resistance of plants. To investigate the role of tomato MAPKs in the NO-mediated defense response, mature green tomatoes (Lycopersicon esculentum Mill. cv. Qian-xi) were treated with a MAPKs inhibitor (1,4-diamino-2,3-dicyano-1,4-bis(o-amino-phenylmercapto) butadiene (U0126)), NO donor sodium nitroprusside (SNP), and SNP plus U0126. Treatment with U0126 increased the incidence of disease and size of lesion areas in the tomato fruits after being inoculated with Botrytis cinerea. NO enhanced the resistance of the tomato fruits against Botrytis cinerea invasion and the activities of nitric oxide synthase, Chitinase, β-1,3-glucanase, polyphenol oxidase, and phenylalanine ammonia-lyase. However, the effects of NO on disease resistance were weakened by the MAPKs inhibitor. Meanwhile, the relative expression of LeMAPK1, LeMAPK2, and LeMAPK3 in the (SNP + U0126)-treated fruits was lower than that in the SNP-treated fruits. The results suggest that LeMAPK1/2/3 are involved in NO-induced disease resistance of tomato fruits against Botrytis cinerea.

  13. Evolutionary history of mitogen-activated protein kinase (MAPK) genes in Lotus, Medicago, and Phaseolus.

    Science.gov (United States)

    Neupane, Achal; Nepal, Madhav P; Benson, Benjamin V; Macarthur, Kenton J; Piya, Sarbottam

    2013-11-01

    Mitogen-Activated Protein Kinase (MAPK) genes encode proteins that mediate various signaling pathways associated with biotic and abiotic stress responses in eukaryotes. The MAPK genes form a 3-tier signal transduction cascade between cellular stimuli and physiological responses. Recent identification of soybean MAPKs and availability of genome sequences from other legume species allowed us to identify their MAPK genes. The main objectives of this study were to identify MAPKs in 3 legume species, Lotus japonicus, Medicago truncatula, and Phaseolus vulgaris, and to assess their phylogenetic relationships. We used approaches in comparative genomics for MAPK gene identification and named the newly identified genes following Arabidopsis MAPK nomenclature model. We identified 19, 18, and 15 MAPKs and 7, 4, and 9 MAPKKs in the genome of Lotus japonicus, Medicago truncatula, and Phaseolus vulgaris, respectively. Within clade placement of MAPKs and MAPKKs in the 3 legume species were consistent with those in soybean and Arabidopsis. Among 5 clades of MAPKs, 4 founder clades were consistent to MAPKs of other plant species and orthologs of MAPK genes in the fifth clade-"Clade E" were consistent with those in soybean. Our results also indicated that some gene duplication events might have occurred prior to eudicot-monocot divergence. Highly diversified MAPKs in soybean relative to those in 3 other legume species are attributable to the polyploidization events in soybean. The identification of the MAPK genes in the legume species is important for the legume crop improvement; and evolutionary relationships and functional divergence of these gene members provide insights into plant genome evolution.

  14. Dephosphorylation and subcellular compartment change of the mitotic Bloom's syndrome DNA helicase in response to ionizing radiation.

    Science.gov (United States)

    Dutertre, Stéphanie; Sekhri, Redha; Tintignac, Lionel A; Onclercq-Delic, Rosine; Chatton, Bruno; Jaulin, Christian; Amor-Guéret, Mounira

    2002-02-22

    Bloom's syndrome is a rare human autosomal recessive disorder that combines a marked genetic instability and an increased risk of developing all types of cancers and which results from mutations in both copies of the BLM gene encoding a RecQ 3'-5' DNA helicase. We recently showed that BLM is phosphorylated and excluded from the nuclear matrix during mitosis. We now show that the phosphorylated mitotic BLM protein is associated with a 3'-5' DNA helicase activity and interacts with topoisomerase III alpha. We demonstrate that in mitosis-arrested cells, ionizing radiation and roscovitine treatment both result in the reversion of BLM phosphorylation, suggesting that BLM could be dephosphorylated through the inhibition of cdc2 kinase. This was supported further by our data showing that cdc2 kinase activity is inhibited in gamma-irradiated mitotic cells. Finally we show that after ionizing radiation, BLM is not involved in the establishment of the mitotic DNA damage checkpoint but is subjected to a subcellular compartment change. These findings lead us to propose that BLM may be phosphorylated during mitosis, probably through the cdc2 pathway, to form a pool of rapidly available active protein. Inhibition of cdc2 kinase after ionizing radiation would lead to BLM dephosphorylation and possibly to BLM recruitment to some specific sites for repair.

  15. Mitochondrial alterations in PINK1 deficient cells are influenced by calcineurin-dependent dephosphorylation of dynamin-related protein 1.

    Directory of Open Access Journals (Sweden)

    Anna Sandebring

    Full Text Available PTEN-induced novel kinase 1 (PINK1 mutations are associated with autosomal recessive parkinsonism. Previous studies have shown that PINK1 influences both mitochondrial function and morphology although it is not clearly established which of these are primary events and which are secondary. Here, we describe a novel mechanism linking mitochondrial dysfunction and alterations in mitochondrial morphology related to PINK1. Cell lines were generated by stably transducing human dopaminergic M17 cells with lentiviral constructs that increased or knocked down PINK1. As in previous studies, PINK1 deficient cells have lower mitochondrial membrane potential and are more sensitive to the toxic effects of mitochondrial complex I inhibitors. We also show that wild-type PINK1, but not recessive mutant or kinase dead versions, protects against rotenone-induced mitochondrial fragmentation whereas PINK1 deficient cells show lower mitochondrial connectivity. Expression of dynamin-related protein 1 (Drp1 exaggerates PINK1 deficiency phenotypes and Drp1 RNAi rescues them. We also show that Drp1 is dephosphorylated in PINK1 deficient cells due to activation of the calcium-dependent phosphatase calcineurin. Accordingly, the calcineurin inhibitor FK506 blocks both Drp1 dephosphorylation and loss of mitochondrial integrity in PINK1 deficient cells but does not fully rescue mitochondrial membrane potential. We propose that alterations in mitochondrial connectivity in this system are secondary to functional effects on mitochondrial membrane potential.

  16. Dephosphorylation of Bcl-10 by calcineurin is essential for canonical NF-κB activation in Th cells.

    Science.gov (United States)

    Frischbutter, Stefan; Gabriel, Christian; Bendfeldt, Hanna; Radbruch, Andreas; Baumgrass, Ria

    2011-08-01

    Antigen-specific stimulation of T helper (Th) cells initiates signaling cascades that ultimately result in the activation of the transcription factors NF-κB, NFAT, and AP-1 which regulate, together with other factors, many T-cell functions such as cytokine production, proliferation, and differentiation. Ordered assembly and different phosphorylation events, along with subcellular translocation of the CARMA1/Bcl-10/MALT1 complex, determine NF-κB activation after T-cell receptor (TCR) triggering. We now provide evidence that inhibition of the Ser/Thr phosphatase calcineurin (CaN) prevents dephosphorylation of Bcl-10. CaN, in constant interaction with the Bcl-10/MALT1 complex, is able to dephosphorylate Bcl-10. The CaN inhibitor cyclosporine A (CsA) converts a transient phosphorylation of Bcl-10 Ser138 during the immediate early phase of T-cell activation into a persistent state. Thus, subsequent processes such as IKKβ phosphorylation, IκBα degradation, p65 nuclear translocation, and DNA binding are diminished. Consistently, CsA treatment does not affect the phosphorylation pattern of the upstream kinase PKCθ. Together, our findings demonstrate that CaN functions as a critical signaling molecule during Th cell activation, regulating Bcl-10 phosphorylation and thereby NF-κB activation.

  17. Human Cdc14B promotes progression through mitosis by dephosphorylating Cdc25 and regulating Cdk1/cyclin B activity.

    Directory of Open Access Journals (Sweden)

    Indra Tumurbaatar

    Full Text Available Entry into and progression through mitosis depends on phosphorylation and dephosphorylation of key substrates. In yeast, the nucleolar phosphatase Cdc14 is pivotal for exit from mitosis counteracting Cdk1-dependent phosphorylations. Whether hCdc14B, the human homolog of yeast Cdc14, plays a similar function in mitosis is not yet known. Here we show that hCdc14B serves a critical role in regulating progression through mitosis, which is distinct from hCdc14A. Unscheduled overexpression of hCdc14B delays activation of two master regulators of mitosis, Cdc25 and Cdk1, and slows down entry into mitosis. Depletion of hCdc14B by RNAi prevents timely inactivation of Cdk1/cyclin B and dephosphorylation of Cdc25, leading to severe mitotic defects, such as delay of metaphase/anaphase transition, lagging chromosomes, multipolar spindles and binucleation. The results demonstrate that hCdc14B-dependent modulation of Cdc25 phosphatase and Cdk1/cyclin B activity is tightly linked to correct chromosome segregation and bipolar spindle formation, processes that are required for proper progression through mitosis and maintenance of genomic stability.

  18. Behavior of trace metals in the hydrothermal plume at two sites on the Izu-Bonin-Mariana Arc

    Science.gov (United States)

    Shitashima, K.

    2004-12-01

    Deep-sea hydrothermal systems play an important role in the oceanic geochemical cycles of trace metals. High concentration of trace metals of the basalt origin is discharged into the deep sea via the hydrothermal plume. The hydrothermal plume is widely diffused to the ocean by mixing with ambient seawater. The processes of input and removal in the diffusing hydrothermal plume differ by individual hydrothermal systems. In this presentation, the behavior of trace metals in the hydrothermal plume of two sites on the Izu-Bonin-Mariana Arc is compared. This study was funded by the O`Archaean ParkO_L project of MEXT. The hydrothermal plume samples were taken from the Suiyo Seamount and the southern Mariana Trough (Pika Site). The mini CTDT-RMS mounted twelve 1.2L Niskin bottles was installed onto the manned submersible. And the hydrothermal plume samples were collected with monitoring the anomaly of temperature and turbidity. The samples were immediately filtered in an onboard clean bench. Unfiltered sample for analysis of total (particulate + dissolved) trace metal and filtered sample for analysis of dissolved trace metal were acidified. Trace metals (Al, Mn, Fe, Cu and Zn) in the hydrothermal plume samples were analyzed by GFAAS. The ranges of concentration of Al, Mn, Fe, Cu and Zn in the hydrothermal plume samples collected from two sites are _`15uM, _`5uM, _`5uM, _` 0.2uM and _`0.6uM, respectively. The particulate phase is predominant form in Al, Fe, Cu and Zn, and Mn shows the superiority of dissolved form. At the Suiyo Seamount, the hydrothermal active site is located in the bottom of caldera. On the other hand, the hydrothermal active site exists on the top of off-ridge seamount at the southern Mariana Trough. The diffusion process of trace metals in the hydrothermal plume to the ocean differed by the topographic factor in two sites. It suggests that trace metals discharged from the vents are hardly diffused to the ocean surmounting the Suiyo Seamount caldera

  19. Resveratrol Induces Apoptosis and Autophagy in T-cell Acute Lymphoblastic Leukemia Cells by Inhibiting Akt/mTOR and Activating p38-MAPK

    Institute of Scientific and Technical Information of China (English)

    GE Jiao; LIU Yan; LI Qiang; GUO Xia; GU Ling; MA Zhi Gui; ZHU Yi Ping

    2013-01-01

    Objective To explore the effects of resveratrol-induced apoptosis and autophagy in T-cell acute lymphoblastic leukemia (T-ALL) cells and potential molecular mechanisms. Methods The anti-proliferation effect of resveratrol-induced, apoptosis and autophagy on T-ALL cells were detected by using MTT test, immunofluorescence, electronic microscope, and flow cytometry, respectively. Western blotting was performed for detecting changes of apoptosis-associated proteins, cell cycle regulatory proteins and state of activation of Akt, mTOR, p70S6K, 4E-BP1, and p38-MAPK. Results Resveratrol inhibited the proliferation and induced apoptosis and autophagy in T-ALL cells in a dose and time-dependent manner. It also induced cell cycle arrest at G0/G1 phase via up regulating cyclin-dependent kinase (CDK) inhibitors p21 and p27 and down regulating cyclin A and cyclin D1. Western blotting revealed that resveratrol significantly decreased the expression of antiapoptotic proteins (Mcl-1 and Bcl-2) and increased the expression of proapoptotic proteins (Bax, Bim, and Bad), and induced cleaved-caspase-3 in a time-dependent manner. Significant increase in ratio of LC3-II/LC3-I and Beclin 1 was also detected. Furthermore, resveratrol induced significant dephosphorylation of Akt, mTOR, p70S6K, and 4E-BP1, but enhanced specific phosphorylation of p38-MAPK which could be blocked by SB203580. When autophagy was suppressed by 3-MA, apoptosis in T-ALL cells induced by resveratrol was enhanced. Conclusion Our findings have suggested that resveratrol induces cell cycle arrest, apoptosis, and autophagy in T-ALL cells through inhibiting Akt/mTOR/p70S6K/4E-BP1 and activating p38-MAPK signaling pathways. Autophagy might play a role as a self-defense mechanism in T-ALL cells treated by resveratrol. Therefore, the reasonable inhibition of autophagy in T-ALL cells may serve as a promising strategy for resveratrol induced apoptosis and can be used as adjuvant chemotherapy for T-ALL.

  20. Poaceae, Secale spp. and Artemisia spp. pollen in the air at two sites of different degrees of urbanisation

    Directory of Open Access Journals (Sweden)

    Aleksandra Kruczek

    2017-03-01

    [b]Results.[/b] The first pollen grains to appear in the air are those produced by rye, followed by those produced by grass and wormwood. The pollen seasons of grasses and wormwood started about one week earlier in Gudowo than in Szczecin, while the pollen season of rye started at almost the same time in the country and in the city. Airborne pollen counts of grasses, rye and wormwood were much higher in the country than in the city. The differences most probably result from the different floristic composition at these two sites and reflect the local contribution of the taxa studied in the country. Conclusions. The risk of allergy caused by the pollen of the taxa analysed was much higher in Gudowo (in the country, than in Szczecin city

  1. Relationships between airborne fungal spore concentration of Cladosporium and the summer climate at two sites in Britain

    Science.gov (United States)

    Hollins, P. D.; Kettlewell, P. S.; Atkinson, M. D.; Stephenson, D. B.; Corden, J. M.; Millington, W. M.; Mullins, J.

    Cladosporium conidia have been shown to be important aeroallergens in many regions throughout the world, but annual spore concentrations vary considerably between years. Understanding these annual fluctuations may be of value in the clinical management of allergies. This study investigates the number of days in summer when spore concentration exceeds the allergenic threshold in relation to regional temperature and precipitation at two sites in England and Wales over 27 years. Results indicate that number of days in summer when the Cladosporium spores are above the allergenic concentration is positively correlated with regional temperature and negatively correlated with precipitation for both sites over the study period. Further analysis used a winter North Atlantic Oscillation index to explore the potential for long-range forecasting of the aeroallergen. For both spore measurement sites, a positive correlation exists between the winter North Atlantic Oscillation index and the number of days in summer above the allergenic threshold for Cladosporium spore concentration.

  2. Poaceae, Secale spp. and Artemisia spp. pollen in the air at two sites of different degrees of urbanisation.

    Science.gov (United States)

    Kruczek, Aleksandra; Puc, Małgorzata; Wolski, Tomasz

    2017-03-21

    Among herbal plants, most cases of allergic reactions, like seasonal inflammation of nasal mucosa, conjunctivitis and pollen asthma, are related to the allergens from grass pollen. As the blossoming and pollination of rye is known to start the pollen season of grasses, information about the airborne rye pollen count permits alerting the people allergic to certain allergens contained in rye pollen. An important cause of allergy is also the pollen from wormwood, blossoming in late summer, as its two main allergens produce cross-reactions with many other plant allergens. The aim of the study was to evaluate the risk of allergic reactions in persons with pollinosis on the basis of the pollen calendar, analysis of concentrations of pollen grains of grass and rye, and comparison of diurnal pattern of airborne pollen grain concentrations at two sites with different degrees of urbanisation (Gudowo in the country and the city of Szczecin) in 2012-2014. The concentration of pollen was measured by the volume method. Length of the pollination season was determined by the method of 98%, assuming that the beginning and the end of the pollen season are the days on which 1% and 99% of the annual sum of pollen appeared. The first pollen grains to appear in the air are those produced by rye, followed by those produced by grass and wormwood. The pollen seasons of grasses and wormwood started about one week earlier in Gudowo than in Szczecin, while the pollen season of rye started at almost the same time in the country and in the city. Airborne pollen counts of grasses, rye and wormwood were much higher in the country than in the city. The differences most probably result from the different floristic composition at these two sites and reflect the local contribution of the taxa studied in the country. The risk of allergy caused by the pollen of the taxa analysed was much higher in Gudowo (in the country), than in Szczecin city.

  3. Evidence for the Involvement of p38 MAPK Activation in Barnacle Larval Settlement

    KAUST Repository

    He, Li-Sheng

    2012-10-24

    The barnacle Balanus ( = Amphibalanus) amphitrite is a major marine fouling animal. Understanding the molecular mechanism of larval settlement in this species is critical for anti-fouling research. In this study, we cloned one isoform of p38 MAPK (Bar-p38 MAPK) from this species, which shares the significant characteristic of containing a TGY motif with other species such as yeast, Drosophila and humans. The activation of p38 MAPK was detected by an antibody that recognizes the conserved dual phosphorylation sites of TGY. The results showed that phospho-p38 MAPK (pp38 MAPK) was more highly expressed at the cyprid stage, particularly in aged cyprids, in comparison to other stages, including the nauplius and juvenile stages. Immunostaining showed that Bar-p38 MAPK and pp38 MAPK were mainly located at the cyprid antennules, and especially the third and fourth segments, which are responsible for substratum exploration during settlement. The expression and localization patterns of Bar-p38 MAPK suggest its involvement in larval settlement. This postulation was also supported by the larval settlement bioassay with the p38 MAPK inhibitor SB203580. Behavioral analysis by live imaging revealed that the larvae were still capable of exploring the surface of the substratum after SB203580 treatment. This shows that the effect of p38 MAPK on larval settlement might be by regulating the secretion of permanent proteinaceous substances. Furthermore, the level of pp38 MAPK dramatically decreased after full settlement, suggesting that Bar-p38 MAPK maybe plays a role in larval settlement rather than metamorphosis. Finally, we found that Bar-p38 MAPK was highly activated when larvae confronted extracts of adult barnacle containing settlement cues, whereas larvae pre-treated with SB203580 failed to respond to the crude adult extracts.

  4. Evidence for the involvement of p38 MAPK activation in barnacle larval settlement.

    Directory of Open Access Journals (Sweden)

    Li-Sheng He

    Full Text Available The barnacle Balanus ( = Amphibalanus amphitrite is a major marine fouling animal. Understanding the molecular mechanism of larval settlement in this species is critical for anti-fouling research. In this study, we cloned one isoform of p38 MAPK (Bar-p38 MAPK from this species, which shares the significant characteristic of containing a TGY motif with other species such as yeast, Drosophila and humans. The activation of p38 MAPK was detected by an antibody that recognizes the conserved dual phosphorylation sites of TGY. The results showed that phospho-p38 MAPK (pp38 MAPK was more highly expressed at the cyprid stage, particularly in aged cyprids, in comparison to other stages, including the nauplius and juvenile stages. Immunostaining showed that Bar-p38 MAPK and pp38 MAPK were mainly located at the cyprid antennules, and especially the third and fourth segments, which are responsible for substratum exploration during settlement. The expression and localization patterns of Bar-p38 MAPK suggest its involvement in larval settlement. This postulation was also supported by the larval settlement bioassay with the p38 MAPK inhibitor SB203580. Behavioral analysis by live imaging revealed that the larvae were still capable of exploring the surface of the substratum after SB203580 treatment. This shows that the effect of p38 MAPK on larval settlement might be by regulating the secretion of permanent proteinaceous substances. Furthermore, the level of pp38 MAPK dramatically decreased after full settlement, suggesting that Bar-p38 MAPK maybe plays a role in larval settlement rather than metamorphosis. Finally, we found that Bar-p38 MAPK was highly activated when larvae confronted extracts of adult barnacle containing settlement cues, whereas larvae pre-treated with SB203580 failed to respond to the crude adult extracts.

  5. Regulation of the MAPK pathway by raf kinase inhibitory protein.

    Science.gov (United States)

    Vandamme, Drieke; Herrero, Ana; Al-Mulla, Fahd; Kolch, Walter

    2014-01-01

    The Raf kinase inhibitor protein 1 (RKIP-1) was the first reported endogenous inhibitor of Raf-1-MEK-ERK/MAPK cascade, by interfering with the phosphorylation of MEK by Raf-1. However, RKIP's functions related to the MAPK signaling are far more complex. Newer data indicate that by modulating different protein-protein interactions, RKIP is involved in fine-tuning cell signaling, modulating ERK dynamics, and regulating cross talk between different pathways. Here, we describe the molecular mechanisms by which RKIP controls MAPK signaling at different levels and vice versa and its regulation via feedback phosphorylation. We also focus on several discrepancies and questions that remain, such as the RKIP binding regulation by Raf-1 N-region phosphorylation, the possible B-Raf inhibition, and the effects of RKIP-lipid binding. We also describe how RKIP's role as key signaling modulator of many cell fate decisions leads to the fact that fine control of RKIP activity and regulation is crucial to avoid pathological processes, such as metastasis, pulmonary arterial hypertension, and heart failure.

  6. Regulation between nitric oxide and MAPK signal transduction in mammals

    Institute of Scientific and Technical Information of China (English)

    TAO Yong; ZHANG Meijia; HONG Haiyan; XIA Guoliang

    2005-01-01

    Nitric oxide (NO) is an important biological messenger in the regulation of tissue homeostasis. It exhibits a wide range of effects during physiological and pathophysiological processes. Typical beneficial properties of NO include the regulation of vascular tone,the protection of cells against apoptosis, the modulation of immune responses, and the killing of microbial pathogens. On the other hand,NO may cause severe vasodilation and myocardial depression during bacterial sepsis or act as a cytotoxic and tissue-damaging molecule in autoimmune diseases. Mitogen-activated protein kinase (MAPK) is a family of serine/threonine protein kinases that are widely distributed in mammalian cells. MAPK cascade plays pivotal roles in gene expression, cell proliferation, differentiation, neuronal survival and programmed cell death under a variety of experimental conditions. MAPKs transduce the signal for the cellular response to extracellular stresses or stimuli. The relation between them, however, has never been reviewed. Based on our researches and other reports in the field, we review their reciprocal regulatory functions.

  7. Analysis of Stomatal Patterning in Selected Mutants of MAPK Pathways

    KAUST Repository

    Felemban, Abrar

    2016-05-01

    Stomata are cellular valves in plants that play an essential role in the regulation of gas exchange and are distributed in the epidermis of aerial organs. In Arabidopsis thaliana, stomatal production and development are coordinated by the mitogen-activated protein kinase (MAPK) signalling pathway, which modulates a variety of other processes, including cell proliferation, regulation of cytokinesis, programed cell death, and response to abiotic and biotic stress. The environment also plays a role in stomatal development, by influencing the frequency at which stomata develop in leaves. This thesis presents an analysis of stomatal development in Arabidopsis mutants in two MAPK pathways: MEKK1-MKK1/MKK2-MPK4, and MAP3K17/18-MKK3. Obtained results demonstrate the effect of stress conditions on stomatal development and specify the involvement of analysed MAPK in stomatal patterning. First, both analysed pathways modulate stomatal patterning in Arabidopsis cotyledons. Second, plant growth-promoting bacteria tested enhance stomatal density and affect guard cell morphology. Third, the sucrose or mannitol treatment increases defects in stomatal patterning. Finally, salt stress or high temperature can suppress stomatal defects in mutants of the MEKK1-MKK1/MKK2-MPK4 pathway.

  8. Cutaneous wound healing through paradoxical MAPK activation by BRAF inhibitors

    Science.gov (United States)

    Escuin-Ordinas, Helena; Li, Shuoran; Xie, Michael W.; Sun, Lu; Hugo, Willy; Huang, Rong Rong; Jiao, Jing; de-Faria, Felipe Meira; Realegeno, Susan; Krystofinski, Paige; Azhdam, Ariel; Komenan, Sara Marie D.; Atefi, Mohammad; Comin-Anduix, Begoña; Pellegrini, Matteo; Cochran, Alistair J.; Modlin, Robert L.; Herschman, Harvey R.; Lo, Roger S.; McBride, William H.; Segura, Tatiana; Ribas, Antoni

    2016-01-01

    BRAF inhibitors are highly effective therapies for the treatment of BRAFV600-mutated melanoma, with the main toxicity being a variety of hyperproliferative skin conditions due to paradoxical activation of the mitogen-activated protein kinase (MAPK) pathway in BRAF wild-type cells. Most of these hyperproliferative skin changes improve when a MEK inhibitor is co-administered, as it blocks paradoxical MAPK activation. Here we show how the BRAF inhibitor vemurafenib accelerates skin wound healing by inducing the proliferation and migration of human keratinocytes through extracellular signal-regulated kinase (ERK) phosphorylation and cell cycle progression. Topical treatment with vemurafenib in two wound-healing mice models accelerates cutaneous wound healing through paradoxical MAPK activation; addition of a mitogen-activated protein kinase kinase (MEK) inhibitor reverses the benefit of vemurafenib-accelerated wound healing. The same dosing regimen of topical BRAF inhibitor does not increase the incidence of cutaneous squamous cell carcinomas in mice. Therefore, topical BRAF inhibitors may have clinical applications in accelerating the healing of skin wounds. PMID:27476449

  9. Chk1 inhibition activates p53 through p38 MAPK in tetraploid cancer cells.

    Science.gov (United States)

    Vitale, Ilio; Senovilla, Laura; Galluzzi, Lorenzo; Criollo, Alfredo; Vivet, Sonia; Castedo, Maria; Kroemer, Guido

    2008-07-01

    We have previously shown that tetraploid cancer cells succumb through a p53-dependent apoptotic pathway when checkpoint kinase 1 (Chk1) is depleted by small interfering RNAs (siRNAs) or inhibited with 7-hydroxystaurosporine (UCN-01). Here, we demonstrate that Chk1 inhibition results in the activating phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK). Depletion of p38 MAPK by transfection with a siRNA targeting the alpha isoform of p38 MAPK (p38alpha MAPK) abolishes the phosphorylation of p53 on serines 15 and 46 that is induced by Chk1 knockdown. The siRNA-mediated downregulation and pharmacological inhibition of p38alpha MAPK (with SB 203580) also reduces cell death induced by Chk1 knockdown or UCN-01. These results underscore the role of p38 MAPK as a pro-apoptotic kinase in the p53-dependant pathway for the therapeutic elimination of polyploidy cells.

  10. MAPK signal pathways in the regulation of cell proliferation in mammalian cells

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    MAPK families play an important role in complex cellular programs like proliferation, differentiation,development, transformation, and apoptosis. At least three MAPK families have been characterized: extracellular signal-regulated kinase (ERK), Jun kinase (JNK/SAPK) and p38 MAPK. The above effects are fulfilled by regulation of cell cycle engine and other cell proliferation related proteins. In this paper we discussed their functions and cooperation with other signal pathways in regulation of cell proliferation.

  11. β-Arrestin 1’s Interaction with TC45 Attenuates Stat signaling by dephosphorylating Stat to inhibit antimicrobial peptide expression

    Science.gov (United States)

    Sun, Jie-Jie; Yang, Hui-Ting; Niu, Guo-Juan; Feng, Xiao-Wu; Lan, Jiang-Feng; Zhao, Xiao-Fan; Wang, Jin-Xing

    2016-01-01

    Impaired phosphatase activity leads to the persistent activation of signal transducers and activators of transcription (Stat). In mammals, Stat family members are often phosphorylated or dephosphorylated by the same enzymes. To date, only one Stat similar to mammalian Stat5a/b has been found in crustaceans and there have been few studies in Stat signal regulation in crustaceans. Here, we report that β-arrestin1 interacts with TC45 (45-kDa form of T cell protein tyrosine phosphatase) in the nucleus to attenuate Stat signaling by promoting dephosphorylation of Stat. Initially, we showed that Stat translocates into the nucleus to induce antimicrobial peptide (AMP) expression after bacterial infection. βArr1 enters the nucleus of hemocytes and recruits TC45 to form the βarr1-TC45-Stat complex, which dephosphorylates Stat efficiently. The interaction of TC45 with Stat decreased and Stat phosphorylation increased in βarr1-silenced shrimp (Marsupenaeus japonicus) after challenge with Vibrio anguillarum. βArr1 directly interacts with Stat in nucleus and accelerates Stat dephosphorylation by recruiting TC45 after V. anguillarum challenge. Further study showed that βarr1 and TC45 also affect AMP expression, which is regulated by Stat. Therefore, βarr1 and TC45 are involved in the anti-V. anguillarum immune response by regulating Stat activity negatively to decrease AMP expression in shrimp. PMID:27782165

  12. Quercetin induces apoptosis in the methotrexate-resistant osteosarcoma cell line U2-OS/MTX300 via mitochondrial dysfunction and dephosphorylation of Akt.

    Science.gov (United States)

    Xie, Xianbiao; Yin, Junqiang; Jia, Qiang; Wang, Jin; Zou, Changye; Brewer, Kari J; Colombo, Chiara; Wang, Yaofei; Huang, Gang; Shen, Jingnan

    2011-09-01

    Quercetin is the most abundant polyphenolic flavonoid found in plants. Several studies suggest that it has potent anticancer effects. The present study examines the apoptosis-inducing activity and the underlying mechanism of action of quercetin in a methotrexate (MTX)-resistant osteosarcoma model. Our results showed that quercetin inhibited cell viability in a dose-dependent manner and there was no cross-resistance between MTX and quercetin in U2-OS/MTX300 cells. The induction of apoptosis was observed by flow cyto-metry and fluorescence staining experiments. Quercetin-induced apoptosis was accompanied by a significant reduction of mitochondrial membrane potential, release of mitochondrial cytochrome c to the cytosol, activation of caspase-3, down-regulation of Bcl-2, p-Bad and up-regulation of Bax. A remarkable dephospho-rylation of Akt was also detected after quercetin treatment. Furthermore, transduction with constitutively active Akt protected against the quercetin-induced dephosphorylation of Akt and Bad as well as poly(ADP-ribose)polymerase (PARP) degradation, while combined treatment with quercetin and LY294002 enhanced the dephosphorylation of Akt, Bad and PARP cleavage in U2-OS/MTX300 cells. Taken together, our results demonstrate that quercetin-induced apoptosis in the MTX-resistant osteosarcoma cells U2-OS/MTX300 was mediated via mitochondrial dysfunction and dephosphorylation of Akt.

  13. PHOSPHORYLATION/DEPHOSPHORYLATION OF MITOCHONDRIAL PROTEINS IN REDOX-SIGNALLING OF HIGHER PLANTS UNDER ABIOTIC STRESS CONDITIONS

    Directory of Open Access Journals (Sweden)

    Subota I.Yu.

    2012-08-01

    . This phosphoprotein phosphatase primarily dephosphorylates the protein of the inner membrane. When the proteins of the outer membrane were added together with the phosphorylated mitoplast proteins, the activity of phosphorylation effectively decreased. Thus, the phosphoprotein phosphatases of the outer membrane could dephosphorylate the inner membrane proteins and vice versa, the inner membrane phosphatases dephosphorylate the outer membrane proteins. Mn-SOD is the first line protection enzyme of the mitochondrial DNA, as this enzyme scavenges superoxide anion and severely reduces the toxic influences of ROS. Till now, the regulation of this enzyme activity is not clear. One of such mechanisms can be the phosphorylation of serine and threonine residues. We studied the effect of different redox agents on the phosphorylation of the mitochondrial Mn-SOD by using the recombinant Mn-SOD protein. It was found out that the oxidizing agent K3[Fe(CN6] led to dephosphorylation of the Mn-SOD, while, at the same time, such reducing agents as sodium dithionite and GSH did not exert any effect on phosphorylation.

  14. Nitric oxide-induced biphasic mechanism of vascular relaxation via dephosphorylation of CPI-17 and MYPT1.

    Science.gov (United States)

    Kitazawa, Toshio; Semba, Shingo; Huh, Yang Hoon; Kitazawa, Kazuyo; Eto, Masumi

    2009-07-15

    Nitric oxide (NO) from endothelium is a major mediator of vasodilatation through cGMP/PKG signals that lead to a decrease in Ca(2+) concentration. In addition, NO-mediated signals trigger an increase in myosin light chain phosphatase (MLCP) activity. To evaluate the mechanism of NO-induced relaxation through MLCP deinhibition, we compared time-dependent changes in Ca(2+), myosin light chain (MLC) phosphorylation and contraction to changes in phosphorylation levels of CPI-17 at Thr38, RhoA at Ser188, and MYPT1 at Ser695, Thr696 and Thr853 in response to sodium nitroprusside (SNP)-induced relaxation in denuded rabbit femoral artery. During phenylephrine (PE)-induced contraction, SNP reduced CPI-17 phosphorylation to a minimal value within 15 s, in parallel with decreases in Ca(2+) and MLC phosphorylation, followed by a reduction of contractile force having a latency period of about 15 s. MYPT1 phosphorylation at Ser695, the PKG-target site, increased concurrently with relaxation. Phosphorylation of RhoA, MYPT1 Thr696 and Thr853 differed significantly at 5 min but not within 1 min of SNP exposure. Inhibition of Ca(2+) release delayed SNP-induced relaxation while inhibition of Ca(2+) channel, BK(Ca) channel or phosphodiesterase-5 did not. Pretreatment of resting artery with SNP suppressed an increase in Ca(2+), contractile force and phosphorylation of MLC, CPI-17, MYPT1 Thr696 and Thr853 at 10 s after PE stimulation, but had no effect on phorbol ester-induced CPI-17 phosphorylation. Together, these results suggest that NO production suppresses Ca(2+) release, which causes an inactivation of PKC and rapid CPI-17 dephosphorylation as well as MLCK inactivation, resulting in rapid MLC dephosphorylation and relaxation.

  15. Aβ promotes VDAC1 channel dephosphorylation in neuronal lipid rafts. Relevance to the mechanisms of neurotoxicity in Alzheimer's disease.

    Science.gov (United States)

    Fernandez-Echevarria, C; Díaz, M; Ferrer, I; Canerina-Amaro, A; Marin, R

    2014-10-10

    Voltage-dependent anion channel (VDAC) is a mitochondrial protein abundantly found in neuronal lipid rafts. In these membrane domains, VDAC is associated with a complex of signaling proteins that trigger neuroprotective responses. Loss of lipid raft integrity may result in disruption of multicomplex association and alteration of signaling responses that may ultimately promote VDAC activation. Some data have demonstrated that VDAC at the neuronal membrane may be involved in the mechanisms of amyloid beta (Aβ)-induced neurotoxicity, through yet unknown mechanisms. Aβ is generated from amyloid precursor protein (APP), and is released to the extracellular space where it may undergo self-aggregation. Aβ aggregate deposition in the form of senile plaques may lead to Alzheimer's disease (AD) neuropathology, although other pathological hallmarks (such as hyper-phosphorylated Tau deposition) also participate in this neurodegenerative process. The present study demonstrates that VDAC1 associates with APP and Aβ in lipid rafts of neurons. Interaction of VDAC1 with APP was observed in lipid rafts from the frontal and entorhinal cortex of human brains affected by AD at early stages (I-IV/0-B of Braak and Braak). Furthermore, Aβ exposure enhanced the dephosphorylation of VDAC1 that correlated with cell death. Both effects were reverted in the presence of tyrosine phosphatase inhibitors. VDAC1 dephosphorylation was corroborated in lipid rafts of AD brains. These results demonstrate that Aβ is involved in alterations of the phosphorylation state of VDAC in neuronal lipid rafts. Modulation of this channel may contribute to the development and progression of AD pathology.

  16. Shp-1 dephosphorylates TRPV1 in dorsal root ganglion neurons and alleviates CFA-induced inflammatory pain in rats.

    Science.gov (United States)

    Xiao, Xing; Zhao, Xiao-Tao; Xu, Ling-Chi; Yue, Lu-Peng; Liu, Feng-Yu; Cai, Jie; Liao, Fei-Fei; Kong, Jin-Ge; Xing, Guo-Gang; Yi, Ming; Wan, You

    2015-04-01

    Transient receptor potential vanilloid 1 (TRPV1) receptors are expressed in nociceptive neurons of rat dorsal root ganglions (DRGs) and mediate inflammatory pain. Nonspecific inhibition of protein-tyrosine phosphatases (PTPs) increases the tyrosine phosphorylation of TRPV1 and sensitizes TRPV1. However, less is known about tyrosine phosphorylation's implication in inflammatory pain, compared with that of serine/threonine phosphorylation. Src homology 2 domain-containing tyrosine phosphatase 1 (Shp-1) is a key phosphatase dephosphorylating TRPV1. In this study, we reported that Shp-1 colocalized with and bound to TRPV1 in nociceptive DRG neurons. Shp-1 inhibitors, including sodium stibogluconate and PTP inhibitor III, sensitized TRPV1 in cultured DRG neurons. In naive rats, intrathecal injection of Shp-1 inhibitors increased both TRPV1 and tyrosine-phosphorylated TRPV1 in DRGs and induced thermal hyperalgesia, which was abolished by pretreatment with TRPV1 antagonists capsazepine, BCTC, or AMG9810. Complete Freund's adjuvant (CFA)-induced inflammatory pain in rats significantly increased the expression of Shp-1, TRPV1, and tyrosine-phosphorylated TRPV1, as well as the colocalization of Shp-1 and TRPV1 in DRGs. Intrathecal injection of sodium stibogluconate aggravated CFA-induced inflammatory pain, whereas Shp-1 overexpression in DRG neurons alleviated it. These results suggested that Shp-1 dephosphorylated and inhibited TRPV1 in DRG neurons, contributing to maintain thermal nociceptive thresholds in normal rats, and as a compensatory mechanism, Shp-1 increased in DRGs of rats with CFA-induced inflammatory pain, which was involved in protecting against excessive thermal hyperalgesia.

  17. Suppression of cancer cell migration and invasion by protein phosphatase 2A through dephosphorylation of mu- and m-calpains.

    Science.gov (United States)

    Xu, Lijun; Deng, Xingming

    2006-11-17

    The mu- and m-calpains are major members of the calpain family that play an essential role in regulating cell motility. We have recently discovered that nicotine-activated protein kinase C iota enhances calpain phosphorylation in association with enhanced calpain activity and accelerated migration and invasion of human lung cancer cells. Here we found that specific disruption of protein phosphatase 2A (PP2A) activity by expression of SV40 small tumor antigen up-regulates phosphorylation of mu- and m-calpains whereas C2-ceramide, a potent PP2A activator, reduces nicotine-induced calpain phosphorylation, suggesting that PP2A may function as a physiological calpain phosphatase. PP2A co-localizes and interacts with mu- and m-calpains. Purified, active PP2A directly dephosphorylates mu- and m-calpains in vitro. Overexpression of the PP2A catalytic subunit (PP2A/C) suppresses nicotine-stimulated phosphorylation of mu- and m-calpains, which is associated with inhibition of calpain activity, wound healing, cell migration, and invasion. By contrast, depletion of PP2A/C by RNA interference enhances calpain phosphorylation, calpain activity, cell migration, and invasion. Importantly, C2-ceramide-induced suppression of calpain phosphorylation results in decreased secretion of mu- and m-calpains from lung cancer cells into culture medium, which may have potential clinic relevance in controlling metastasis of lung cancer. These findings reveal a novel role for PP2A as a physiological calpain phosphatase that not only directly dephosphorylates but also inactivates mu- and m-calpains, leading to suppression of migration and invasion of human lung cancer cells.

  18. Inhibition of Raf/MAPK signaling in Xenopus oocyte extracts by Raf-1-specific peptides.

    Science.gov (United States)

    Radziwill, G; Steinhusen, U; Aitken, A; Moelling, K

    1996-10-01

    Raf-1 is an upstream element of the mitogen-activated protein kinase (MAPK) pathway which leads to cell proliferation and differentiation. In this study Raf-1 derived peptides comprising the conserved amino acid residues Arg89 and Ser259, involved in binding of activated Ras and 14-3-3 proteins, respectively, were shown to interfere with MAPK activation in extracts from immature Xenopus oocytes. Lipids prepared from oocyte extracts can stimulate MAPK in a Ras- and protein kinase C-independent manner. This lipid-induced MAPK activation is blocked by a Raf-1 derived peptide comprising Ser259.

  19. Critical role of p38 MAPK in IL-4-induced alternative activation of peritoneal macrophages.

    Science.gov (United States)

    Jiménez-Garcia, Lidia; Herránz, Sandra; Luque, Alfonso; Hortelano, Sonsoles

    2015-01-01

    Alternative activation of macrophages plays an important role in a range of physiological and pathological processes. This alternative phenotype, also known as M2 macrophages, is induced by type 2 cytokines such as IL-4. The binding of IL-4 to its receptor leads to activation of two major signaling pathways: STAT-6 and PI3K. However, recent studies have described that p38 MAPK might play a role in IL-4-dependent signaling in some cells, although its role in macrophages is still controversial. In this study, we investigated whether p38 MAPK plays a role in the polarization of macrophages in mice. Our results reveal that IL-4 induces phosphorylation of p38 MAPK in thioglycollate-elicited murine peritoneal macrophages, in addition to STAT-6 and PI3K activation. Furthermore, p38 MAPK inactivation, by gene silencing or pharmacological inhibition, suppressed IL-4-induced typical M2 markers, indicating the involvement of p38 MAPK in the signaling of IL-4 leading to M2-macrophage polarization. Moreover, p38 MAPK inhibition blocked phosphorylation of STAT-6 and Akt, suggesting that p38 MAPK is upstream of these signaling pathways. Finally, we show that in an in vivo model of chitin-induced M2 polarization, p38 MAPK inhibition also diminished activation of M2 markers. Taken together, our data establish a new role for p38 MAPK during IL-4-induced alternative activation of macrophages. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Quercitrin attenuates osteoporosis in ovariectomized rats by regulating mitogen-activated protein kinase (MAPK) signaling pathways.

    Science.gov (United States)

    Xing, Li-Zhi; Ni, Huai-Jun; Wang, Yu-Ling

    2017-03-13

    MAPK signaling pathways are crucial in regulating osteogenesis, a genetic disorder affecting the bones. Quercitrin, a type of flavonoid, is widely distributed in nature and involved in many pharmacological activities. But its osteoprotective functions and mechanism in osteoporosis are far from being understood clearly. In this paper, the MAPK upregulation was observed in the ovariectomy-induced bone loss. Quercitrin was found to downregulate MAPK signaling pathways and prevent the ovariectomy-induced deterioration of bone mineral density (BMD), trabecular microstructure, and bone mechanical characteristics. In this study, quercitrin was seen to prevent the progression of the postmenopausal osteoporosis among the rats, which may be mediated by the downregulated MAPK signaling pathways.

  1. Study of the dung beetle (Coleoptera: Scarabaeidae) community at two sites: Atlantic forest and clear-cut, Pernambuco, Brazil.

    Science.gov (United States)

    Silva, F A B; Costa, C M Q; Moura, R C; Farias, A I

    2010-04-01

    The aim of this study was to compare the dung beetle (Coleoptera, Scarabaeidae: Scarabaeinae) community structure at two sites in the Charles Darwin Ecological Refuge in Igarassu, Pernambuco, Brazil. Dung beetles were collected in 2006 using monthly samples from 48 pitfall traps baited with human dung and bovine carrion. The dung beetle communities from the study sites were compared in terms of abundance, species richness, and diversity (Shannon index). Seasonality was analyzed by Spearman correlation between rainfall data and community parameters. In total, 2,560 individuals belonging to 40 species, 16 genera, and 6 tribes were collected. Species richness was higher for the clear-cut area compared with the forest habitat. Estimators of species richness suggested a total richness of 42-47 species in the entire study area. A positive correlation was observed between monthly rainfall and total abundance of individuals for the clear-cut area but not for the forest habitat. This study contributes to a better understanding of Scarabaeinae ecology in the Atlantic rainforest of northeastern Brazil.

  2. High concentrations and dry deposition of reactive nitrogen species at two sites in the North China Plain

    Energy Technology Data Exchange (ETDEWEB)

    Shen, J.L.; Tang, A.H.; Liu, X.J.; Fangmeier, A.; Goulding, K.T.W.; Zhang, F.S. [China Agricultural University, Beijing (China)

    2009-11-15

    Atmospheric concentrations of major reactive nitrogen (N{sub r}) species were quantified using passive samplers, denuders, and particulate samplers at Dongbeiwang and Quzhou, North China Plain (NCP) in a two-year study. Average concentrations of NH{sub 3}, NO{sub 2}, HNO{sub 3}, pNH{sub 4}{sup +} and pNO{sub 3}{sup -} were 12.0, 12.9, 0.6, 10.3, and 4.7 {mu} g N m{sup -3} across the two sites, showing different seasonal patterns of these N, species. For example, the highest NH{sub 3} concentration occurred in summer while NO{sub 2} concentrations were greater in winter, both of which reflected impacts of N fertilization (summer) and coal-fueled home heating (winter). Based on measured N{sub r} concentrations and their deposition velocities taken from the literature, annual N dry deposition was up to 55 kg N ha{sup -1}. Such high concentrations and deposition rates of N{sub r} species in the NCP indicate very serious air pollution from anthropogenic sources and significant atmospheric N input to crops.

  3. Uric acid stimulates proliferative pathways in vascular smooth muscle cells through the activation of p38 MAPK, p44/42 MAPK and PDGFRβ.

    Science.gov (United States)

    Kırça, M; Oğuz, N; Çetin, A; Uzuner, F; Yeşilkaya, A

    2017-04-01

    Hyperuricemia and angiotensin II (Ang II) may have a pathogenetic role in the development of hypertension and atherosclerosis as well as cardiovascular disease (CVD) and its prognosis. The purpose of this study was to investigate whether uric acid can induce proliferative pathways of vascular smooth muscle cell (VSMC) that are thought to be responsible for the development of CVD. The phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK), p44/42 mitogen-activated protein kinase (p44/42 MAPK) and platelet-derived growth factor receptor β (PDGFRβ) was measured by Elisa and Western blot techniques to determine the activation of proliferative pathways in primary cultured VSMCs from rat aorta. Results demonstrated that uric acid can stimulate p38 MAPK, p44/42 MAPK and PDGFRβ phosphorylation in a time- and concentration-dependent manner. Furthermore, treatment of VSMCs with the angiotensin II type I receptor (AT1R) inhibitor losartan suppressed p38 MAPK and p44/42 MAPK induction by uric acid. The stimulatory effect of uric acid on p38 MAPK was higher compared to that of Ang II. The results of this study show for the first time that uric acid-induced PDGFRβ phosphorylation plays a crucial role in the development of CVDs and that elevated uric acid levels could be a potential therapeutical target in CVD patients.

  4. The inhibition of MAPK potentiates the anti-angiogenic efficacy of mTOR inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Dormond-Meuwly, Anne; Roulin, Didier; Dufour, Marc; Benoit, Michael; Demartines, Nicolas [Department of Visceral Surgery, Centre Hospitalier Universitaire Vaudois and University of Lausanne, Pavillon 3, Av. de Beaumont, 1011 Lausanne (Switzerland); Dormond, Olivier, E-mail: olivier.dormond@chuv.ch [Department of Visceral Surgery, Centre Hospitalier Universitaire Vaudois and University of Lausanne, Pavillon 3, Av. de Beaumont, 1011 Lausanne (Switzerland)

    2011-04-22

    Highlights: {yields} Targeting mTOR in endothelial cell activates MAPK. {yields} Blocking MAPK enhances the anti-angiogenic effects of mTOR inhibitors. {yields} The anti-angiogenic efficacy of ATP-competitive inhibitors of mTOR is superior to that of rapamycin. -- Abstract: The mammalian target of rapamycin (mTOR) which is part of two functionally distinct complexes, mTORC1 and mTORC2, plays an important role in vascular endothelial cells. Indeed, the inhibition of mTOR with an allosteric inhibitor such as rapamycin reduces the growth of endothelial cell in vitro and inhibits angiogenesis in vivo. Recent studies have shown that blocking mTOR results in the activation of other prosurvival signals such as Akt or MAPK which counteract the growth inhibitory properties of mTOR inhibitors. However, little is known about the interactions between mTOR and MAPK in endothelial cells and their relevance to angiogenesis. Here we found that blocking mTOR with ATP-competitive inhibitors of mTOR or with rapamycin induced the activation of the mitogen-activated protein kinase (MAPK) in endothelial cells. Downregulation of mTORC1 but not mTORC2 had similar effects showing that the inhibition of mTORC1 is responsible for the activation of MAPK. Treatment of endothelial cells with mTOR inhibitors in combination with MAPK inhibitors reduced endothelial cell survival, proliferation, migration and tube formation more significantly than either inhibition alone. Similarly, in a tumor xenograft model, the anti-angiogenic efficacy of mTOR inhibitors was enhanced by the pharmacological blockade of MAPK. Taken together these results show that blocking mTORC1 in endothelial cells activates MAPK and that a combined inhibition of MAPK and mTOR has additive anti-angiogenic effects. They also provide a rationale to target both mTOR and MAPK simultaneously in anti-angiogenic treatment.

  5. Measurement of dimeric inhibin using a modified two-site immunoradiometric assay specific for oxidized (Met O) inhibin.

    Science.gov (United States)

    Knight, P G; Muttukrishna, S

    1994-06-01

    Several years ago we developed a novel two-site immunoradiometric assay (IRMA) for dimeric inhibin. However, relative to the purified 32 kDa bovine inhibin standard used at that time, the immunopotencies of crude inhibin-containing samples were much less than their biopotencies estimated by pituitary cell bioassay. In attempting to improve assay performance and resolve this discrepancy we recently discovered that introduction of a preassay oxidation step to the IRMA results in a dramatic increase in the immunopotencies of inhibin-containing test samples (e.g.: bovine, human, porcine follicular fluid (FF)) and of a new (purified in 1993) 32 kDa bovine inhibin standard. However, the oxidation step did not affect the immunopotency of our original standard (purified in 1987), indicating that this material had undergone spontaneous oxidation during long-term storage, thus accounting for its higher immunopotency in our original IRMA and providing an explanation for the discrepancy between immunoactivity and bioactivity referred to above. These findings, together with other observations on the behaviour of oxidized and non-oxidized samples of inhibin, related peptide fragments and inhibin-containing samples in the IRMA and alpha subunit radioimmunoassay (RIA), indicate that the anti-beta A82-114 monoclonal antibody (E4) used as tracer in the IRMA binds selectively to the oxidized (Met O89,91,108) form of the peptide. This property of the antibody can be exploited to advantage by incorporating simple modifications to existing inhibin/activin immunoassays to ensure that all samples and standards are fully oxidized before antibody addition.(ABSTRACT TRUNCATED AT 250 WORDS)

  6. Foliar response of an Ailanthus altissima clone in two sites with different levels of ozone-pollution

    Energy Technology Data Exchange (ETDEWEB)

    Gravano, Elisabetta; Giulietti, Valentina; Desotgiu, Rosanna; Bussotti, Filippo; Grossoni, Paolo; Gerosa, Giacomo; Tani, Corrado

    2003-01-01

    Ozone sensitivity of Ailanthus altissima leaves is due to low leaf density and large intercellular spaces. - Potted plants of Ailanthus altissima, produced by root suckers coming from a single symptomatic mother tree, were placed in two sites in the vicinity of Florence (central Italy), with different levels of ozone pollution. These plants were kept in well watered conditions during the period May-September 1999. In the high pollution site (Settignano-SET) the level of ozone exposure (AOT40) reached at the end of the season a value of 31 ppm h, whereas in the 'low pollution' site (Cascine-CAS) the exposure to ozone was 11 ppm h. A. altissima showed foliar symptoms in early July at SET and in the second half of July at CAS when exposure values reached 5 ppm h at both sites. However, at the end of August the conditions of the plantlets were rather similar in both sites. Microscopic and ultrastructural analysis were performed at the first onset of symptoms at SET (the CAS leaflets were asymptomatic). Observing the upper leaf surface where the brown stipples were visible, it was found that the cells of the palisade mesophyll displayed loss of chlorophyll and the organelles in the cytoplasm were damaged. Swelling of thylacoids was observed in the CAS leaflets, thus indicating the possible onset of a pre-visual damage. The injured cells were separated from the healthy ones by a layer of callose. We conclude that the sensitivity to ozone of A. altissima leaves is related to its leaf structure, with low leaf density and large intercellular spaces. Cell walls, as well as acting as mechanical barriers against the spread of ozone within the cell, also provide important detoxifying processes.

  7. Foliar response of an Ailanthus altissima clone in two sites with different levels of ozone-pollution.

    Science.gov (United States)

    Gravano, Elisabetta; Giulietti, Valentina; Desotgiu, Rosanna; Bussotti, Filippo; Grossoni, Paolo; Gerosa, Giacomo; Tani, Corrado

    2003-01-01

    Potted plants of Ailanthus altissima, produced by root suckers coming from a single symptomatic mother tree, were placed in two sites in the vicinity of Florence (central Italy), with different levels of ozone pollution. These plants were kept in well watered conditions during the period May-September 1999. In the high pollution site (Settignano-SET) the level of ozone exposure (AOT40) reached at the end of the season a value of 31 ppm h, whereas in the "low pollution" site (Cascine-CAS) the exposure to ozone was 11 ppm h. A. altissima showed foliar symptoms in early July at SET and in the second half of July at CAS when exposure values reached 5 ppm h at both sites. However, at the end of August the conditions of the plantlets were rather similar in both sites. Microscopic and ultrastructural analysis were performed at the first onset of symptoms at SET (the CAS leaflets were asymptomatic). Observing the upper leaf surface where the brown stipples were visible, it was found that the cells of the palisade mesophyll displayed loss of chlorophyll and the organelles in the cytoplasm were damaged. Swelling of thylacoids was observed in the CAS leaflets, thus indicating the possible onset of a pre-visual damage. The injured cells were separated from the healthy ones by a layer of callose. We conclude that the sensitivity to ozone of A. altissima leaves is related to its leaf structure, with low leaf density and large intercellular spaces. Cell walls, as well as acting as mechanical barriers against the spread of ozone within the cell, also provide important detoxifying processes.

  8. Efficacy and tolerability of one-site versus two-site phaco-trabeculectomy: a meta-analysis of randomized controlled clinical trials

    Institute of Scientific and Technical Information of China (English)

    LIU He-nan; CHEN Xiao-long; LI Xun; NIE Qing-zhu; ZHU Ying

    2010-01-01

    Background Phacotrabeculectomy can be performed using one-site or two-site incisions.This meta-analysis evaluated the efficacy and tolerability of one-site versus two-site phacotrabecuiectomy in the treatment of patients with coexisting cataract and glaucoma.Methods A comprehensive literature search was performed according to the Cochrane Collaboration methodology toidentify randomized controlled clinical trials comparing one-site with two-site phacotrabeculectomy.Studies meeting our predefined criteria were included in the meta-analysis.Efficacy estimates were measured by weighted mean difference (WMD) for the percentage intraocular pressure (IOP) reduction from baseline to end point, relative risk (RR) for the proportion of patients with a best-corrected visual acuity (BCVA) of 0.5 or better after surgery and complete success rates.Tolerability estimates were measured by RR for adverse events.All of outcomes were reported with 95% confidence interval (95% CI).Data were synthesised by Stata 10.1 for Windows.Results Two-site phacotrabeculectomy was associated with greater reductions in IOP than the one-site procedure (WMD: -5.99, 95% CI: -10.74-1.24, P=0.01).A greater proportion of patients also achieved a BCVA of 0.5 or better (RR:0.91, 95% CI: 0.74-1.12, P=0.36) and the target IOP without anti-glaucoma medication at the study end point (RR: 0.94,95% CI: 0.83-1.07, P=0.34) after two-site than one-site phacotrabeculectomy, but the differences were not significant.There were no significant differences in adverse events between two surgical procedures.Conclusions Two-site phacotrabeculectomy is superior to one-site phacotrabeculectomy in reducing IOP, but other post-operative effects are similar.One-site and two-site phacotrabeculectomies have similar adverse event rates.

  9. EETs mediate cardioprotection of salvianolic acids through MAPK signaling pathway

    Directory of Open Access Journals (Sweden)

    Shoubao Wang

    2013-02-01

    Full Text Available Salvianolic acids, including salvianolic acid A (SAA and salvianolic acid B (SAB, are the main water-soluble bioactive compounds isolated from the Chinese medicinal herb Danshen and have been shown to exert in vitro and in vivo cardiovascular protection. Recent studies suggest that epoxyeicosatrienoic acids (EETs, the primary cytochrome P450 2J (CYP2J epoxygenase metabolites of arachidonic acid, are involved in the progression of ischemic injury in diverse organs. Here, we investigated the relation between the protective effects of salvianolic acids and EETs/sEH as well as MAPK signaling pathway. In the present study, the rat acute myocardial infarction (AMI model was established by the left anterior descending coronary artery occlusion. Our results showed that salvianolic acids significantly reduced ST-segment elevation and serum levels of CK-MB, LDH, and ALT in AMI rats, and significantly attenuated the caspase 3 expression and reduced the ratio of Bax/Bcl-2. ELISA measurement showed that salvianolic acids significantly increased the 14,15-EET levels in blood and heart, and attenuated hydrolase activity of sEH in heart of AMI rat. Western blotting analysis suggested that salvianolic acids significantly attenuated the phosphorylation of JNK and p38, and increased phosphorylation of ERK in heart. In conclusion, these results indicate that EETs/sEH and MAPK signaling pathways are important processes in cardioprotection of salvianolic acids.

  10. Relaxation oscillations and hierarchy of feedbacks in MAPK signaling

    Science.gov (United States)

    Kochańczyk, Marek; Kocieniewski, Paweł; Kozłowska, Emilia; Jaruszewicz-Błońska, Joanna; Sparta, Breanne; Pargett, Michael; Albeck, John G.; Hlavacek, William S.; Lipniacki, Tomasz

    2017-01-01

    We formulated a computational model for a MAPK signaling cascade downstream of the EGF receptor to investigate how interlinked positive and negative feedback loops process EGF signals into ERK pulses of constant amplitude but dose-dependent duration and frequency. A positive feedback loop involving RAS and SOS, which leads to bistability and allows for switch-like responses to inputs, is nested within a negative feedback loop that encompasses RAS and RAF, MEK, and ERK that inhibits SOS via phosphorylation. This negative feedback, operating on a longer time scale, changes switch-like behavior into oscillations having a period of 1 hour or longer. Two auxiliary negative feedback loops, from ERK to MEK and RAF, placed downstream of the positive feedback, shape the temporal ERK activity profile but are dispensable for oscillations. Thus, the positive feedback introduces a hierarchy among negative feedback loops, such that the effect of a negative feedback depends on its position with respect to the positive feedback loop. Furthermore, a combination of the fast positive feedback involving slow-diffusing membrane components with slower negative feedbacks involving faster diffusing cytoplasmic components leads to local excitation/global inhibition dynamics, which allows the MAPK cascade to transmit paracrine EGF signals into spatially non-uniform ERK activity pulses.

  11. KRAS and MAPK1 Gene Amplification in Type II Ovarian Carcinomas

    Directory of Open Access Journals (Sweden)

    Noriyuki Ishikawa

    2013-07-01

    Full Text Available In this study, we examined the clinical significance of KRAS and MAPK1 amplification and assessed whether these amplified genes were potential therapeutic targets in type II ovarian carcinoma. Using fluorescence in situ hybridization, immunohistochemistry, and retrospectively collected clinical data, KRAS and MAPK1 amplifications were identified in 9 (13.2% and 5 (7.4% of 68 type II ovarian carcinoma tissue samples, respectively. Interestingly, co-amplification of KRAS and MAPK1 seemed to be absent in the type II ovarian carcinomas tested, except one case. Active phospho-ERK1/2 was identified in 26 (38.2% out of 68 type II ovarian carcinomas and did not correlate with KRAS or MAPK1 amplification. There was no significant relationship between KRAS amplification and overall or progression-free survival in patients with type II ovarian carcinoma. However, patients with MAPK1 amplification had significantly poorer progression-free survival than patients without MAPK1 amplification. Moreover, type II ovarian carcinoma cells with concomitant KRAS amplification and mutation exhibited dramatic growth reduction following treatment with the MEK inhibitor PD0325901. These findings indicate that KRAS/MAPK1 amplification is critical for the growth of a subset of type II ovarian carcinomas. Additionally, RAS/RAF/MEK/ERK pathway-targeted therapy may benefit selected patients with type II ovarian carcinoma harboring KRAS/MAPK1 amplifications.

  12. Melanoma MAPK pathway proteins and associated tumour suppressors: p16 is an independent prognostic biomarker by tissue microarrays

    DEFF Research Database (Denmark)

    Lade-Keller, Johanne; Guldberg, Per; Hamilton-Dutoit, Stephen Jacques

    2013-01-01

    Melanoma MAPK pathway proteins and associated tumour suppressors: p16 is an independent prognostic biomarker by tissue microarrays......Melanoma MAPK pathway proteins and associated tumour suppressors: p16 is an independent prognostic biomarker by tissue microarrays...

  13. MAPKs and Signal Transduction in the Control of Gastrointestinal Epithelial Cell Proliferation and Differentiation

    Directory of Open Access Journals (Sweden)

    Luciana H. Osaki

    2013-05-01

    Full Text Available Mitogen-activated protein kinase (MAPK pathways are activated by several stimuli and transduce the signal inside cells, generating diverse responses including cell proliferation, differentiation, migration and apoptosis. Each MAPK cascade comprises a series of molecules, and regulation takes place at different levels. They communicate with each other and with additional pathways, creating a signaling network that is important for cell fate determination. In this review, we focus on ERK, JNK, p38 and ERK5, the major MAPKs, and their interactions with PI3K-Akt, TGFβ/Smad and Wnt/β-catenin pathways. More importantly, we describe how MAPKs regulate cell proliferation and differentiation in the rapidly renewing epithelia that lines the gastrointestinal tract and, finally, we highlight the recent findings on nutritional aspects that affect MAPK transduction cascades.

  14. Single-Cell Analysis Reveals that Insulation Maintains Signaling Specificity between Two Yeast MAPK Pathways with Common Components

    OpenAIRE

    Patterson, Jesse C.; Klimenko, Evguenia S.; Thorner, Jeremy

    2010-01-01

    Eukaryotic cells use multiple mitogen-activated protein kinase (MAPK) cascades to evoke appropriate responses to external stimuli. In Saccharomyces cerevisiae, the MAPK Fus3 is activated by pheromone-binding G protein-coupled receptors to promote mating, whereas the MAPK Hog1 is activated by hyperosmotic stress to elicit the high osmolarity glycerol (HOG) response. Although these MAPK pathways share several upstream components, exposure to either pheromone or osmolyte alone triggers only the ...

  15. DMPD: DUSP meet immunology: dual specificity MAPK phosphatases in control of theinflammatory response. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17114416 DUSP meet immunology: dual specificity MAPK phosphatases in control of theinfl...ml) (.csml) Show DUSP meet immunology: dual specificity MAPK phosphatases in control of theinflammatory resp...onse. PubmedID 17114416 Title DUSP meet immunology: dual specificity MAPK phospha...tases in control of theinflammatory response. Authors Lang R, Hammer M, Mages J. Publication J Immunol. 2006

  16. Temporal variation of sandy beach macrofauna at two sites with distinct environmental conditions on Cassino Beach, extreme southern Brazil

    Directory of Open Access Journals (Sweden)

    Pedro de Sá Rodrigues da Silva

    2008-12-01

    Full Text Available Temporal variations of the macrofauna of sandy beaches have been related to variations in the beach morphodynamics and also to the population dynamics of dominant species. The aim of this article is to describe the temporal variation of the intertidal macrofauna at two sites with distinct environmental condition on Cassino Beach, extreme southern Brazil. At each site three transect lines 50 m apart were defined perpendicular to the shore line, from which samples were collected monthly in triplicate at 4 intertidal levels (10 m apart from June 2004 to May 2005. During winter a generally low density was observed, due to the absence of recruitments and to the mud deposition, which occurred just before sampling (in April 2004, and to low intensity stranding events. Spring witnessed a population explosion of Scolelepis gaucha, a migration of Mesodesma mactroides adults from the subtidal zone, and a strong stranding event. In the summer, recruitment of M. mactroides, Donax hanleyanus and Emerita brasiliensis was observed. Fall was characterized by low densities, except for D. hanleyanus recruitment. The macrofauna at both sites showed a striking seasonal variation in density and diversity, perhaps attributable to the recruitment of numerically dominant species and physical disturbances (stranding and mud deposition.Variações sazonais da macrofauna bentônica de praias arenosas têm sido relacionadas com variações da morfodinâmica da praia e também aos recrutamentos das espécies dominantes. Este trabalho objetiva avaliar a variabilidade temporal da macrofauna da zona entremarés de dois locais com distintas características ambientais na praia do Cassino, extremo sul do Brasil. Em cada local foram demarcadas três transversais (separadas por 50m perpendiculares à linha de água, nas quais amostras foram coletadas em triplicata em 4 níveis entremarés (separados por 10 m, entre junho/2004 e maio/2005. Durante o inverno ocorreram baixas

  17. Compressive stress induces dephosphorylation of the myosin regulatory light chain via RhoA phosphorylation by the adenylyl cyclase/protein kinase A signaling pathway.

    Directory of Open Access Journals (Sweden)

    Kenji Takemoto

    Full Text Available Mechanical stress that arises due to deformation of the extracellular matrix (ECM either stretches or compresses cells. The cellular response to stretching has been actively studied. For example, stretching induces phosphorylation of the myosin regulatory light chain (MRLC via the RhoA/RhoA-associated protein kinase (ROCK pathway, resulting in increased cellular tension. In contrast, the effects of compressive stress on cellular functions are not fully resolved. The mechanisms for sensing and differentially responding to stretching and compressive stress are not known. To address these questions, we investigated whether phosphorylation levels of MRLC were affected by compressive stress. Contrary to the response in stretching cells, MRLC was dephosphorylated 5 min after cells were subjected to compressive stress. Compressive loading induced activation of myosin phosphatase mediated via the dephosphorylation of myosin phosphatase targeting subunit 1 (Thr853. Because myosin phosphatase targeting subunit 1 (Thr853 is phosphorylated only by ROCK, compressive loading may have induced inactivation of ROCK. However, GTP-bound RhoA (active form increased in response to compressive stress. The compression-induced activation of RhoA and inactivation of its effector ROCK are contradictory. This inconsistency was due to phosphorylation of RhoA (Ser188 that reduced affinity of RhoA to ROCK. Treatment with the inhibitor of protein kinase A that phosphorylates RhoA (Ser188 induced suppression of compression-stimulated MRLC dephosphorylation. Incidentally, stretching induced phosphorylation of MRLC, but did not affect phosphorylation levels of RhoA (Ser188. Together, our results suggest that RhoA phosphorylation is an important process for MRLC dephosphorylation by compressive loading, and for distinguishing between stretching and compressing cells.

  18. PDH-E1alpha dephosphorylation and activation in human skeletal muscle during exercise: effect of intralipid infusion.

    Science.gov (United States)

    Pilegaard, Henriette; Birk, Jesper B; Sacchetti, Massimo; Mourtzakis, Marina; Hardie, D Graham; Stewart, Greg; Neufer, P Darrell; Saltin, Bengt; van Hall, Gerrit; Wojtaszewski, Jorgen F P

    2006-11-01

    To investigate pyruvate dehydrogenase (PDH)-E1alpha subunit phosphorylation and whether free fatty acids (FFAs) regulate PDH activity, seven subjects completed two trials: saline (control) and intralipid/heparin (intralipid). Each infusion trial consisted of a 4-h rest followed by a 3-h two-legged knee extensor exercise at moderate intensity. During the 4-h resting period, activity of PDH in the active form (PDHa) did not change in either trial, yet phosphorylation of PDH-E1alpha site 1 (PDH-P1) and site 2 (PDH-P2) was elevated in the intralipid compared with the control trial. PDHa activity increased during exercise similarly in the two trials. After 3 h of exercise, PDHa activity remained elevated in the intralipid trial but returned to resting levels in the control trial. Accordingly, in both trials PDH-P1 and PDH-P2 decreased during exercise, and the decrease was more marked during intralipid infusion. Phosphorylation had returned to resting levels at 3 h of exercise only in the control trial. Thus, an inverse association between PDH-E1alpha phosphorylation and PDHa activity exists. Short-term elevation in plasma FFA at rest increases PDH-E1alpha phosphorylation, but exercise overrules this effect of FFA on PDH-E1alpha phosphorylation leading to even greater dephosphorylation during exercise with intralipid infusion than with saline.

  19. Phosphoproteomics Reveals Regulatory T Cell-Mediated DEF6 Dephosphorylation That Affects Cytokine Expression in Human Conventional T Cells

    KAUST Repository

    Joshi, Rubin N.

    2017-09-25

    Regulatory T cells (Tregs) control key events of immune tolerance, primarily by suppression of effector T cells. We previously revealed that Tregs rapidly suppress T cell receptor (TCR)-induced calcium store depletion in conventional CD4CD25 T cells (Tcons) independently of IP levels, consequently inhibiting NFAT signaling and effector cytokine expression. Here, we study Treg suppression mechanisms through unbiased phosphoproteomics of primary human Tcons upon TCR stimulation and Treg-mediated suppression, respectively. Tregs induced a state of overall decreased phosphorylation as opposed to TCR stimulation. We discovered novel phosphosites (T595_S597) in the DEF6 (SLAT) protein that were phosphorylated upon TCR stimulation and conversely dephosphorylated upon coculture with Tregs. Mutation of these DEF6 phosphosites abrogated interaction of DEF6 with the IP receptor and affected NFAT activation and cytokine transcription in primary Tcons. This novel mechanism and phosphoproteomics data resource may aid in modifying sensitivity of Tcons to Treg-mediated suppression in autoimmune disease or cancer.

  20. PRL-3 Promotes the Malignant Progression of Melanoma via Triggering Dephosphorylation and Cytoplasmic Localization of NHERF1.

    Science.gov (United States)

    Fang, Xian-Ying; Song, Ran; Chen, Wei; Yang, Yuan-Yuan; Gu, Yan-Hong; Shu, Yong-Qian; Wu, Xu-Dong; Wu, Xue-Feng; Sun, Yang; Shen, Yan; Xu, Qiang

    2015-09-01

    Phosphatase of regenerating liver-3 (PRL-3) has been reported to have a critical role in metastatic progression of cancers. Here, we investigate how PRL-3 increases the malignant degree of melanoma cells. The expression of PRL-3 increased gradually during the malignant progression of melanoma. The phosphorylation of Akt was elevated in highly malignant melanoma cells, which was accompanied by a decrease in nuclear phosphatase and tensin homolog (PTEN). The phosphorylation of NHERF1 in the serine site was regulated by PRL-3 and showed cytoplasmic translocation upon dephosphorylation, which resulted in a decrease in nuclear PTEN. The co-translocation of NHERF1 and PTEN from the nucleus to the cytoplasm was observed during the malignant progression of melanoma cells. Tumor growth was inhibited significantly, and the survival was prolonged upon knockdown of cytoplasmic NHERF1 in B16BL6 cells prior to the inoculation into mice. Taken together, to our knowledge previously unreported, we have identified NHERF1 as a potential substrate of PRL-3. Its phosphorylation status as well as its change in cellular localization and association with PTEN correlated with the malignant progression of melanoma. Our data provide an explanation for how PRL-3 promotes the malignant progression of melanoma, as well as a diagnostic marker or therapeutic target for malignant melanoma.

  1. Dipalmitoleoylphosphoethanolamine as a PP2A Enhancer Obstructs Insulin Signaling by Promoting Ser/Thr Dephosphorylation of Akt

    Directory of Open Access Journals (Sweden)

    Ayako Tsuchiya

    2014-08-01

    Full Text Available Background/Aims: The phospholipid phosphatidylethanolamine is implicated in the regulation of a variety of cellular processes. The present study investigated the effect of phosphatidylethanolamines such as 1,2-diarachidonoyl-sn-glycero-3-phosphoethanolamine (DAPE, 1,2-dilinoleoyl-sn-glycero-3-phosphoethanolamine (DLPE, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE, and 1,2-dipalmitoleoyl-sn-glycero-3-phosphoethanolamine (DPPE on protein phosphatases, Akt1/2 activity, GLUT4 mobilizations, and glucose uptake into cells. Methods: Activity of protein phosphatase 2A (PP2A was assayed under the cell-free conditions, and Western blotting, intracellular GLUT4 trafficking, and glucose uptake into cells were monitored using differentiated 3T3-L1-GLUT4myc adipocytes. Results: Of the investigated phosphatidylethanolamines, DLPE and DPPE significantly enhanced PP2A activity. DPPE inhibited insulin-induced phosphorylation of Akt1/2 at Thr308/309 and Ser473/474 in differentiated 3T3-L1-GLUT4myc adipocytes. DPPE also inhibited insulin-stimulated GLUT4 translocation to the cell surface and reduced insulin-stimulated glucose uptake into adipocytes. Conclusion: The results of the present study indicate that the PP2A enhancer DPPE obstructs insulin signaling by promoting serine/threonine dephosphorylation of Akt1/2, resulting in the suppression of GLUT4 translocation to the cell surface and glucose uptake into adipocytes.

  2. p38 MAPK inhibition alleviates experimental acute pancreatitis in mice

    Institute of Scientific and Technical Information of China (English)

    Ming-HuaCao; JingXu; Hai-DongCai; Zhong-WeiLv; Ya-JingFeng; KunLi; Chun-QiuChen; Yong-YuLi

    2015-01-01

    BACKGROUND: The mitogen-activated protein kinases (MAPKs) signaling pathway is involved in inflammatory process. However, the mechanism is not clear. The present study was to investi-gate the role of p38 MAPK in acute pancreatitis in mice. METHODS: Mice were divided into 4 groups: saline control; acute  pancreatitis  induced  with  repeated  injections  of  ceru-lein;  control  plus  p38  MAPK  inhibitor  SB203580;  and  acute pancreatitis plus SB203580. The pancreatic histology, pancre-atic enzymes, cytokines, myeloperoxidase activity, p38 MAPK and heat shock protein (HSP) 60 and 70 were evaluated. RESULTS: Repeated  injections  of  cerulein  resulted  in  acute pancreatitis  in  mice,  accompanying  with  the  activation  of p38  MAPK  and  overexpression  of  HSP60  and  HSP70  in  the pancreatic tissues. Treatment with SB203580 significantly in-hibited the activation of p38 MAPK, and furthermore, inhib-ited the expression of HSP60 and HSP70 in the pancreas, the inflammatory  cytokines  in  the  serum,  and  myeloperoxidase activity in the lung. CONCLUSION: The p38 MAPK signaling pathway is involved in  the  regulation  of  inflammatory  response  and  the  expres-sion of HSP60 and HSP70 in acute pancreatitis.

  3. Involvement of ATM/ATR-p38 MAPK cascade in MNNG induced G1-S arrest

    Institute of Scientific and Technical Information of China (English)

    Ke-Qing Zhu; Suo-Jiang Zhang

    2003-01-01

    AIM: To understand the effect of low concentration of Nmethyl-N′-nitro-nitrosoguanidine (MNNG), which is a widely distributed environmental mutagen and carcinogen especially for human gastric cancer, on DNA damage and to study its possible pathway in regulating cell cycle arrest.METHODS: The DNA damage effect was measured by Comet assay. A specific phospho-(Ser/Thr) ATM/ATR substrate antibody was used to detect the damage sensor by Western blot. p38 kinase activity was measured by direct kinase assay,and immunoprecipitation for the possible connection between ATM/ATR and p38 MAPK. Flow cytometry analysis and p38MAPK specific inhibitor SB203580 were combined to detect the possible cell cycle arrest by p38 MAPK.RESULTS: With the same low concentration MNNG exposure (0.2μM 2.5 h), Comet assays indicated that strand breaks accumulated, Western blot and kinase assay showed ATM/ATR and p38 kinase activated, immunoprecipitation showed phospho-ATM/ATR substrate antibody combined with both p38 MAPK antibody and phospho-p38 MAPK antibody, p38MAPK pathway was involved in the G1-S arrest.CONCLUSION: Activation of ATM/ATR by MNNG induced DNA damage leads to activation of p38 MAPK, which involves in the G1 checkpoint in mammalian cells.

  4. Context-Dependent Effects of Amplified MAPK Signaling during Lung Adenocarcinoma Initiation and Progression

    Directory of Open Access Journals (Sweden)

    Michelle Cicchini

    2017-02-01

    Full Text Available Expression of oncogenic KrasG12D initiates lung adenomas in a mitogen-activated protein kinase (MAPK signal-dependent manner from only a subset of cell types in the adult mouse lung. Amplification of MAPK signaling is associated with progression to malignant adenocarcinomas, but whether this is a cause or a consequence of disease progression is not known. To better understand the effects of MAPK signaling downstream of KrasG12D expression, we capitalized on the ability of Braf inhibition to selectively amplify MAPK pathway signaling in KrasG12D-expressing epithelial cells. MAPK signal amplification indeed promoted the rapid progression of established adenomas to malignant adenocarcinomas. However, we observed, surprisingly, a greater number of overall tumor-initiating events after MAPK signal amplification, due to induced proliferation of cell types that are normally refractory to KrasG12D-induced transformation. Thus, MAPK signaling in the lung is thresholded not only during malignant progression but also at the moment of tumor initiation.

  5. Impact of MAPK pathway activation in BRAFV600 melanoma on T cell and Dendritic Cell function

    Directory of Open Access Journals (Sweden)

    Patrick Alexander Ott

    2013-10-01

    Full Text Available Constitutive upregulation of the MAPK pathway by a BRAFV600 mutation occurs in about half of melanomas. This leads to increased oncogenic properties such as tumor cell invasion, metastatic potential, and resistance to apoptosis. Blockade of the MAPK pathway with highly specific kinase inhibitors induces unprecedented tumor response rates in patients with advanced BRAFV600 mutant melanoma. Immune checkpoint blockade with monoclonal antibodies targeting CTLA-4 and PD-1/PD-L1 has also demonstrated striking anti-tumor activity in patients with advanced melanoma. Tumor responses are likely limited by multiple additional layers of immune suppression in the tumor microenvironment. There is emerging preclinical and clinical evidence suggesting that MAPK inhibition has a beneficial effect on the immunosuppressive tumor microenvironment, providing a strong rationale for combined immunotherapy and MAPK pathway inhibition in melanoma. The T cell response has been the main focus in the studies reported to date. Since dendritic cells (DCs are important in the induction of tumor-specific T cell responses, the impact of MAPK pathway activation in melanoma on DC function is critical for the melanoma directed immune response. BRAFV600E melanoma cells modulate DC through the MAPK pathway because its blockade in melanoma cells can reverse suppression of DC function. As both MEK/BRAF inhibition and immune checkpoint blockade have recently taken center stage in the treatment of melanoma, a deeper understanding of how MAPK pathway inhibition affects the tumor immune response is needed.

  6. Role of amygdala MAPK activation on immobility behavior of forced swim rats.

    Science.gov (United States)

    Huang, Tung-Yi; Lin, Chih-Hung

    2006-10-01

    The role of amygdala mitogen-activated protein kinase (MAPK) in rats during a forced swim test was investigated. The variation of amygdala MAPK level was studied in control rats and early-life maternally deprived rats. A forced swim test was carried out to estimate the immobility level. The data showed that the immobility time of rats that received maternal deprivation in early life was longer than that of control rats and Western blot analysis also showed that the amygdala phospho-MAPK level in maternally deprived rats was almost two times higher than in control rats. Intra-amygdala infusion of PD098059 or U0126, MEK inhibitors, suppressed immobility behavior during the forced swim test in both rats. Western blot analysis also showed that the amygdala MAPK activities in both rats infused with MEK inhibitors were also suppressed in parallel with expression of immobility behavior. The suppressed MAPK activities as well as the restoration of immobility time returned to the original level 48 h later. These results suggest that amygdala MAPK activation might play a role in the regulation of immobility behavior in rats during the forced swim test. Moreover, it could provide a hint that amygdala MAPK activation might be involved in the formation of depression-like behavior.

  7. Impact of MAPK Pathway Activation in BRAFV600 Melanoma on T Cell and Dendritic Cell Function

    Directory of Open Access Journals (Sweden)

    Patrick A. Ott

    2013-10-01

    Full Text Available Constitutive upregulation of the MAPK pathway by a BRAFV600 mutation occurs in about half of melanomas. This leads to increased oncogenic properties such as tumor cell invasion, metastatic potential, and resistance to apoptosis. Blockade of the MAPK pathway with highly specific kinase inhibitors induces unprecedented tumor response rates in patients with advanced BRAFV600 mutant melanoma. Immune checkpoint blockade with monoclonal antibodies targeting cytotoxic T-lymphocyte antigen 4 and programed death-1/PD-L1 has also demonstrated striking anti-tumor activity in patients with advanced melanoma. Tumor responses are likely limited by multiple additional layers of immune suppression in the tumor microenvironment. There is emerging preclinical and clinical evidence suggesting that MAPK inhibition has a beneficial effect on the immunosuppressive tumor microenvironment, providing a strong rationale for combined immunotherapy and MAPK pathway inhibition in melanoma. The T cell response has been the main focus in the studies reported to date. Since dendritic cells (DCs are important in the induction of tumor-specific T cell responses, the impact of MAPK pathway activation in melanoma on DC function is critical for the melanoma directed immune response. BRAFV600E melanoma cells modulate DCs through the MAPK pathway because its blockade in melanoma cells can reverse suppression of DC function. As both MEK/BRAF inhibition and immune checkpoint blockade have recently taken center stage in the treatment of melanoma, a deeper understanding of how MAPK pathway inhibition affects the tumor immune response is needed.

  8. Use of the Performance Diagnostic Checklist to Select an Intervention Designed to Increase the Offering of Promotional Stamps at Two Sites of a Restaurant Franchise

    Science.gov (United States)

    Rodriguez, Manuel; Wilder, David A.; Therrien, Kelly; Wine, Byron; Miranti, Reylissa; Daratany, Kenneth; Salume, Gloria; Baranovsky, Greg; Rodriquez, Matias

    2006-01-01

    The performance diagnostic checklist (PDC) was administered to examine the variables influencing the offering of promotional stamps by employees at two sites of a restaurant franchise. PDC results suggested that a lack of appropriate antecedents, equipment and processes, and consequences were responsible for the deficits. Based on these results,…

  9. Involvement of AP-1 in p38MAPK signaling pathway in osteoblast apoptosis induced by high glucose.

    Science.gov (United States)

    Feng, Z P; Deng, H C; Jiang, R; Du, J; Cheng, D Y

    2015-04-10

    We investigated the effect of p38MAPK/AP-1 (activator protein-1) signaling on the apoptosis of osteoblasts induced by high glucose. A lentivirus vector of small hairpin RNA (shRNA) targeting p38MAPK was constructed in vitro. Osteoblasts MC3T3-E1 cultured in vitro were treated with vehicle, high glucose, p38MAPK-shRNA transfection, p38MAPK inhibitor, and unrelated shRNA transfection. Apoptosis, protein levels of p38MAPK, and activities of AP-1 in MC3T3-E1 osteoblasts were measured using TUNEL and flow cytometry, Western blot analysis, and an electrophoretic mobility shift assay. Compared with the vehicle group, high glucose induced apoptosis of MC3T3-E1 osteoblasts and activated p38MAPK and AP-1. p38MAPK-shRNA transfection blocked the effect of high glucose stimulation, and the p38MAPK inhibitor showed similar effects as those observed in p38MAPK transfection. Unrelated shRNA had no effect on these changes in MC3T3-E1 osteoblasts induced by high glucose. Therefore, our results suggest that p38MAPK-shRNA reduce apoptosis of MC3T3-E1 osteoblasts induced by high glucose by inhibiting the p38MAPK-AP-1 signaling pathway.

  10. Ancient signals: comparative genomics of plant MAPK and MAPKK gene families

    DEFF Research Database (Denmark)

    Hamel, Louis-Philippe; Nicole, Marie-Claude; Sritubtim, Somrudee;

    2006-01-01

    MAPK signal transduction modules play crucial roles in regulating many biological processes in plants, and their components are encoded by highly conserved genes. The recent availability of genome sequences for rice and poplar now makes it possible to examine how well the previously described...... Arabidopsis MAPK and MAPKK gene family structures represent the broader evolutionary situation in plants, and analysis of gene expression data for MPK and MKK genes in all three species allows further refinement of those families, based on functionality. The Arabidopsis MAPK nomenclature appears sufficiently...... robust to allow it to be usefully extended to other well-characterized plant systems....

  11. Decursin Isolated from Angelica gigas Nakai Rescues PC12 Cells from Amyloid β-Protein-Induced Neurotoxicity through Nrf2-Mediated Upregulation of Heme Oxygenase-1: Potential Roles of MAPK

    Directory of Open Access Journals (Sweden)

    Li Li

    2013-01-01

    Full Text Available Decursin (D, purified from Angelica gigas Nakai, has been proven to exert neuroprotective property. Previous study revealed that D reduced Aβ25‒35-induced cytotoxicity in PC12 cells. Our study explored the underlying mechanisms by which D mediates its therapeutic effects in vitro. Pretreatment of cells with D diminished intracellular generation of ROS in response to Aβ25‒35. Western blot revealed that D significantly increased the expression and activity of HO-1, which was correlated with its protection against Aβ25‒35-induced injury. Addition of ZnPP, an HO-1 competitive inhibitor, significantly attenuated its protective effect in Aβ25‒35-treated cells, indicating the vital role of HO-1 resistance to oxidative injury. Moreover, D induced Nrf2 nuclear translocation, the upstream of HO-1 expression. While investigating the signaling pathways responsible for HO-1 induction, D activated ERK and dephosphorylated p38 in PC12 cells. Addition of U0126, a selective inhibitor of ERK, blocked D-induced Nrf2 activation and HO-1 induction and meanwhile reversed the protection of D against Aβ25‒35-induced cell death. These findings suggest D augments cellular antioxidant defense capacity through both intrinsic free radical scavenging activity and activation of MAPK signal pathways that leads to Nrf2 activation, and subsequently HO-1 induction, thereby protecting the PC12 cells from Aβ25‒35-induced oxidative cytotoxicity.

  12. A novel effect of MARCKS phosphorylation by activated PKC: the dephosphorylation of its serine 25 in chick neuroblasts.

    Directory of Open Access Journals (Sweden)

    Andrea Toledo

    Full Text Available MARCKS (Myristoylated Alanine-Rich C Kinase Substrate is a peripheral membrane protein, especially abundant in the nervous system, and functionally related to actin organization and Ca-calmodulin regulation depending on its phosphorylation by PKC. However, MARCKS is susceptible to be phosphorylated by several different kinases and the possible interactions between these phosphorylations have not been fully studied in intact cells. In differentiating neuroblasts, as well as some neurons, there is at least one cell-type specific phosphorylation site: serine 25 (S25 in the chick. We demonstrate here that S25 is included in a highly conserved protein sequence which is a Cdk phosphorylatable region, located far away from the PKC phosphorylation domain. S25 phosphorylation was inhibited by olomoucine and roscovitine in neuroblasts undergoing various states of cell differentiation in vitro. These results, considered in the known context of Cdks activity in neuroblasts, suggest that Cdk5 is the enzyme responsible for this phosphorylation. We find that the phosphorylation by PKC at the effector domain does not occur in the same molecules that are phosphorylated at serine 25. The in situ analysis of the subcellular distribution of these two phosphorylated MARCKS variants revealed that they are also segregated in different protein clusters. In addition, we find that a sustained stimulation of PKC by phorbol-12-myristate-13-acetate (PMA provokes the progressive disappearance of phosphorylation at serine 25. Cells treated with PMA, but in the presence of several Ser/Thr phosphatase (PP1, PP2A and PP2B inhibitors indicated that this dephosphorylation is achieved via a phosphatase 2A (PP2A form. These results provide new evidence regarding the existence of a novel consequence of PKC stimulation upon the phosphorylated state of MARCKS in neural cells, and propose a link between PKC and PP2A activity on MARCKS.

  13. MKP1 phosphatase mediates G1-specific dephosphorylation of H3Serine10P in response to DNA damage

    Energy Technology Data Exchange (ETDEWEB)

    Sharma, Ajit K.; Khan, Shafqat A.; Sharda, Asmita; Reddy, Divya V; Gupta, Sanjay, E-mail: sgupta@actrec.gov.in

    2015-08-15

    Highlights: • Reversible reduction of H3S10 phosphorylation after DNA damage is G1 phase specific. • Dynamic balance between MAP kinases, MKP1 and MSK1 regulate H3S10P during DDR. • MKP1 associates with chromatin bearing γH2AX in response to DNA damage. • Inhibition of MKP1 activity with specific inhibitor promotes radiation-induced cell death. - Abstract: Histone mark, H3S10 phosphorylation plays a dual role in a cell by maintaining relaxed chromatin for active transcription in interphase and condensed chromatin state in mitosis. The level of H3S10P has also been shown to alter on DNA damage; however, its cell cycle specific behavior and regulation during DNA damage response is largely unexplored. In the present study, we demonstrate G1 cell cycle phase specific reversible loss of H3S10P in response to IR-induced DNA damage is mediated by opposing activities of phosphatase, MKP1 and kinase, MSK1 of the MAP kinase pathway. We also show that the MKP1 recruits to the chromatin in response to DNA damage and correlates with the decrease of H3S10P, whereas MKP1 is released from chromatin during recovery phase of DDR. Furthermore, blocking of H3S10 dephosphorylation by MKP1 inhibition impairs DNA repair process and results in poor survival of WRL68 cells. Collectively, our data proposes a pathway regulating G1 cell cycle phase specific reversible reduction of H3S10P on IR induced DNA damage and also raises the possibility of combinatorial modulation of H3S10P with specific inhibitors to target the cancer cells in G1-phase of cell cycle.

  14. The MAP(K) of fear: from memory consolidation to memory extinction.

    Science.gov (United States)

    Cestari, Vincenzo; Rossi-Arnaud, Clelia; Saraulli, Daniele; Costanzi, Marco

    2014-06-01

    The highly conserved mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) signaling cascade is involved in several intracellular processes ranging from cell differentiation to proliferation, as well as in synaptic plasticity. In the last two decades, the role of MAPK/ERK in long-term memory formation in mammals, particularly in fear-related memories, has been extensively investigated. In this review we describe knowledge advancement on the role of MAPK/ERK in orchestrating the intracellular processes that lead to the consolidation, reconsolidation and extinction of fear memories. In doing so, we report studies in which the specific role of MAP/ERK in switching from memory formation to memory erasure has been suggested. The possibility to target MAPK/ERK in developing and/or refining pharmacological approaches to treat psychiatric disorders in which fear regulation is defective has also been envisaged. Copyright © 2013 Elsevier Inc. All rights reserved.

  15. Interferon alpha regulates MAPK and STAT1 pathways in human hepatoma cells

    Directory of Open Access Journals (Sweden)

    Ren Hao

    2011-04-01

    Full Text Available Abstract Background Signaling events triggered by interferon (IFN account for the molecular mechanisms of antiviral effect. JAK-STAT pathway plays a critical role in IFN signaling, and other pathways are also implicated in IFN-mediated antiviral effect. Changes in mitogen-activated protein kinase (MAPK and STAT1 pathways were evaluated in human hepatoma cells Huh7 and HepG2 upon IFN alpha treatment. Results Phosphorylation of ERK was significantly and specifically up-regulated, whereas enhanced phosphorylation of upstream kinase MEK was unobservable upon IFN alpha treatment. A mild increase in p38 MAPK, SAPK/JNK and downstream target ATF-2 phosphorylation was detectable after exposure to IFN alpha, indicating differential up-regulation of the MAPK signaling cascades. Moreover, STAT1 phosphorylation was strongly enhanced by IFN alpha. Conclusion IFN alpha up-regulates MAPK and STAT1 pathways in human hepatoma cells, and may provide useful information for understanding the IFN signaling.

  16. Genome-wide survey of yeast mutations leading to activation of the yeast cell integrity MAPK pathway: Novel insights into diverse MAPK outcomes

    Directory of Open Access Journals (Sweden)

    Arias Patricia

    2011-08-01

    Full Text Available Abstract Background The yeast cell wall integrity mitogen-activated protein kinase (CWI-MAPK pathway is the main regulator of adaptation responses to cell wall stress in yeast. Here, we adopt a genomic approach to shed light on two aspects that are only partially understood, namely, the characterization of the gene functional catalog associated with CWI pathway activation and the extent to which MAPK activation correlates with transcriptional outcomes. Results A systematic yeast mutant deletion library was screened for constitutive transcriptional activation of the CWI-related reporter gene MLP1. Monitoring phospho-Slt2/Mpk1 levels in the identified mutants revealed sixty-four deletants with high levels of phosphorylation of this MAPK, including mainly genes related to cell wall construction and morphogenesis, signaling, and those with unknown function. Phenotypic analysis of the last group of mutants suggests their involvement in cell wall homeostasis. A good correlation between levels of Slt2 phosphorylation and the magnitude of the transcriptional response was found in most cases. However, the expression of CWI pathway-related genes was enhanced in some mutants in the absence of significant Slt2 phosphorylation, despite the fact that functional MAPK signaling through the pathway was required. CWI pathway activation was associated to increased deposition of chitin in the cell wall - a known survival compensatory mechanism - in about 30% of the mutants identified. Conclusion We provide new insights into yeast genes related to the CWI pathway and into how the state of activation of the Slt2 MAPK leads to different outcomes, discovering the versatility of this kind of signaling pathways. These findings potentially have broad implications for understanding the functioning of other eukaryotic MAPKs.

  17. Aldosterone-induced brain MAPK signaling and sympathetic excitation are angiotensin II type-1 receptor dependent.

    Science.gov (United States)

    Zhang, Zhi-Hua; Yu, Yang; Wei, Shun-Guang; Felder, Robert B

    2012-02-01

    Angiotensin II (ANG II)-induced mitogen-activated protein kinase (MAPK) signaling upregulates angiotensin II type-1 receptors (AT(1)R) in hypothalamic paraventricular nucleus (PVN) and contributes to AT(1)R-mediated sympathetic excitation in heart failure. Aldosterone has similar effects to increase AT(1)R expression in the PVN and sympathetic drive. The present study was undertaken to determine whether aldosterone also activates the sympathetic nervous system via MAPK signaling and, if so, whether its effect is independent of ANG II and AT(1)R. In anesthetized rats, a 4-h intravenous infusion of aldosterone induced increases (P < 0.05) in phosphorylated (p-) p44/42 MAPK in PVN, PVN neuronal excitation, renal sympathetic nerve activity (RSNA), mean blood pressure (MBP), and heart rate (HR). Intracerebroventricular or bilateral PVN microinjection of the p44/42 MAPK inhibitor PD-98059 reduced the aldosterone-induced RSNA, HR, and MBP responses. Intracerebroventricular pretreatment (5 days earlier) with pooled small interfering RNAs targeting p44/42 MAPK reduced total and p-p44/42 MAPK, aldosterone-induced c-Fos expression in the PVN, and the aldosterone-induced increases in RSNA, HR, and MBP. Intracerebroventricular infusion of either the mineralocorticoid receptor antagonist RU-28318 or the AT(1)R antagonist losartan blocked aldosterone-induced phosphorylation of p44/42 MAPK and prevented the increases in RSNA, HR, and MBP. These data suggest that aldosterone-induced sympathetic excitation depends upon that AT(1)R-induced MAPK signaling in the brain. The short time course of this interaction suggests a nongenomic mechanism, perhaps via an aldosterone-induced transactivation of the AT(1)R as described in peripheral tissues.

  18. Involvement of MAPK proteins in bystander effects induced by chemicals and ionizing radiation

    Energy Technology Data Exchange (ETDEWEB)

    Asur, Rajalakshmi [Department of Biological Sciences, Wayne State University, 5047 Gullen Mall, Suite 1370, Detroit, MI 48202-3917 (United States); Balasubramaniam, Mamtha [Research Institute - Biostatistics, William Beaumont Hospital, 3911 W. Thirteen Mile Road, Royal Oak, MI 48073 (United States); Marples, Brian [Department of Radiation Oncology, William Beaumont Hospital, 3811 W. Thirteen Mile Road, Royal Oak, MI 48073 (United States); Thomas, Robert A. [Department of Biological Sciences, Wayne State University, 5047 Gullen Mall, Suite 1370, Detroit, MI 48202-3917 (United States); Tucker, James D., E-mail: jtucker@biology.biosci.wayne.edu [Department of Biological Sciences, Wayne State University, 5047 Gullen Mall, Suite 1370, Detroit, MI 48202-3917 (United States)

    2010-04-01

    Many studies have examined bystander effects induced by ionizing radiation, however few have evaluated the ability of chemicals to induce similar effects. We previously reported the ability of two chemicals, mitomycin C (MMC) and phleomycin (PHL) to induce bystander effects in normal human lymphoblastoid cell lines. The focus of the current study was to determine the involvement of the MAPK proteins in bystander effects induced by physical and chemical DNA damaging agents and to evaluate the effects of MAPK inhibition on bystander-induced caspase 3/7 activation. The phosphorylation levels of the MAPK proteins ERK1/2, JNK, and p38, were measured from 1 to 24 h following direct or bystander exposure to MMC, PHL or radiation. We observed transient phosphorylation, at early time points, of all 3 proteins in bystander cells. We also evaluated the effect of MAPK inhibition on bystander-induced caspase 3/7 activity to determine the role of MAPK proteins in bystander-induced apoptosis. We observed bystander-induced activation of caspase 3/7 in bystander cells. Inhibition of MAPK proteins resulted in a decrease in caspase 3/7 activity at the early time points, and the caspase activity increased (in the case of ERK inhibition) or returned to basal levels (in the case of JNK or p38 inhibition) between 12 and 24 h. PHL is considered to be a radiomimetic agent, however in the present study PHL behaved more like a chemical and not like radiation in terms of MAPK phosphorylation. These results point to the involvement of MAPK proteins in the bystander effect induced by radiation and chemicals and provide additional evidence that this response is not limited to radiation but is a generalized stress response in cells.

  19. Essential roles of Cdc42 and MAPK in cadmium-induced apoptosis in Litopenaeus vannamei

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Ting; Wang, Wei-Na, E-mail: weina63@aliyun.com; Gu, Mei-Mei; Xie, Chen-Ying; Xiao, Yu-Chao; Liu, Yuan; Wang, Lei

    2015-06-15

    Highlights: • Cd{sup 2+} induces Cdc42 and MAPKs pathway related gene of Litopenaeus vannamei up-regulation. • Reduction of THC, increase of ROS production and apoptotic cell rate were observed when the shrimps exposure to Cd{sup 2+}. • DsRNA-suppression of LvCdc42 and MAPKs during Cd{sup 2+} stress reduces the ROS production and apoptosis. • We conclude that LvCdc42 and MAPKs play key roles in Cd{sup 2+} stress responses of shrimps. - Abstract: Cadmium, one of the most toxic heavy metals in aquatic environments, has severe effects on marine invertebrates and fishes. The MAPK signaling pathway plays a vital role in stress responses of animals. The mitogen-activated protein kinase (MAPK) signaling pathway plays a vital role in animals’ stress responses, including mediation of apoptosis induced by the Rho GTPase Cdc42. However, there is limited knowledge about its function in shrimps, although disorders exacerbated by environmental stresses (including heavy metal pollution) have caused serious mortality in commercially cultured shrimps. Thus, we probed roles of Cdc42 in Litopenaeus vannamei shrimps (LvCdc42) during cadmium exposure by inhibiting its expression using dsRNA-mediated RNA interference. The treatment successfully reduced expression levels of MAPKs (including p38, JNK, and ERK). Cadmium exposure induced significant increases in expression levels of LvCdc42 and MAPKs, accompanied by reductions in total hemocyte counts (THC) and increases in apoptotic hemocyte ratios and ROS production. However, all of these responses were much weaker in LvCdc42-suppressed shrimps, in which mortality rates were higher than in controls. Our results suggest that the MAPK pathway plays a vital role in shrimps’ responses to Cd{sup 2+}. They also indicate that LvCdc42 in shrimps participates in its regulation, and thus plays key roles in ROS production, regulation of apoptosis and associated stress responses.

  20. Calcium signaling and the MAPK cascade are required for sperm activation in Caenorhabditis elegans.

    Science.gov (United States)

    Liu, Zhiyu; Wang, Bin; He, Ruijun; Zhao, Yanmei; Miao, Long

    2014-02-01

    In nematode, sperm activation (or spermiogenesis), a process in which the symmetric and non-motile spermatids transform into polarized and crawling spermatozoa, is critical for sperm cells to acquire fertilizing competence. SPE-8 dependent and SPE-8 independent pathways function redundantly during sperm activation in both males and hermaphrodites of Caenorhabditis elegans. However, the downstream signaling for both pathways remains unclear. Here we show that calcium signaling and the MAPK cascade are required for both SPE-8 dependent and SPE-8 independent sperm activation, implying that both pathways share common downstream signaling components during sperm activation. We demonstrate that activation of the MAPK cascade is sufficient to activate spermatids derived from either wild-type or spe-8 group mutant males and that activation of the MAPK cascade bypasses the requirement of calcium signal to induce sperm activation, indicating that the MAPK cascade functions downstream of or parallel with the calcium signaling during sperm activation. Interestingly, the persistent activation of MAPK in activated spermatozoa inhibits Major Sperm Protein (MSP)-based cytoskeleton dynamics. We demonstrate that MAPK plays dual roles in promoting pseudopod extension during sperm activation but also blocking the MSP-based, amoeboid motility of the spermatozoa. Thus, though nematode sperm are crawling cells, morphologically distinct from flagellated sperm, and the molecular machinery for motility of amoeboid and flagellated sperm is different, both types of sperm might utilize conserved signaling pathways to modulate sperm maturation.

  1. Enhancement of tunability of MAPK cascade due to coexistence of processive and distributive phosphorylation mechanisms.

    Science.gov (United States)

    Sun, Jianqiang; Yi, Ming; Yang, Lijian; Wei, Wenbin; Ding, Yiming; Jia, Ya

    2014-03-04

    The processive phosphorylation mechanism becomes important when there is macromolecular crowding in the cytoplasm. Integrating the processive phosphorylation mechanism with the traditional distributive one, we propose a mixed dual-site phosphorylation (MDP) mechanism in a single-layer phosphorylation cycle. Further, we build a degree model by applying the MDP mechanism to a three-layer mitogen-activated protein kinase (MAPK) cascade. By bifurcation analysis, our study suggests that the crowded-environment-induced pseudoprocessive mechanism can qualitatively change the response of this biological network. By adjusting the degree of processivity in our model, we find that the MAPK cascade is able to switch between the ultrasensitivity, bistability, and oscillatory dynamical states. Sensitivity analysis shows that the theoretical results remain unchanged within a reasonably chosen variation of parameter perturbation. By scaling the reaction rates and also introducing new connections into the kinetic scheme, we further construct a proportion model of the MAPK cascade to validate our findings. Finally, it is illustrated that the spatial propagation of the activated MAPK signal can be improved (or attenuated) by increasing the degree of processivity of kinase (or phosphatase). Our research implies that the MDP mechanism makes the MAPK cascade become a flexible signal module, and the coexistence of processive and distributive phosphorylation mechanisms enhances the tunability of the MAPK cascade.

  2. MAPK signaling promotes axonal degeneration by speeding the turnover of the axonal maintenance factor NMNAT2

    Science.gov (United States)

    Walker, Lauren J; Summers, Daniel W; Sasaki, Yo; Brace, EJ; Milbrandt, Jeffrey; DiAntonio, Aaron

    2017-01-01

    Injury-induced (Wallerian) axonal degeneration is regulated via the opposing actions of pro-degenerative factors such as SARM1 and a MAPK signal and pro-survival factors, the most important of which is the NAD+ biosynthetic enzyme NMNAT2 that inhibits activation of the SARM1 pathway. Here we investigate the mechanism by which MAPK signaling facilitates axonal degeneration. We show that MAPK signaling promotes the turnover of the axonal survival factor NMNAT2 in cultured mammalian neurons as well as the Drosophila ortholog dNMNAT in motoneurons. The increased levels of NMNAT2 are required for the axonal protection caused by loss of MAPK signaling. Regulation of NMNAT2 by MAPK signaling does not require SARM1, and so cannot be downstream of SARM1. Hence, pro-degenerative MAPK signaling functions upstream of SARM1 by limiting the levels of the essential axonal survival factor NMNAT2 to promote injury-dependent SARM1 activation. These findings are consistent with a linear molecular pathway for the axonal degeneration program. DOI: http://dx.doi.org/10.7554/eLife.22540.001 PMID:28095293

  3. PPARy phosphorylation mediated by JNK MAPK: a potential role in macrophage-derived foam cell formation

    Institute of Scientific and Technical Information of China (English)

    Ran YIN; Yu-gang DONG; Hong-lang LI

    2006-01-01

    Aim: To investigate whether oxidized low-density lipoprotein (ox-LDL) modulates peroxisome proliferator-activated receptor γ (PPARγ) activity through phosphorylation in macrophages, and the effect of PPARy phosphorylation on macrophages-derived foam cell formation. Methods: After exposing the cultured THP-1 cells to ox-LDL in the presence or absence of different mitogen-activated protein kinase (MAPK) inhibitors, PPARγ and phosphorylated PPARγ protein levels were detected by Western blot. MAPK activity was analyzed using MAP Kinase Assay Kit. Intracellular cholesterol accumulation was assessed by Oil red O staining and cholesterol oxidase enzymatic method. The Mrna level of PPARγ target gene was determined by reverse transcription-polymerase chain reaction (RT-PCR). Results: ox-LDL evaluated PPARγ phosphorylation status and subsequently decreased PPARγ target gene expression in a dose-dependent manner. Ox-LDL also induced MAPK activation. Treatment of THP-1 cells with c-Jun N-terminal kinase-, but not p38- or extracellular signal-regulated kinase-MAPK inhibitor, significantly suppressed PPARγ phosphorylation induced by ox-LDL, which in turn inhibited foam cell formation. Conclusion: In addition to its ligand-dependent activation, ox-LDL modulates PPARγ activity through phosphorylation, which is mediated by MAPK activation. PPARγ phosphorylation mediated by MAPK facilitates foam cell formation from macrophages exposed to ox-LDL.

  4. The Role of p38 MAPK and Its Substrates in Neuronal Plasticity and Neurodegenerative Disease

    Directory of Open Access Journals (Sweden)

    Sônia A. L. Corrêa

    2012-01-01

    Full Text Available A significant amount of evidence suggests that the p38-mitogen-activated protein kinase (MAPK signalling cascade plays a crucial role in synaptic plasticity and in neurodegenerative diseases. In this review we will discuss the cellular localisation and activation of p38 MAPK and the recent advances on the molecular and cellular mechanisms of its substrates: MAPKAPK 2 (MK2 and tau protein. In particular we will focus our attention on the understanding of the p38 MAPK-MK2 and p38 MAPK-tau activation axis in controlling neuroinflammation, actin remodelling and tau hyperphosphorylation, processes that are thought to be involved in normal ageing as well as in neurodegenerative diseases. We will also give some insight into how elucidating the precise role of p38 MAPK-MK2 and p38 MAPK-tau signalling cascades may help to identify novel therapeutic targets to slow down the symptoms observed in neurodegenerative diseases such as Alzheimer's and Parkinson's disease.

  5. The Role of p38 MAPK in the Development of Diabetic Cardiomyopathy

    Directory of Open Access Journals (Sweden)

    Shudong Wang

    2016-06-01

    Full Text Available Diabetic cardiomyopathy (DCM is a major complication of diabetes that contributes to an increase in mortality. A number of mechanisms potentially explain the development of DCM including oxidative stress, inflammation and extracellular fibrosis. Mitogen-activated protein kinase (MAPK-mediated signaling pathways are common among these pathogenic responses. Among the diverse array of kinases, extensive attention has been given to p38 MAPK due to its capacity for promoting or inhibiting the translation of target genes. Growing evidence has indicated that p38 MAPK is aberrantly expressed in the cardiovascular system, including the heart, under both experimental and clinical diabetic conditions and, furthermore, inhibition of p38 MAPK activation in transgenic animal model or with its pharmacologic inhibitor significantly prevents the development of DCM, implicating p38 MAPK as a novel diagnostic indicator and therapeutic target for DCM. This review summarizes our current knowledge base to provide an overview of the impact of p38 MAPK signaling in diabetes-induced cardiac remodeling and dysfunction.

  6. Stimulation of p38 MAPK by hormal preconditioning with atrial natriuretic peptide

    Institute of Scientific and Technical Information of China (English)

    Alexandra K. Kiemer; Stefanie Kulhanek-Heinze; Tobias Gerwig; Alexander L. Gerbes; Angelika M. Vollmar

    2002-01-01

    AIM: Stress-activated signaling pathways responsiblefor hepatic ischemia reperfusion injury and theirmodulation by protective interventions are widelyunknown. Preconditioning of rat livers with AtrialNatriuretic Peptide (ANP) attenuates ischemiareperfusion injury (Gerbes et al. Hepatology 1998, 28:1309-1317). Since ANP has recently been shown to bea regulator of the p38 MAPK pathway in endothelialcells (Kiemer et al. Circ Res 2002, 90:874-881), aim ofthis study was to investigate activities of MAPK duringischemia and reperfusion and effects of ANP on MAPK.METHODS: Rat livers were perfused with KH-buffer inthe presence or absence of ANP for 20 min, kept in coldUW solution for 24 h, and reperfused for up to 120 min.Activities of p38 MAPK and JNK was determined by invitro phosphorylation assays using MBP and c-jun assubstrates. After SDS/PAGE electrophoresis, gels werequantified by phosphorimaging.RESULTS: Activity of p38 MAPK in control organsdecreased in the course of ischemia and reperfusionby 85%, whereas ANP increased p38 activity by up to30-fold. JNK activation of control livers increased in thecourse of ischemia and reperfusion by up to three-fold.This increase in JNK activity was slightly elevated inANP preconditioned organs.CONCLUSION: This work represents a systematicinvestigation of MAPK activation during liver ischemiaand reperfusion. Employing ANP, for the first time apharmacological approach to modulate these centralsignal transduction molecules is presented.

  7. The MAPK pathway as an apoptosis enhancer in melanoma.

    Science.gov (United States)

    Haydn, Johannes M; Hufnagel, Anita; Grimm, Johannes; Maurus, Katja; Schartl, Manfred; Meierjohann, Svenja

    2014-07-15

    Inhibition of RAF/MEK/ERK signaling is beneficial for many patients with BRAF(V600E)-mutated melanoma. However, primary and secondary resistances restrict long-lasting therapy success. Combination therapies are therefore urgently needed. Here, we evaluate the cellular effect of combining a MEK inhibitor with a genotoxic apoptosis inducer. Strikingly, we observed that an activated MAPK pathway promotes in several melanoma cell lines the pro-apoptotic response to genotoxic stress, and MEK inhibition reduces intrinsic apoptosis. This goes along with MEK inhibitor induced increased RAS and P-AKT levels. The protective effect of the MEK inhibitor depends on PI3K signaling, which prevents the induction of pro-apoptotic PUMA that mediates apoptosis after DNA damage. We could show that the MEK inhibitor dependent feedback loop is enabled by several factors, including EGF receptor and members of the SPRED family. The simultaneous knockdown of SPRED1 and SPRED2 mimicked the effects of MEK inhibitor such as PUMA repression and protection from apoptosis. Our data demonstrate that MEK inhibition of BRAF(V600E)-positive melanoma cells can protect from genotoxic stress, thereby achieving the opposite of the intended anti-tumorigenic effect of the combination of MEK inhibitor with inducers of intrinsic apoptosis.

  8. A novel cardioprotective p38-MAPK/mTOR pathway.

    Science.gov (United States)

    Hernández, Gonzalo; Lal, Hind; Fidalgo, Miguel; Guerrero, Ana; Zalvide, Juan; Force, Thomas; Pombo, Celia M

    2011-12-10

    Despite intensive study, the mechanisms regulating activation of mTOR and the consequences of that activation in the ischemic heart remain unclear. This is particularly true for the setting of ischemia/reperfusion (I/R) injury. In a mouse model of I/R injury, we observed robust mTOR activation, and its inhibition by rapamycin increased injury. Consistent with the in-vivo findings, mTOR activation was also protective in isolated cardiomyocytes exposed to two models of I/R. Moreover, we identify a novel oxidant stress-activated pathway regulating mTOR that is critically dependent on p38-MAPK and Akt. This novel p38-regulated pathway signals downstream through REDD1, Tsc2, and 14-3-3 proteins to activate mTOR and is independent of AMPK. The protective role of p38/Akt and mTOR following oxidant stress is a general phenomenon since we observed it in a wide variety of cell types. Thus we have identified a novel protective pathway in the cardiomyocyte involving p38-mediated mTOR activation. Furthermore, the p38-dependent protective pathway might be able to be selectively modulated to enhance cardio-protection while not interfering with the inhibition of the better-known detrimental p38-dependent pathways.

  9. Protein phosphatase PPM1G regulates protein translation and cell growth by dephosphorylating 4E binding protein 1 (4E-BP1).

    Science.gov (United States)

    Liu, Jianyu; Stevens, Payton D; Eshleman, Nichole E; Gao, Tianyan

    2013-08-09

    Protein translation initiation is a tightly controlled process responding to nutrient availability and mitogen stimulation. Serving as one of the most important negative regulators of protein translation, 4E binding protein 1 (4E-BP1) binds to translation initiation factor 4E and inhibits cap-dependent translation in a phosphorylation-dependent manner. Although it has been demonstrated previously that the phosphorylation of 4E-BP1 is controlled by mammalian target of rapamycin in the mammalian target of rapamycin complex 1, the mechanism underlying the dephosphorylation of 4E-BP1 remains elusive. Here, we report the identification of PPM1G as the phosphatase of 4E-BP1. A coimmunoprecipitation experiment reveals that PPM1G binds to 4E-BP1 in cells and that purified PPM1G dephosphorylates 4E-BP1 in vitro. Knockdown of PPM1G in 293E and colon cancer HCT116 cells results in an increase in the phosphorylation of 4E-BP1 at both the Thr-37/46 and Ser-65 sites. Furthermore, the time course of 4E-BP1 dephosphorylation induced by amino acid starvation or mammalian target of rapamycin inhibition is slowed down significantly in PPM1G knockdown cells. Functionally, the amount of 4E-BP1 bound to the cap-dependent translation initiation complex is decreased when the expression of PPM1G is depleted. As a result, the rate of cap-dependent translation, cell size, and protein content are increased in PPM1G knockdown cells. Taken together, our study has identified protein phosphatase PPM1G as a novel regulator of cap-dependent protein translation by negatively controlling the phosphorylation of 4E-BP1.

  10. Tau phosphorylation in human, primate, and rat brain: evidence that a pool of tau is highly phosphorylated in vivo and is rapidly dephosphorylated in vitro.

    Science.gov (United States)

    Garver, T D; Harris, K A; Lehman, R A; Lee, V M; Trojanowski, J Q; Billingsley, M L

    1994-12-01

    The extent of tau phosphorylation is thought to regulate the binding of tau to microtubules: Highly phosphorylated tau does not bind to tubules, whereas dephosphorylated tau can bind to microtubules. It is interesting that the extent of tau phosphorylation in vivo has not been accurately determined. Tau was rapidly isolated from human temporal neocortex and hippocampus, rhesus monkey temporal neocortex, and rat temporal neocortex and hippocampus under conditions that minimized dephosphorylation. In brain slices, we observed that tau isolated under such conditions largely existed in several phosphorylated states, including a pool that was highly phosphorylated; this was determined using epitope-specific monoclonal and polyclonal antibodies. This highly phosphorylated tau was dephosphorylated during a 120-min time course in vitro, presumably as a result of neuronal phosphatase activity. The slow-mobility forms of tau were shifted to faster-mobility forms following in vitro incubation with alkaline phosphatase. Laser densitometry was used to estimate the percent of tau in slow-mobility, highly phosphorylated forms. Approximately 25% of immunoreactive tau was present as slow-mobility (66- and 68-kDa) forms of tau. The percentage of immunoreactive tau in faster-mobility pools (42-54 kDa) increased in proportion to the decrease in content of 66-68-kDa tau as a function of neuronal phosphatases or alkaline phosphatase treatment. These data suggest that the turnover of phosphorylated sites on tau is rapid and depends on neuronal phosphatases. Furthermore, tau is highly phosphorylated in normal-appearing human, primate, and rodent brain.(ABSTRACT TRUNCATED AT 250 WORDS)

  11. Intrathecal MK-801 inhibits formalin-induced activation of spinal p38-MAPK in rats

    Institute of Scientific and Technical Information of China (English)

    Zhifeng Peng; Xin Zhao; Xing Jin; Xiaochun Yan; Xiaorong Yang; Ce Zhang

    2008-01-01

    BACKGROUND: p38 mitogen-activated protein kinase (MAPK) plays an instrumental role in signal transduction from the cell surface to the nucleus, while subcutaneous injection of formalin can induce increased activation of spinal p38 MAPK. However, the mechanisms underlying the formalin-induced activation of spinal p38 MAPK in rats are unclear. OBJECTIVE: To observe the effects of N-methyl-D-aspartic acid (NMDA) receptor antagonist MK-801 on the formalin-induced activation of spinal p38 MAPK in rats. DESIGN, TIME AND SETTING: This randomized grouping, controlled animal experiment was performed at the Department of Physiology and Neurobiology, Shanxi Medical University between May and November 2007. MATERIALS: Forty eight healthy, adult Wistar rats were randomly divided into two groups: formalin + normal saline (n = 12) and formalin + MK-801 (n = 36). The formalin + MK-801 group was further divided into three subgroups according to the dosage of MK-801 (10, 50, and 100 nmol/L, 12 rats for each subgroup) METHODS: Following anesthesia, polyethylene tubing filled with sterile normal saline was implanted into the subarachnoid cavity. On postoperative days 5-8, rats received a 15 minute perfusion of normal saline or MK-801 (10, 50, and 100 nmol/L) in the formalin + normal saline and formalin + MK-801 groups, respectively, followed by formalin injection for the induction of nociceptive behavior. MAIN OUTCOME MEASURES: Detection of total p38 MAPK and of phosphorylated p38 MAPK by western Blot analysis; observation of nociceptive behaviors in the 1 hour after formalin injection. RESULTS: Western Blot analysis revealed that injection of formalin had no effect on total p38 MAPK expression but resulted in increased phosphorylation of p38 MAPK in the spinal cord. This increase was apparent after 5 minutes, peaked at 20 minutes, and thereafter descended and reached control levels after 45 minutes. Pretreatment with MK-801 (10, 50, 100 nmol/L) resulted in a dose-dependent reduction

  12. Protein Phosphatase 1c Associated with the Cardiac Sodium Calcium Exchanger 1 Regulates Its Activity by Dephosphorylating Serine 68-phosphorylated Phospholemman.

    Science.gov (United States)

    Hafver, Tandekile Lubelwana; Hodne, Kjetil; Wanichawan, Pimthanya; Aronsen, Jan Magnus; Dalhus, Bjørn; Lunde, Per Kristian; Lunde, Marianne; Martinsen, Marita; Enger, Ulla Helene; Fuller, William; Sjaastad, Ivar; Louch, William Edward; Sejersted, Ole Mathias; Carlson, Cathrine Rein

    2016-02-26

    The sodium (Na(+))-calcium (Ca(2+)) exchanger 1 (NCX1) is an important regulator of intracellular Ca(2+) homeostasis. Serine 68-phosphorylated phospholemman (pSer-68-PLM) inhibits NCX1 activity. In the context of Na(+)/K(+)-ATPase (NKA) regulation, pSer-68-PLM is dephosphorylated by protein phosphatase 1 (PP1). PP1 also associates with NCX1; however, the molecular basis of this association is unknown. In this study, we aimed to analyze the mechanisms of PP1 targeting to the NCX1-pSer-68-PLM complex and hypothesized that a direct and functional NCX1-PP1 interaction is a prerequisite for pSer-68-PLM dephosphorylation. Using a variety of molecular techniques, we show that PP1 catalytic subunit (PP1c) co-localized, co-fractionated, and co-immunoprecipitated with NCX1 in rat cardiomyocytes, left ventricle lysates, and HEK293 cells. Bioinformatic analysis, immunoprecipitations, mutagenesis, pulldown experiments, and peptide arrays constrained PP1c anchoring to the K(I/V)FF motif in the first Ca(2+) binding domain (CBD) 1 in NCX1. This binding site is also partially in agreement with the extended PP1-binding motif K(V/I)FF-X5-8Φ1Φ2-X8-9-R. The cytosolic loop of NCX1, containing the K(I/V)FF motif, had no effect on PP1 activity in an in vitro assay. Dephosphorylation of pSer-68-PLM in HEK293 cells was not observed when NCX1 was absent, when the K(I/V)FF motif was mutated, or when the PLM- and PP1c-binding sites were separated (mimicking calpain cleavage of NCX1). Co-expression of PLM and NCX1 inhibited NCX1 current (both modes). Moreover, co-expression of PLM with NCX1(F407P) (mutated K(I/V)FF motif) resulted in the current being completely abolished. In conclusion, NCX1 is a substrate-specifying PP1c regulator protein, indirectly regulating NCX1 activity through pSer-68-PLM dephosphorylation.

  13. Linear B-cell epitope mapping of MAPK3 and MAPK4 from Leishmania braziliensis: implications for the serodiagnosis of human and canine leishmaniasis.

    Science.gov (United States)

    Menezes-Souza, Daniel; de Oliveira Mendes, Tiago Antônio; de Araújo Leão, Ana Carolina; de Souza Gomes, Matheus; Fujiwara, Ricardo Toshio; Bartholomeu, Daniella Castanheira

    2015-02-01

    The correct and early identification of humans and dogs infected with Leishmania are key steps in the control of leishmaniasis. Additionally, a method with high sensitivity and specificity at low cost that allows the screening of a large number of samples would be extremely valuable. In this study, we analyzed the potential of mitogen-activated protein kinase 3 (MAPK3) and mitogen-activated protein kinase 4 (MAPK4) proteins from Leishmania braziliensis to serve as antigen candidates for the serodiagnosis of human visceral and tegumentary leishmaniasis, as well as canine visceral disease. Moreover, we mapped linear B-cell epitopes in these proteins and selected those epitopes with sequences that were divergent in the corresponding orthologs in Homo sapiens, in Canis familiaris, and in Trypanosoma cruzi. We compared the performance of these peptides with the recombinant protein using ELISA. Both MAPK3 and MAPK4 recombinant proteins showed better specificity in the immunodiagnosis of human and canine leishmaniasis than soluble parasite antigens and the EIE-leishmaniose-visceral-canina-bio-manguinhos (EIE-LVC) kit. Furthermore, the performance of this serodiagnosis assay was improved using synthetic peptides corresponding to B-cell epitopes derived from both proteins.

  14. Xenon preconditioning differently regulates p44/42 MAPK (ERK 1/2) and p46/54 MAPK (JNK 1/2 and 3) in vivo

    NARCIS (Netherlands)

    N.C. Weber; J. Stursberg; N.M. Wirthle; O. Toma; W. Schlack; B. Preckel

    2006-01-01

    Background. Xenon (Xe) induces preconditioning (PC) of the rat heart in vivo via activation of p38 mitogen-activated protein kinase (MAPK). The role of ERK 1/2 and JNK 1/2 and 3 in Xe-PC has yet not been determined. Methods. For infarct size measurements, anaesthetized rats were subjected to 25 min

  15. Identification of novel PAMP-triggered phosphorylation and dephosphorylation events in arabidopsis thaliana by quantitative phosphoproteomic analysis

    KAUST Repository

    Rayapuram, Naganand

    2014-04-04

    Signaling cascades rely strongly on protein kinase-mediated substrate phosphorylation. Currently a major challenge in signal transduction research is to obtain high confidence substrate phosphorylation sites and assign them to specific kinases. In response to bacterial flagellin, a pathogen-associated molecular pattern (PAMP), we searched for rapidly phosphorylated proteins in Arabidopsis thaliana by combining multistage activation (MSA) and electron transfer dissociation (ETD) fragmentation modes, which generate complementary spectra and identify phosphopeptide sites with increased reliability. Of a total of 825 phosphopeptides, we identified 58 to be differentially phosphorylated. These peptides harbor kinase motifs of mitogen-activated protein kinases (MAPKs) and calcium-dependent protein kinases (CDPKs), as well as yet unknown protein kinases. Importantly, 12 of the phosphopeptides show reduced phosphorylation upon flagellin treatment. Since protein abundance levels did not change, these results indicate that flagellin induces not only various protein kinases but also protein phosphatases, even though a scenario of inhibited kinase activity may also be possible. © 2014 American Chemical Society.

  16. A specific mechanomodulatory role for p38 MAPK in embryonic joint articular surface cell MEK-ERK pathway regulation.

    Science.gov (United States)

    Lewthwaite, Jo C; Bastow, Edward R; Lamb, Katherine J; Blenis, John; Wheeler-Jones, Caroline P D; Pitsillides, Andrew A

    2006-04-21

    Mechanisms regulating cell behavior and extracellular matrix composition in response to mechanical stimuli remain unresolved. Our previous studies have established that the MEK-ERK cascade plays a specific role in the mechano-dependent joint formation process by promoting the assembly of pericellular matrices reliant upon hyaluronan (HA) for their integrity. Here we demonstrate: (i) novel cross-talk between p38 MAPK and MEK-ERK signaling pathways that is specific for mechanical stimuli and (ii) a role for p38 MAPK in facilitating HA production by cells derived from the articular surface of embryonic chick tibiotarsal joints. We find that p38 MAPK blockade restricts pericellular assembly of HA-rich matrices and reduces basal as well as mechanical strain-induced release of HA. p38 MAPK blockers potentiated early strain-induced increases but restricted sustained increases in MEK/ERK phosphorylation at later times; c-Fos hyperphosphorylation at threonine 325 was found to parallel this p38 MAPK-mediated modulation of ERK activation. In contrast, p38 MAPK inhibitors had no detectable effect on the ERK activation induced by fibroblast growth factor 2 or pervanadate, a phosphatase inhibitor, and MEK inhibitors did not influence p38 MAPK phosphorylation, confirming both the specificity and unidirectionality of p38 MAPK-ERK cross-talk. Immunochemical and immunoblotting studies revealed constitutive p38 MAPK activation in cells at, or derived from, developing articular joint surfaces. Unlike the MEK-ERK pathway, however, p38 MAPK was not further stimulated by mechanical stimulation in vitro. Thus, p38 MAPK specifically facilitates ERK activation and downstream signaling in response to mechanical stimuli. These results suggest that constitutively active p38 MAPK serves an essential, permissive role in mechanically induced changes in ERK activation and in the accumulation of HA-rich extracellular matrices that serve a key role in joint development.

  17. Soybean MAPK, GMK1 Is Dually Regulated by Phosphatidic Acid and Hydrogen Peroxide and Translocated to Nucleus during Salt Stress

    OpenAIRE

    Im, Jong Hee; Lee, Hyoungseok; Kim, Jitae; Kim, Ho Bang; An, Chung Sun

    2012-01-01

    Mitogen-activated protein kinase (MAPK) is activated by various biotic and abiotic stresses. Salt stress induces two well-characterized MAPK activating signaling molecules, phosphatidic acid (PA) via phospholipase D and phospholipase C, and reactive oxygen species (ROS) via nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase. In our previous study, the activity of soybean MAPK, GMK1 was strongly induced within 5 min of 300 mM NaCl treatment and this early activity was regulated by PA....

  18. In silico identification and characterization of the MAPK family members of unicellular model eukaryote Tetrahymena thermophila.

    Science.gov (United States)

    Yıldız, Mehmet Taha; Arslanyolu, Muhittin

    2014-10-01

    The biological function and evolutionary diversity of the mitogen-activated protein kinase (MAPK) family have mostly been studied in fungi, animals and plants, with very limited information from lower eukaryotes. This study aimed to describe the MAPKs of unicellular Tetrahymena thermophila. Eight members of the T. thermophila MAPK (TtMPK) gene family, in addition to previously reported TtMPK1, TtMPK2 and TtMPK3, were identified bioinformatically using a T. thermophila genome database. Phylogenetic analysis assigned the TtMPKs into two major groups, ERK1/2-like (TtMPK1, 2, 3, 5, 6, 7, 8, and 9) as stress-responsive MAPKs for biotic and abiotic stresses, and ERK7/8-like (TtMPK4, 10, and 11) as cell-cycle-associated protein kinases for biotic factors. Semi-quantitative RT-PCR analysis of the TtMPKs showed high mRNA expression at 30°C; however, only TtMPK5 and TtMPK6 showed high expression at 37°C. Osmotic shock by 100mM NaCl only increased the expression of TtMPK2, whereas 20mM NaCl reduced the expression of all MPKs to almost zero. The results suggested that T. thermophila MAPKs are among the closest representatives of the ancestors of the eukaryotic MAPK family. Although no functional characterization of MPKs was performed, this study is the first report of the genome-wide MAPK family in T. thermophila. Copyright © 2014 Elsevier GmbH. All rights reserved.

  19. Regulation of hepatitis C virus replication and gene expression by the MAPK-ERK pathway.

    Science.gov (United States)

    Pei, Rongjuan; Zhang, Xiaoyong; Xu, Song; Meng, Zhongji; Roggendorf, Michael; Lu, Mengji; Chen, Xinwen

    2012-10-01

    The mitogen activated protein kinases-extracellular signal regulated kinases (MAPK-ERK) pathway is involved in regulation of multiple cellular processes including the cell cycle. In the present study using a Huh7 cell line Con1 with an HCV replicon, we have shown that the MAPK-ERK pathway plays a significant role in the modulation of HCV replication and protein expression and might influence IFN-α signalling. Epithelial growth factor (EGF) was able to stimulate ERK activation and decreased HCV RNA load while a MAPK-ERK pathway inhibitor U0126 led to an elevated HCV RNA load and higher NS5A protein amounts in Con1 cells. It could be further demonstrated that the inhibition of the MAPK-ERK pathway facilitated the translation directed by the HCV internal ribosome entry site. Consistently, a U0126 treatment enhanced activity of the HCV reporter replicon in transient transfection assays. Thus, the MAPK-ERK pathway plays an important role in the regulation of HCV gene expression and replication. In addition, cyclin-dependent kinases (CDKs) downstream of ERK may also be involved in the modulation of HCV replication since roscovitine, an inhibitor of CDKs had a similar effect to that of U0126. Modulation of the cell cycle progression by cell cycle inhibitor or RNAi resulted consistently in changes of HCV RNA levels. Further, the replication of HCV replicon in Con1 cells was inhibited by IFN-α. The inhibitory effect of IFN-α could be partly reversed by pre-incubation of Con-1 cells with inhibitors of the MAPK-ERK pathway and CDKs. It could be shown that the MAPK-ERK inhibitors are able to partially modulate the expression of interferon-stimulated genes.

  20. MAPK15 upregulation promotes cell proliferation and prevents DNA damage in male germ cell tumors

    Science.gov (United States)

    Ilardi, Gennaro; Acunzo, Mario; Nigita, Giovanni; Sasdelli, Federica; Celetti, Angela; Strambi, Angela; Staibano, Stefania; Croce, Carlo Maria; Chiariello, Mario

    2016-01-01

    Germ cell tumors (GCT) are the most common malignancies in males between 15 and 35 years of age. Despite the high cure rate, achieved through chemotherapy and/or surgery, the molecular basis of GCT etiology is still largely obscure. Here, we show a positive correlation between MAPK15 (ERK8; ERK7) expression and specific GCT subtypes, with the highest levels found in the aggressive embryonal carcinomas (EC). Indeed, in corresponding cellular models for EC, MAPK15 enhanced tumorigenicity in vivo and promoted cell proliferation in vitro, supporting a role for this kinase in human GCT. At molecular level, we demonstrated that endogenous MAPK15 is necessary to sustain cell cycle progression of EC cells, by limiting p53 activation and preventing the triggering of p53-dependent mechanisms resulting in cell cycle arrest. To understand MAPK15-dependent mechanisms impinging on p53 activation, we demonstrate that this kinase efficiently protects cells from DNA damage. Moreover, we show that the ability of MAPK15 to control the autophagic process is necessary for basal management of DNA damage and for tumor formation controlled by the kinase. In conclusion, our findings suggest that MAPK15 overexpression may contribute to the malignant transformation of germ cells by controlling a “stress support” autophagic pathway, able to prevent DNA damage and the consequent activation of the p53 tumor suppressor. Moreover, in light of these results, MAPK15-specific inhibitors might represent new tools to enhance the therapeutic index of cytotoxic therapy in GCT treatment, and to increase the sensitivity to DNA-damaging drugs in other chemotherapy-resistant human tumors. PMID:26988910

  1. Hepatocyte cytoskeleton during ischemia and reperfusion influence of ANP-mediated p38 MAPK activation

    Institute of Scientific and Technical Information of China (English)

    Melanie Keller; Alexander L Gerbes; Stefanie Kulhanek-Heinze; Tobias Gerwig; Uwe Grützner; Nico van Rooijen; Angelika M Vollmar; Alexandra K Kiemer

    2005-01-01

    AIM: To determine functional consequences of this activation, whereby we focused on a potential regulation of the hepatocyte cytoskeleton during ischemia and reperfusion.METHODS: For in vivo experiments, animals received ANP (5 μg/kg) intravenously. In a different experimental setting, isolated rat livers were perfused with KH-buffer ±ANP (200 nmol/L)±SB203580 (2 μmol/L). Liverswere then kept under ischemic conditions for 24 h, and either transplanted or reperfused. Actin, Hsp27, and phosphorylated Hsp27 were determined by Western blotting, p38 MAPK activity by in vitro phosphorylation assay. F-actin distribution was determined by confocal microscopy.RESULTS: We first confirmed that ANP preconditioning leads to an activation of p38 MAPK and observedalterations of the cytoskeleton in hepatocytes of ANPpreconditioned organs. ANP induced an increase of hepatic F-actin after ischemia, which could be prevented by the p38 MAPK inhibitor SB203580 but had no effect on bile flow. After ischemia untreated livers showed a translocation of Hsp27 towards the cytoskeleton and an increase in total Hsp27, whereas ANP preconditioning prohibited translocation but caused an augmentation of Hsp27 phosphorylation. This effect is also mediated via p38 MAPK, since it was abrogated by the p38 MAPK inhibitor SB203580.CONCLUSION: This study reveals that ANP-mediated p38 MAPK activation leads to changes in hepatocyte cytoskeleton involving an elevation of phosphorylated Hsp27 and thereby for the first time shows functional consequences of ANP-induced hepatic p38 MAPK activation.

  2. Regulation of Hepatitis C Virus Replication and Gene Expression by the MAPK-ERK Pathway

    Institute of Scientific and Technical Information of China (English)

    Rongjuan Pei; Xiaoyong Zhang; Song Xu; Zhongji Meng; Michael Roggendorf; Mengji Lu; Xinwen Chen

    2012-01-01

    The mitogen activated protein kinases-extracellular signal regulated kinases (MAPK-ERK) pathway is involved in regulation of multiple cellular processes including the cell cycle.In the present study using a Huh7 cell line Con1 with an HCV replicon,we have shown that the MAPK-ERK pathway plays a significant role in the modulation of HCV replication and protein expression and might influence IFN-α signalling.Epithelial growth factor (EGF) was able to stimulate ERK activation and decreased HCV RNA load while a MAPK-ERK pathway inhibitor U0126 led to an elevated HCV RNA load and higher NS5A protein amounts in Con1 cells.It could be further demonstrated that the inhibition of the MAPK-ERK pathway facilitated the translation directed by the HCV internal ribosome entry site.Consistently,a U0126 treatment enhanced activity of the HCV reporter replicon in transient transfection assays.Thus,the MAPK-ERK pathway plays an important role in the regulation of HCV gene expression and replication.In addition,cyclin-dependent kinases (CDKs) downstream of ERK may also be involved in the modulation of HCV replication since roscovitine,an inhibitor of CDKs had a similar effect to that of U0126.Modulation of the cell cycle progression by cell cycle inhibitor or RNAi resulted consistently in changes of HCV RNA levels.Further,the replication of HCV replicon in Conl cells was inhibited by IFN-α.The inhibitory effect of IFN-α could be partly reversed by pre-incubation of Con-1 cells with inhibitors of the MAPK-ERK pathway and CDKs.It could be shown that the MAPK-ERK inhibitors are able to partially modulate the expression of interferon-stimulated genes.

  3. MAPK14/p38α confers irinotecan resistance to TP53-defective cells by inducing survival autophagy.

    Science.gov (United States)

    Paillas, Salome; Causse, Annick; Marzi, Laetitia; de Medina, Philippe; Poirot, Marc; Denis, Vincent; Vezzio-Vie, Nadia; Espert, Lucile; Arzouk, Hayat; Coquelle, Arnaud; Martineau, Pierre; Del Rio, Maguy; Pattingre, Sophie; Gongora, Céline

    2012-07-01

    Recently we have shown that the mitogen-activated protein kinase (MAPK) MAPK14/p38α is involved in resistance of colon cancer cells to camptothecin-related drugs. Here we further investigated the cellular mechanisms involved in such drug resistance and showed that, in HCT116 human colorectal adenocarcinoma cells in which TP53 was genetically ablated (HCT116-TP53KO), overexpression of constitutively active MAPK14/p38α decreases cell sensitivity to SN-38 (the active metabolite of irinotecan), inhibits cell proliferation and induces survival-autophagy. Since autophagy is known to facilitate cancer cell resistance to chemotherapy and radiation treatment, we then investigated the relationship between MAPK14/p38α, autophagy and resistance to irinotecan. We demonstrated that induction of autophagy by SN38 is dependent on MAPK14/p38α activation. Finally, we showed that inhibition of MAPK14/p38α or autophagy both sensitizes HCT116-TP53KO cells to drug therapy. Our data proved that the two effects are interrelated, since the role of autophagy in drug resistance required the MAPK14/p38α. Our results highlight the existence of a new mechanism of resistance to camptothecin-related drugs: upon SN38 induction, MAPK14/p38α is activated and triggers survival-promoting autophagy to protect tumor cells against the cytotoxic effects of the drug. Colon cancer cells could thus be sensitized to drug therapy by inhibiting either MAPK14/p38 or autophagy.

  4. Ratiometric analysis in hyperpolarized NMR (I): test of the two-site exchange model and the quantification of reaction rate constants.

    Science.gov (United States)

    Li, Lin Z; Kadlececk, Stephen; Xu, He N; Daye, Dania; Pullinger, Benjamin; Profka, Harrilla; Chodosh, Lewis; Rizi, Rahim

    2013-10-01

    Conventional methods for the analysis of in vivo hyperpolarized (13) C NMR data from the lactate dehydrogenase (LDH) reaction usually make assumptions on the stability of rate constants and/or the validity of the two-site exchange model. In this study, we developed a framework to test the validity of the assumption of stable reaction rate constants and the two-site exchange model in vivo via ratiometric fitting of the time courses of the signal ratio L(t)/P(t). Our analysis provided evidence that the LDH enzymatic kinetics observed by hyperpolarized NMR are in near-equilibrium and satisfy the two-site exchange model for only a specific time window. In addition, we quantified both the forward and reverse exchange rate constants of the LDH reaction for the transgenic and mouse xenograft models of breast cancer using the ratio fitting method developed, which includes only two modeling parameters and is less sensitive to the influence of instrument settings/protocols, such as flip angles, degree of polarization and tracer dosage. We further compared the ratio fitting method with a conventional two-site exchange modeling method, i.e. the differential equation fitting method, using both the experimental and simulated hyperpolarized NMR data. The ratio fitting method appeared to fit better than the differential equation fitting method for the reverse rate constant on the mouse tumor data, with less relative errors on average, whereas the differential equation fitting method also resulted in a negative reverse rate constant for one tumor. The simulation results indicated that the accuracy of both methods depends on the width of the transport function, noise level and rate constant ratio; one method may be more accurate than the other based on the experimental/biological conditions aforementioned. We were able to categorize our tumor models into specific conditions of the computer simulation and to estimate the errors of rate quantification. We also discussed possible

  5. Experimental Verification of the Chemical Sensitivity of Two-Site Double Core-Hole States Formed by an X-ray FEL

    CERN Document Server

    Salen, P; Schmidt, H T; Thomas, R D; Larsson, M; Feifel, R; Piancastelli, M N; Fang, L; Murphy, B; Osipov, T; Berrah, N; Kukk, E; Ueda, K; Bozek, J D; Bostedt, C; Wada, S; Richter, R; Feyer, V; Prince, K C

    2012-01-01

    We have performed X-ray two-photon photoelectron spectroscopy (XTPPS) using the Linac Coherent Light Source (LCLS) X-ray free-electron laser (FEL) in order to study double core-hole (DCH) states of CO2, N2O and N2. The experiment verifies the theory behind the chemical sensitivity of two-site (ts) DCH states by comparing a set of small molecules with respect to the energy shift of the tsDCH state and by extracting the relevant parameters from this shift.

  6. CSTP1, a novel protein phosphatase, blocks cell cycle, promotes cell apoptosis, and suppresses tumor growth of bladder cancer by directly dephosphorylating Akt at Ser473 site.

    Directory of Open Access Journals (Sweden)

    De-Xiang Zhuo

    Full Text Available Akt/protein kinase B is a pivotal component downstream of phosphatidylinositol 3-kinase (PI3K pathway, whose activity regulates the balance between cell survival and apoptosis. Phosphorylation of Akt occurs at two key sites either at Thr308 site in the activation loop or at Ser473 site in the hydrophobic motif. The phosphorylated form of Akt (pAkt is activated to promote cell survival. The mechanisms of pAkt dephosphorylation and how the signal transduction of Akt pathway is terminated are still largely unknown. In this study, we identified a novel protein phosphatase CSTP1(complete s transactivated protein 1, which interacts and dephosphorylates Akt specifically at Ser473 site in vivo and in vitro, blocks cell cycle progression and promotes cell apoptosis. The effects of CSTP1 on cell survival and cell cycle were abrogated by depletion of phosphatase domain of CSTP1 or by expression of a constitutively active form of Akt (S473D, suggesting Ser473 site of Akt as a primary cellular target of CSTP1. Expression profile analysis showed that CSTP1 expression is selectively down-regulated in non-invasive bladder cancer tissues and over-expression of CSTP1 suppressed the size of tumors in nude mice. Kaplan-Meier curves revealed that decreased expression of CSTP1 implicated significantly reduced recurrence-free survival in patients suffered from non-invasive bladder cancers.

  7. p38 MAPK Inhibition Improves Synaptic Plasticity and Memory in Angiotensin II-dependent Hypertensive Mice

    Science.gov (United States)

    Dai, Hai-long; Hu, Wei-yuan; Jiang, Li-hong; Li, Le; Gaung, Xue-feng; Xiao, Zhi-cheng

    2016-01-01

    The pathogenesis of hypertension-related cognitive impairment has not been sufficiently clarified, new molecular targets are needed. p38 MAPK pathway plays an important role in hypertensive target organ damage. Activated p38 MAPK was seen in AD brain tissue. In this study, we found that long-term potentiation (LTP) of hippocampal CA1 was decreased, the density of the dendritic spines on the CA1 pyramidal cells was reduced, the p-p38 protein expression in hippocampus was elevated, and cognitive function was impaired in angiotensin II-dependent hypertensive C57BL/6 mice. In vivo, using a p38 heterozygous knockdown mice (p38KI/+) model, we showed that knockdown of p38 MAPK in hippocampus leads to the improvement of cognitive function and hippocampal synaptic plasticity in angiotensin II-dependent p38KI/+ hypertensive mice. In vitro, LTP was improved in hippocampal slices from C57BL/6 hypertensive mice by treatment with p38MAPK inhibitor SKF86002. Our data demonstrated that p38 MAPK may be a potential therapeutic target for hypertension-related cognitive dysfunction. PMID:27283322

  8. Effects of acrylonitrile on lymphocyte lipid rafts and RAS/RAF/MAPK/ERK signaling pathways.

    Science.gov (United States)

    Li, X J; Li, B; Huang, J S; Shi, J M; Wang, P; Fan, W; Zhou, Y L

    2014-09-26

    Acrylonitrile (ACN) is a widely used chemical in the production of plastics, resins, nitriles, acrylic fibers, and synthetic rubber. Previous epidemiological investigations and animal studies have confirmed that ACN affects the lymphocytes and spleen. However, the immune toxicity mechanism is unknown. Lipid rafts are cell membrane structures that are rich in cholesterol and involved in cell signal transduction. The B cell lymophoma-10 (Bcl10) protein is a joint protein that is important in lymphocyte development and signal pathways. This study was conducted to examine the in vitro effects of ACN. We separated lipid rafts, and analyzed Bcl10 protein and caveolin. Western blotting was used to detect mitogen-activated protein kinase (MAPK) and phosphorylated MAPK levels. The results indicated that with increasing ACN concentration, the total amount of Bcl10 remained stable, but was concentrated mainly in part 4 to part 11 in electrophoretic band district which is high density in gradient centrifugation. Caveolin-1 was evaluated as a lipid raft marker protein; caveolin-1 content and position were relatively unchanged. Western blotting showed that in a certain range, MAPK protein was secreted at a higher level. At some ACN exposure levels, MAPK protein secretion was significantly decreased compared to the control group (P lipid raft structures, causing Bcl10 protein and lipid raft separation and restraining Ras-Raf-MAPK-extracellular signal-regulated kinase signaling pathways.

  9. Tumor heterogeneity and plasticity as elusive drivers for resistance to MAPK pathway inhibition in melanoma.

    Science.gov (United States)

    Roesch, A

    2015-06-04

    Despite the recent success of MAPK signaling-targeted drugs in melanoma, the majority of patients with metastatic melanoma still undergo disease progression after initial tumor shrinkage indicating gradually developing therapy resistance. This review will give an overview on currently suggested concepts of resistance to MAPK pathway inhibitors in melanoma with particular focus on inter- and intraindividual as well as intratumor heterogeneity. The high plasticity of melanoma cells promotes both the clonal evolution of genetic resistance, for example, because of mutations in the MAPK or PI3K/AKT/PTEN pathways, and the emergence of cell phenotypes that functionally and metabolically overcome MAPK inhibition. Like a 'moving target', melanoma cells are shifting between different metabolic, cell cycle and differentiation states reflecting a highly dynamic potential to adapt to exogenous stressors including drugs. The introduction of MAPK inhibitors into the clinics has tremendously pushed the field of melanoma research not just because of the historic therapeutic success but also by providing a new tool to study human melanoma in its natural microenvironment.

  10. Mechanistic role of p38 MAPK in gastric cancer dissemination in a rodent model peritoneal metastasis.

    Science.gov (United States)

    Graziosi, Luigina; Mencarelli, Andrea; Santorelli, Chiara; Renga, Barbara; Cipriani, Sabrina; Cavazzoni, Emanuel; Palladino, Giuseppe; Laufer, Stefan; Burnet, Michael; Donini, Annibale; Fiorucci, Stefano

    2012-01-15

    Peritoneal dissemination is a highly frequent complication of poorly differentiated gastric cancers for which no effective therapies are available. Constitutive activation of mitogen-activated protein kinases (MAPKs) signaling cascades is recognized as a causative factor in the malignant transformation of several carcinoma cell types. In the present study we provide evidence that p38 MAPK inhibition protects against gastric cancer cells dissemination in a mouse model of peritoneal carcinomatosis. Administering mice with ML3403 and SB203580, potent and selective p38 MAPK inhibitors, attenuate the formation of neoplastic foci induced by intraperitoneal inoculation of gastric cancer cells. By gene array analysis we found that such a protective effect correlates with a robust downregulation in the expression of CXC chemokine receptor-4, Fms-related tyrosine kinase 4 (FLT4), the non-receptor spleen tyrosine kinase (SYK) and the collagen α2(IV) (COL4A2) in neoplasic foci. Inhibition of p38 MAPK in vivo increased the sensitivity of tumor cells to cisplatin and associated with a robust downregulation in the expression of the multidrug resistance (MDR)-1, a well defined marker of resistance to chemotherapy. In summary, p38 MAPK inhibition by a small molecule is beneficial in preventing the peritoneal dissemination of poorly differentiated gastric cancer cells by acting at multiple check-points in the process of attachment and diffusion of tumor cells in the peritoneum.

  11. Live-cell microscopy reveals small molecule inhibitor effects on MAPK pathway dynamics.

    Directory of Open Access Journals (Sweden)

    Daniel J Anderson

    Full Text Available Oncogenic mutations in the mitogen activated protein kinase (MAPK pathway are prevalent in human tumors, making this pathway a target of drug development efforts. Recently, ATP-competitive Raf inhibitors were shown to cause MAPK pathway activation via Raf kinase priming in wild-type BRaf cells and tumors, highlighting the need for a thorough understanding of signaling in the context of small molecule kinase inhibitors. Here, we present critical improvements in cell-line engineering and image analysis coupled with automated image acquisition that allow for the simultaneous identification of cellular localization of multiple MAPK pathway components (KRas, CRaf, Mek1 and Erk2. We use these assays in a systematic study of the effect of small molecule inhibitors across the MAPK cascade either as single agents or in combination. Both Raf inhibitor priming as well as the release from negative feedback induced by Mek and Erk inhibitors cause translocation of CRaf to the plasma membrane via mechanisms that are additive in pathway activation. Analysis of Erk activation and sub-cellular localization upon inhibitor treatments reveals differential inhibition and activation with the Raf inhibitors AZD628 and GDC0879 respectively. Since both single agent and combination studies of Raf and Mek inhibitors are currently in the clinic, our assays provide valuable insight into their effects on MAPK signaling in live cells.

  12. cAMP Modulates Macrophage Development by Suppressing M-CSF-Induced MAPKs Activation

    Institute of Scientific and Technical Information of China (English)

    Ning Zhu; Jian Cui; Chunxia Qiao; Yan Li; Yuanfang Ma; Jiyan Zhang; Beifen Shen

    2008-01-01

    M-CSF is a key cytokine in macrophage development by inducing MAPKs activation, and cAMP can inhibit MAPKs activation induced by inflammatory stimuli. To explore the effects of cAMP on M-CSF-induced MAPKs activation and on macrophage development, the model of bone marrow-derived murine macrophages (BMMs) was used. The effects of cAMP on M-CSF-induced MAPKs activation were analyzed by Western blotting assay, and the effects of cAMP on CD14 and F4/80 expression during macrophage development were examined by FACS analysis.Macrophage morphology showed the successful establishment of the model of macrophage development. Western blotting assay revealed that M-CSF activated ERK, JNK and p38 in both mature and immature macrophages, and cAMP inhibited M-CSF-induced ERK, JNK and p38 activation in a time-dependent manner. FACS analysis revealed that macrophage development was impaired with cAMP pretreatment. In conclusion, cAMP modulates macrophage development by suppressing M-CSF-induced MAPKs activation.

  13. Absence of ERK5/MAPK7 delays tumorigenesis in Atm-/- mice.

    Science.gov (United States)

    Granados-Jaén, Alba; Angulo-Ibáñez, Maria; Rovira-Clavé, Xavier; Gamez, Celina Paola Vasquez; Soriano, Francesc X; Reina, Manuel; Espel, Enric

    2016-11-15

    Ataxia-telangiectasia mutated (ATM) is a cell cycle checkpoint kinase that upon activation by DNA damage leads to cell cycle arrest and DNA repair or apoptosis. The absence of Atm or the occurrence of loss-of-function mutations in Atm predisposes to tumorigenesis. MAPK7 has been implicated in numerous types of cancer with pro-survival and pro-growth roles in tumor cells, but its functional relation with tumor suppressors is not clear. In this study, we show that absence of MAPK7 delays death due to spontaneous tumor development in Atm-/- mice. Compared with Atm-/- thymocytes, Mapk7-/-Atm-/- thymocytes exhibited an improved response to DNA damage (increased phosphorylation of H2AX) and a restored apoptotic response after treatment of mice with ionizing radiation. These findings define an antagonistic function of ATM and MAPK7 in the thymocyte response to DNA damage, and suggest that the lack of MAPK7 inhibits thymic lymphoma growth in Atm-/- mice by partially restoring the DNA damage response in thymocytes.

  14. Plasticity of the MAPK signaling network in response to mechanical stress.

    Directory of Open Access Journals (Sweden)

    Andrea M Pereira

    Full Text Available Cells display versatile responses to mechanical inputs and recent studies have identified the mitogen-activated protein kinase (MAPK cascades mediating the biological effects observed upon mechanical stimulation. Although, MAPK pathways can act insulated from each other, several mechanisms facilitate the crosstalk between the components of these cascades. Yet, the combinatorial complexity of potential molecular interactions between these elements have prevented the understanding of their concerted functions. To analyze the plasticity of the MAPK signaling network in response to mechanical stress we performed a non-saturating epistatic screen in resting and stretched conditions employing as readout a JNK responsive dJun-FRET biosensor. By knocking down MAPKs, and JNK pathway regulators, singly or in pairs in Drosophila S2R+ cells, we have uncovered unexpected regulatory links between JNK cascade kinases, Rho GTPases, MAPKs and the JNK phosphatase Puc. These relationships have been integrated in a system network model at equilibrium accounting for all experimentally validated interactions. This model allows predicting the global reaction of the network to its modulation in response to mechanical stress. It also highlights its context-dependent sensitivity.

  15. Regulation of the MAPK family members: role of subcellular localization and architectural organization.

    Science.gov (United States)

    Fanger, G R

    1999-07-01

    The members of the mitogen-activated protein kinase (MAPK) family are regulated by a diverse array of extracellular cues ranging from cytokines, growth factors and neuropeptides, which activate cell surface receptors, to stresses such as cold, heat, osmolarity changes and irradiation. The MAPK pathways control genetic expression by modifying transcription factor activity and cue important cell fate decisions including survival, proliferation, and programmed cell death (apoptosis). One interesting feature of the MAPK pathways is that the components are evolutionarily conserved from yeast to human, and many of the pathways are similarly organized and regulated. Unlike previously imagined, architectural organization or the multimeric organization of signaling proteins into complexes which are localized to distinct subcellular regions is an important mechanism that influences the regulation of these pathways. In addition, extracellular stimuli can induce relocalization of specific signal transduction proteins. The formation of multimeric signaling complexes, as well as the dynamic movement of signaling proteins, contribute to determine signaling specificity and efficacy. This review describes what is currently known about the subcellular localization of MAPK pathway signaling proteins and the relocalization that occurs during events associated with activation of the MAPK family members.

  16. Human adenovirus type 19 infection of corneal cells induces p38 MAPK-dependent interleukin-8 expression

    Directory of Open Access Journals (Sweden)

    Chodosh James

    2008-01-01

    Full Text Available Abstract Background Human adenovirus type 19 (HAdV-19 is a major cause of epidemic keratoconjunctivitis, the only ocular adenoviral infection associated with prolonged corneal inflammation. In this study, we investigated the role of p38 mitogen-activated protein kinase (MAPK in HAdV-19 infection, with particular attention to the role of p38 MAPK in the transcriptional control of interleukin-8 (IL-8, a chemokine previously shown to be central to the initiation of adenovirus keratitis. Results We found that infection of corneal cells with HAdV-19 led to activation of p38 MAPK and its downstream targets, HSP-27 and ATF-2, within 15 to 30 minutes post-infection. Infection also induced phosphorylation of IκB and NFκB in a p38 MAPK-dependent fashion. Furthermore, HAdV-19 induced an interaction between p38 MAPK and NFκB-p65, followed by nuclear translocation of activated NFκB-p65 and its binding to the IL-8 promoter. The interaction between p38 MAPK and NFκB-p65 was inhibited in concentration-dependent fashion by SB203580, a chemical inhibitor of p38 MAPK, but not by SP600125, an inhibitor of JNK – another MAPK implicated in chemokine expression by HAdV-19 infected cells. IL-8 gene expression in HAdV-19 infection was significantly reduced in the presence of sequence-specific p38 MAPK siRNA but not control siRNA. Conclusion These results provide the first direct evidence for transcriptional regulation of IL-8 in HAdV-19 infected cells through the activation of the p38 MAPK signaling pathway. The p38 MAPK pathway may play a biologically important role in regulation of IL-8 gene expression in the adenovirus-infected cornea.

  17. MKK3 Was Involved in Larval Settlement of the Barnacle Amphibalanus amphitrite through Activating the Kinase Activity of p38MAPK

    KAUST Repository

    Zhang, Gen

    2013-07-29

    The p38 mitogen-activated protein kinase (p38MAPK) plays a key role in larval settlement of the barnacle Amphibalanus amphitrite. To study the signaling pathway associated with p38MAPK during larval settlement, we sought to identify the upstream kinase of p38MAPK. Three MKKs (MKK3, MKK4 and MKK7) and three MAPKs (p38MAPK, ERK and JNK) in A. amphitrite were cloned and recombinantly expressed in E. coli. Through kinase assays, we found that MKK3, but not MKK4 or MKK7, phosphorylated p38MAPK. Furthermore, MKK3 activity was specific to p38MAPK, as it did not phosphorylate ERK or JNK. To further investigate the functional relationship between MKK3 and p38MAPK in vivo, we studied the localization of phospho-MKK3 (pMKK3) and MKK3 by immunostaining. Consistent with the patterns of p38MAPK and phospho-p38MAPK (pp38MAPK), pMKK3 and MKK3 mainly localized to the antennules of the cyprids. Western blot analysis revealed that pMKK3 levels, like pp38MAPK levels, were elevated at cyprid stage, compared to nauplii and juvenile stages. Moreover, pMKK3 levels increased after treatment with adult barnacle crude extracts, suggesting that MKK3 might mediate the stimulatory effects of adult barnacle extracts on the p38MAPK pathway. © 2013 Zhang et al.

  18. Control of the cell cycle progression by the MAPK Hog1

    Directory of Open Access Journals (Sweden)

    Josep Clotet

    2013-02-01

    Full Text Available Eukaryotic cells coordinate various intracellular activities in response to environmental stresses, activating an adaptive program to maximize the probability of survival and proliferation. Cells transduce diverse cellular stimuli by multiple mitogen-activated protein kinase (MAPK cascades. MAPK are key signal transduction kinases required to respond to stress. A prototypical member of the MAPK family is the yeast high osmolarity glycerol (Hog1. Activation of Hog1 results in the generation of a set of adaptive responses that leads to the modulation of several aspects of cell physiology that are essential for cell survival, such as gene expression, translation, and morphogenesis. This review focuses on the control of cell cycle progression by Hog1 which is critical for cell survival in response to stress conditions.

  19. MAPK/ERK signaling regulates insulin sensitivity to control glucose metabolism in Drosophila.

    Directory of Open Access Journals (Sweden)

    Wei Zhang

    2011-12-01

    Full Text Available The insulin/IGF-activated AKT signaling pathway plays a crucial role in regulating tissue growth and metabolism in multicellular animals. Although core components of the pathway are well defined, less is known about mechanisms that adjust the sensitivity of the pathway to extracellular stimuli. In humans, disturbance in insulin sensitivity leads to impaired clearance of glucose from the blood stream, which is a hallmark of diabetes. Here we present the results of a genetic screen in Drosophila designed to identify regulators of insulin sensitivity in vivo. Components of the MAPK/ERK pathway were identified as modifiers of cellular insulin responsiveness. Insulin resistance was due to downregulation of insulin-like receptor gene expression following persistent MAPK/ERK inhibition. The MAPK/ERK pathway acts via the ETS-1 transcription factor Pointed. This mechanism permits physiological adjustment of insulin sensitivity and subsequent maintenance of circulating glucose at appropriate levels.

  20. Involvement of p38MAPK on the antinociceptive action of myricitrin in mice.

    Science.gov (United States)

    Meotti, Flavia Carla; Posser, Thaís; Missau, Fabiana Cristina; Pizzolatti, Moacir Geraldo; Leal, Rodrigo Bainy; Santos, Adair R S

    2007-09-15

    Previous studies from our group investigated the analgesic and anti-inflammatory properties of the flavonoid myricitrin. Here, we demonstrated the role of interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha) and mitogen-activated protein kinases (MAPKs) on the antinociceptive action of myricitrin. The nociceptive response was evaluated by monitoring biting behaviour following intratecal (i.t.) administration of IL-1beta and TNF-alpha in mice. Western blot analyses of total and phosphorylated MAPKs: p38(MAPK), extracellular-signal regulated kinase (ERK1/2) and c-Jun amino-terminal kinases (JNK1/2) from the spinal cord of mice injected with cytokines were measured. Myricitrin (0.03-30mg/kg) or vehicle (control) was administered 30 min beforehand by intraperitoneal (i.p.) injection. Myricitrin pre-treatment prevented cytokine-induced biting behaviour. The calculated ID(50) of myricitrin were 6.8 (4.6-9.0) and 2.6 (0.3-4.9) mg/kg and maximal inhibition of 83+/-9 and 100+/-0% for IL-1beta and TNF-alpha, respectively. Intrathecal injection of IL-1beta and TNF-alpha significantly increased p38(MAPK) phosphorylation and this was inhibited by myricitrin treatment. Cytokines administration did not alter ERK1/2 and JNK1/2 phosphorylation. Myricitrin prevented cytokine-induced biting behaviour and inhibited p38(MAPK) phosphorylation in response to cytokines stimulation. Taken together, it suggests that the mechanism for antinociceptive action of myricitrin in response to cytokines may involve a blockage on p38(MAPK) pathway. This finding could explain, at least in part, the antinociceptive action of this flavonoid in process like neuropathic and inflammatory chronic pain.

  1. MAPK/ERK signaling pathway-induced hyper-O-GlcNAcylation enhances cancer malignancy.

    Science.gov (United States)

    Zhang, Xinling; Ma, Leina; Qi, Jieqiong; Shan, Hui; Yu, Wengong; Gu, Yuchao

    2015-12-01

    Dysregulated MAPK/ERK signaling is implicated in one-third of human tumors and represents an attractive target for the development of anticancer drugs. Similarly, elevated protein O-GlcNAcylation and O-GlcNAc transferase (OGT) are detected in various cancers and serve as attractive novel cancer-specific therapeutic targets. However, the potential connection between them remains unexplored. Here, a positive correlation was found between the activated MAPK/ERK signaling and hyper-O-GlcNAcylation in various cancer types and inhibition of the MAPK/ERK signaling by 10 µM U0126 significantly decreased the expression of OGT and O-GlcNAcylation in H1299, BPH-1 and DU145 cells; then, the pathway analysis of the potential regulators of OGT obtained from the UCSC Genome Browser was done, and ten downstream targets of ERK pathway were uncovered; the following results showed that ELK1, one of the ten targets of ERK pathway, mediated ERK signaling-induced OGT upregulation; finally, the MTT assay and the soft agar assay showed that the inhibition of MAPK/ERK signaling reduced the promotion effect of hyper-O-GlcNAcylation on cancer cell proliferation and anchorage-independent growth. Taken together, our data originally provided evidence for the regulatory mechanism of hyper-O-GlcNAcylation in tumors, which will be helpful for the development of anticancer drugs targeting to hyper-O-GlcNAcylation. This study also provided a new mechanism by which MAPK/ERK signaling-enhanced cancer malignancy. Altogether, the recently discovered oncogenic factor O-GlcNAc was linked to the classical MAPK/ERK signaling which is essential for the maintenance of malignant phenotype of cancers.

  2. Peroxiredoxins in Regulation of MAPK Signalling Pathways; Sensors and Barriers to Signal Transduction

    Science.gov (United States)

    Latimer, Heather R.; Veal, Elizabeth A.

    2016-01-01

    Peroxiredoxins are highly conserved and abundant peroxidases. Although the thioredoxin peroxidase activity of peroxiredoxin (Prx) is important to maintain low levels of endogenous hydrogen peroxide, Prx have also been shown to promote hydrogen peroxide-mediated signalling. Mitogen activated protein kinase (MAPK) signalling pathways mediate cellular responses to a variety of stimuli, including reactive oxygen species (ROS). Here we review the evidence that Prx can act as both sensors and barriers to the activation of MAPK and discuss the underlying mechanisms involved, focusing in particular on the relationship with thioredoxin. PMID:26813660

  3. The Vibrio parahaemolyticus Type III Secretion Systems manipulate host cell MAPK for critical steps in pathogenesis.

    LENUS (Irish Health Repository)

    Matlawska-Wasowska, Ksenia

    2010-12-01

    Vibrio parahaemolyticus is a food-borne pathogen causing inflammation of the gastrointestinal epithelium. Pathogenic strains of this bacterium possess two Type III Secretion Systems (TTSS) that deliver effector proteins into host cells. In order to better understand human host cell responses to V. parahaemolyticus, the modulation of Mitogen Activated Protein Kinase (MAPK) activation in epithelial cells by an O3:K6 clinical isolate, RIMD2210633, was investigated. The importance of MAPK activation for the ability of the bacterium to be cytotoxic and to induce secretion of Interleukin-8 (IL-8) was determined.

  4. MLK4β functions as a negative regulator of MAPK signaling and cell invasion

    OpenAIRE

    Abi Saab, W F; Brown, M S; Chadee, D N

    2012-01-01

    Mixed lineage kinase (MLK) 4, or MLK4, is a member of the MLK family of mitogen-activated protein kinase kinase kinases (MAP3Ks). Typically, MAP3Ks function to activate the mitogen-activated protein kinase (MAPK)-signaling pathways and regulate different cellular responses. However, here we report that MLK4β, unlike the other MLKs, negatively regulates the activities of the MAPKs, p38, c-Jun N-terminal kinase and extracellular signal-regulated kinase, and the MAP2Ks, MEK3 and 6. Our results s...

  5. Clinical significance of the phosphorylation of MAPK and protein expression of cyclin D1 in human osteosarcoma tissues.

    Science.gov (United States)

    Wu, Jian; Cui, Lei-Lei; Yuan, Jun; Wang, Yuan; Song, Shu

    2017-04-01

    The aim of the present study was to investigate the significance of the phosphorylation of mitogen-activated protein kinase (MAPK) and the protein expression of cyclin D1 in human osteosarcoma tissues. Human osteosarcoma tissue samples were collected from 30 patients, benign bone tumor samples were collected from 30 patients, and normal bone tissues were collected from 10 individuals as controls. Immunohistochemistry was performed to measure the levels of phosphorylated (p)-MAPK and cyclin D1 protein in cases of human osteosarcoma. The results showed that the positive rates of MAPK and cyclin D1 in osteosarcoma were 86.67% (26/30) and 73.00% (22/30), respectively. The positive staining rates of MAPK and cyclin D1 in benign bone tumor tissues were 10.00% (3/30) and 3.30% (1/30), respectively. The positive rate in the normal bone tissues was 0% (0/30), which was significantly lower, compared with that of the cancerous bone tissue. The positive rates of MAPK and cyclin D1 in osteosarcoma were increased (P<0.05), and the expression of cyclin D1 and p‑MAPK were positively correlated. The phosphorylation of MAPK may be important in the development of osteosarcoma, and the overactivation of MAPK may induce high expression of cyclin D1 and induce tumor cells to proliferate continuously.

  6. PDGF-BB-mediated activation of p42(MAPK) is independent of PDGF beta-receptor tyrosine phosphorylation.

    Science.gov (United States)

    Cartel, N J; Liu, J; Wang, J; Post, M

    2001-10-01

    Herein, we investigated the activity of mitogen-activated protein kinase (MAPK), a key component of downstream signaling events, which is activated subsequent to platelet-derived growth factor (PDGF)-BB stimulation. Specifically, p42(MAPK) activity peaked 60 min after addition of PDGF-BB, declined thereafter, and was determined not to be a direct or necessary component of glycosaminoglycan (GAG) synthesis. PDGF-BB also activated MAPK kinase 2 (MAPKK2) but had no effect on MAPKK1 and Raf-1 activity. Chemical inhibition of Janus kinase, phosphatidylinositol 3-kinase, Src kinase, or tyrosine phosphorylation inhibition of the PDGF beta-receptor (PDGFR-beta) did not abrogate PDGF-BB-induced p42(MAPK) activation or its threonine or tyrosine phosphorylation. A dominant negative cytoplasmic receptor for hyaluronan-mediated motility variant 4 (RHAMMv4), a regulator of MAPKK-MAPK interaction and activation, did not inhibit PDGF-BB-induced p42(MAPK) activation nor did a construct expressing PDGFR-beta with cytoplasmic tyrosines mutated to phenylalanine. However, overexpression of a dominant negative PDGFR-beta lacking the cytoplasmic signaling domain abrogated p42(MAPK) activity. These results suggest that PDGF-BB-mediated activation of p42(MAPK) requires the PDGFR-beta but is independent of its tyrosine phosphorylation.

  7. Cross-reactivities of mammalian MAPKs antibodies in rotifer and copepod: Application in mechanistic studies in aquatic ecotoxicology.

    Science.gov (United States)

    Kang, Hye-Min; Jeong, Chang-Bum; Lee, Young Hwan; Cui, Yan-Hong; Kim, Duck-Hyun; Lee, Min-Chul; Kim, Hui-Su; Han, Jeonghoon; Hwang, Dae-Sik; Lee, Su-Jae; Lee, Jae-Seong

    2016-12-21

    The mitogen-activated protein kinases (MAPKs) family is known to mediate various biological processes in response to diverse environmental pollutants. Although MAPKs are well characterized and studied in vertebrates, in invertebrates the cross-reactivities of MAPKs antibodies were not clearly known in response to environmental pollutants due to limited information of antibody epitopes with material resources for invertebrates. In this paper, we performed phylogenetic analysis of MAPKs genes in the marine rotifer Brachionus koreanus and the copepods Paracyclopina nana and Tigriopus japonicus. Also in rotifer and copepods, several studies of Western blot of MAPK signaling pathways were shown in response to environmental pollutants, including multi-walled carbon nanotubes (MWCNTs), water-accommodated fractions (WAFs) of crude oil, and microplastics. This paper will provide a better understanding of the underlying mechanistic scenario in terms of cross-reactivities of mammalian antibodies in rotifer and copepod. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. Overexpression of the mitogen-activated protein kinase gene OsMAPK33 enhances sensitivity to salt stress in rice (Oryza sativa L.)

    Indian Academy of Sciences (India)

    Seong-Kon Lee; Beom-Gi Kim; Taek-Ryoun Kwon; Mi-Jeong Jeong; Sang-Ryeol Park; Jung-Won Lee; Myung-Ok Byun; Hawk-Bin Kwon; Benjamin F Matthews; Choo-Bong Hong; Soo-Chul Park

    2011-03-01

    Mitogen-activated protein kinases (MAPK) signalling cascades are activated by extracellular stimuli such as environmental stresses and pathogens in higher eukaryotic plants. To know more about MAPK signalling in plants, a MAPK cDNA clone, OsMAPK33, was isolated from rice. The gene is mainly induced by drought stress. In phylogenetic analysis, OsMAPK33 (Os02g0148100) showed approximately 47–93% identity at the amino acid level with other plant MAPKs. It was found to exhibit organ-specific expression with relatively higher expression in leaves as compared with roots or stems, and to exist as a single copy in the rice genome. To investigate the biological functions of OsMAPK33 in rice MAPK signalling, transgenic rice plants that either overexpressed or suppressed OsMAPK33 were made. Under dehydration conditions, the suppressed lines showed lower osmotic potential compared with that of wild-type plants, suggesting a role of OsMAPK33 in osmotic homeostasis. Nonetheless, the suppressed lines did not display any significant difference in drought tolerance compared with their wild-type plants. With increased salinity, there was still no difference in salt tolerance between OsMAPK33-suppressed lines and their wild-type plants. However, the overexpressing lines showed greater reduction in biomass accumulation and higher sodium uptake into cells, resulting in a lower K+/Na+ ratio inside the cell than that in the wild-type plants and OsMAPK33-suppressed lines. These results suggest that OsMAPK33 could play a negative role in salt tolerance through unfavourable ion homeostasis. Gene expression profiling of OsMAPK33 transgenic lines through rice DNA chip analysis showed that OsMAPK33 altered expression of genes involved in ion transport. Further characterization of downstream components will elucidate various biological functions of this novel rice MAPK.

  9. Determination of black carbon in fine particles using a semi-continuous method at two sites in the city of Guadalajara, Mexico, during 2007.

    Science.gov (United States)

    Hernández-Mena, Leonel; Saldarriaga-Noreña, Hugo; Murillo-Tovar, Mario A; Amador-Muñoz, Omar; López-López, Alberto; Waliszewski, Stefan M

    2011-09-01

    The black carbon is a pollutant species primarily emitted from the combustion of fossil fuels (diesel). Their concentrations associated to PM2.5 were monitoring at two sites in the city of Guadalajara. From January to May (except April), downtown site shown 2.7, 2.6, 4.0 and 2.3 times higher monthly concentrations. The dry season two showed higher concentrations respect to at least one of the others seasons (p < 0.0001) at each site, probably due to atmospheric conditions less favorable for the dispersal of pollutants. During the 24 h period were observed at the year two peaks of concentrations: the highest morning peak and lower night peak, both probably related to anthropogenic activity.

  10. Microchannel conductivity measurements in microchip for on line monitoring of dephosphorylation rates of organic phosphates using paramagnetic-beads linked alkaline phosphatase.

    Science.gov (United States)

    Kechadi, Mohammed; Sotta, Bruno; Gamby, Jean

    2015-01-01

    This paper presents the use of polymer coated microelectrodes for the realtime conductivity monitoring in a microchannel photoablated through the polymer without contact. Based on this strategy, a small conductometry sensor has been developed to record in time conductivity variation when an enzymatic reaction occurs through the channel. The rate constant determination, k2, for the dephosphorylation of organic phosphate-alkaline phosphatase-superparamagnetic beads complex using chemically different substrates such as adenosine monoesterphosphate, adenosine diphosphate and adenosine triphosphate was taken as an example to demonstrate selectivity and sensivity of the detection scheme. The k2 value measured for each adenosine phosphate decreases from 39 to 30 s(-1) in proportion with the number (3, 2 and 1) of attached phosphate moiety, thus emphasizing the steric hindrance effect on kinetics.

  11. Protein phosphatase complex PP5/PPP2R3C dephosphorylates P-glycoprotein/ABCB1 and down-regulates the expression and function.

    Science.gov (United States)

    Katayama, Kazuhiro; Yamaguchi, Miho; Noguchi, Kohji; Sugimoto, Yoshikazu

    2014-04-01

    P-glycoprotein (P-gp)/ABCB1 is a key molecule of multidrug resistance in cancer. Protein phosphatase (PP) 2A, regulatory subunit B, gamma (PPP2R3C), which is a regulatory subunit of PP2A and PP5, was identified as a binding candidate to P-gp. Immunoprecipitation-western blotting revealed that PP5 and PPP2R3C were coprecipitated with P-gp, while PP2A was not. PP5/PPP2R3C dephosphorylated protein kinase A/protein kinase C-phosphorylation of P-gp. Knockdown of PP5 and/or PPP2R3C increased P-gp expression and lowered the sensitivity to vincristine and doxorubicin. Consequently, our results indicate that PP5/PPP2R3C negatively regulates P-gp expression and function.

  12. iNOS participates in apoptosis of spinal cord neurons via p-BAD dephosphorylation following ischemia/reperfusion (I/R) injury in rat spinal cord.

    Science.gov (United States)

    Li, Yiming; Gu, Jun; Liu, Yuwen; Long, Hao; Wang, Guannan; Yin, Guoyong; Fan, Jin

    2013-06-17

    The pro-apoptotic effect of nitric oxide (NO) has been reported both in vivo and in vitro. Previous studies have revealed that NO, especially which produced by inducible nitric oxide synthase (iNOS), has an important effect on apoptosis of neurons in spinal cord ischemia/reperfusion (I/R) injury. To investigate the role of iNOS in this process, a randomized, controlled study was designed using a classical rat model of ischemic spinal cord injury. Fifty-four male Sprague-Dawley rats were randomly divided into three different groups: a sham-operated group (n=6), a vehicle group (I/R, n=24), and an iNOS inhibitor (aminoguanidine: AG) group (I/R+AG, n=24). Rats were sacrificed 6, 12, 24 and 72 h after reperfusion. We examined neurological motor function evaluated by 'Tarlov's score', assessed alterations in the morphology of spinal cord neurons by transmission electron microscopy (TEM), analyzed expression of iNOS at the levels of mRNA and protein, evaluated local concentrations and cellular locations of other key regulatory proteins, and investigated protein-protein interactions. In the vehicle group, iNOS expression, dephosphorylation of p-BAD (Ser 136), disassociation of BAD from p-BAD/14-3-3 dimers, and release of cytochrome c were all increased compared with the sham group. But in the AG group, all the performances above were decreased compared with the vehicle group. Similarly, rats in the sham group got a maximum score of 5 by Tarlov's motor scores evaluation. While the scores were higher in the AG group compared to the vehicle group because iNOS was inhibited. These results indicate that the activity of iNOS plays a critical role in the apoptosis of spinal cord neurons by influencing the dephosphorylation of p-BAD (Ser 136) and the interaction between BAD and 14-3-3.

  13. Barrier to auto integration factor becomes dephosphorylated during HSV-1 Infection and Can Act as a host defense by impairing viral DNA replication and gene expression.

    Science.gov (United States)

    Jamin, Augusta; Thunuguntla, Prasanth; Wicklund, April; Jones, Clinton; Wiebe, Matthew S

    2014-01-01

    BAF (Barrier to Autointegration Factor) is a highly conserved DNA binding protein that senses poxviral DNA in the cytoplasm and tightly binds to the viral genome to interfere with DNA replication and transcription. To counteract BAF, a poxviral-encoded protein kinase phosphorylates BAF, which renders BAF unable to bind DNA and allows efficient viral replication to occur. Herein, we examined how BAF phosphorylation is affected by herpes simplex virus type 1 (HSV-1) infection and tested the ability of BAF to interfere with HSV-1 productive infection. Interestingly, we found that BAF phosphorylation decreases markedly following HSV-1 infection. To determine whether dephosphorylated BAF impacts HSV-1 productive infection, we employed cell lines stably expressing a constitutively unphosphorylated form of BAF (BAF-MAAAQ) and cells overexpressing wild type (wt) BAF for comparison. Although HSV-1 production in cells overexpressing wtBAF was similar to that in cells expressing no additional BAF, viral growth was reduced approximately 80% in the presence of BAF-MAAAQ. Experiments were also performed to determine the mechanism of the antiviral activity of BAF with the following results. BAF-MAAAQ was localized to the nucleus, whereas wtBAF was dispersed throughout cells prior to infection. Following infection, wtBAF becomes dephosphorylated and relocalized to the nucleus. Additionally, BAF was associated with the HSV-1 genome during infection, with BAF-MAAAQ associated to a greater extent than wtBAF. Importantly, unphosphorylated BAF inhibited both viral DNA replication and gene expression. For example, expression of two regulatory proteins, ICP0 and VP16, were substantially reduced in cells expressing BAF-MAAAQ. However, other viral genes were not dramatically affected suggesting that expression of certain viral genes can be differentially regulated by unphosphorylated BAF. Collectively, these results suggest that BAF can act in a phosphorylation-regulated manner to impair

  14. RhoA and p38 MAPK mediate apoptosis induced by cellular cholesterol depletion.

    Science.gov (United States)

    Calleros, Laura; Lasa, Marina; Rodríguez-Alvarez, Francisco J; Toro, María J; Chiloeches, Antonio

    2006-07-01

    Cholesterol is essential for cell viability, and homeostasis of cellular cholesterol is crucial to various cell functions. Here we examined the effect of cholesterol depletion on apoptosis and the mechanisms underlying this effect in NIH3T3 cells. We show that chronic cholesterol depletion achieved with lipoprotein-deficient serum (LPDS) and 25-hydroxycholesterol (25-HC) treatment resulted in a significant increase in cellular apoptosis and caspase-3 activation. This effect is not due to a deficiency of nonsterol isoprenoids, intermediate metabolites of the cholesterol biosynthetic pathway, but rather to low cholesterol levels, since addition of cholesterol together with LPDS and 25-HC nearly abolished apoptosis, whereas addition of farnesyl pyrophosphate or geranylgeranyl-pyrophosphate did not reverse the cell viability loss induced by LPDS plus 25-HC treatment. These effects were accompanied by an increase in ERK, JNK and p38 MAPK activity. However, only the inhibition of p38 MAPK with the specific inhibitor SB203580 or the overexpression of a kinase defective MKK6 resulted in a significant decrease in apoptosis and caspase-3 cleavage induced by cholesterol depletion. Furthermore, LPDS plus 25-HC increased RhoA activity, and this effect was reversed by addition of exogenous cholesterol. Finally, overexpression of the dominant negative N19RhoA inhibited p38 MAPK phosphorylation and apoptosis induced by low cholesterol levels. Together, our results demonstrate that cholesterol depletion induces apoptosis through a RhoA- and p38 MAPK-dependent mechanism.

  15. Phosphofructokinase-P Modulates P44/42 MAPK Levels in HeLa Cells.

    Science.gov (United States)

    Cardim Pires, Thyago Rubens; Albanese, Jamille Mansur; Schwab, Michael; Marette, André; Carvalho, Renato Sampaio; Sola-Penna, Mauro; Zancan, Patricia

    2017-05-01

    It is known that interfering with glycolysis leads to profound modification of cancer cell proliferation. However, energy production is not the major reason for this correlation. Here, using HeLa cells as a model for cancer, we demonstrate that phosphofructokinase-P (PFK-P), which is overexpressed in diverse types of cancer including HeLa cells, modulates expression of P44/42 mitogen-activated protein kinase (MAPK). Silencing of PFK-P did not alter HeLa cell viability or energy production, including the glycolytic rate. On the other hand, silencing of PFK-P induced the downregulation of p44/42 MAPK, augmenting the sensitivity of HeLa cells to different drugs. Conversely, overexpression of PFK-P promotes the upregulation of p44/42 MAPK, making the cells more resistant to the drugs. These results indicate that overexpression of PFK-P by cancer cells is related to activation of survival pathways via upregulation of MAPK and suggest PFK-P as a promising target for cancer therapy. J. Cell. Biochem. 118: 1216-1226, 2017. © 2016 Wiley Periodicals, Inc.

  16. In Vitro Manganese Exposure Disrupts MAPK Signaling Pathways in Striatal and Hippocampal Slices from Immature Rats

    Directory of Open Access Journals (Sweden)

    Tanara Vieira Peres

    2013-01-01

    Full Text Available The molecular mechanisms mediating manganese (Mn-induced neurotoxicity, particularly in the immature central nervous system, have yet to be completely understood. In this study, we investigated whether mitogen-activated protein kinases (MAPKs and tyrosine hydroxylase (TH could represent potential targets of Mn in striatal and hippocampal slices obtained from immature rats (14 days old. The aim of this study was to evaluate if the MAPK pathways are modulated after subtoxic Mn exposure, which do not significantly affect cell viability. The concentrations of manganese chloride (MnCl2; 10–1,000 μM caused no change in cell viability in slices exposed for 3 or 6 hours. However, Mn exposure significantly increased extracellular signal-regulated kinase (ERK 1/2, as well as c-Jun N-terminal kinase (JNK 1/2/3 phosphorylation at both 3 and 6 hours incubations, in both brain structures. Furthermore, Mn exposure did not change the total content or phosphorylation of TH at the serine 40 site in striatal slices. Thus, Mn at concentrations that do not disrupt cell viability causes activation of MAPKs (ERK1/2 and JNK1/2/3 in immature hippocampal and striatal slices. These findings suggest that altered intracellular MAPKs signaling pathways may represent an early event concerning the effects of Mn in the immature brain.

  17. The effect of p38MAPK on cyclic stretch in human facial hypertrophic scar fibroblast differentiation.

    Directory of Open Access Journals (Sweden)

    Qi-cui Du

    Full Text Available Hypertrophic scars (HTS, the excessive deposition of scar tissue by fibroblasts, is one of the most common skin disorders. Fibroblasts derived from surgical scar tissue produce high levels of α-smooth muscle actin (α-SMA and transforming growth factor-β1 (TGF-β1. However, the molecular mechanisms for this phenomenon is poorly understood. Thus, the purpose of this study was to evaluate the molecular mechanisms of HTS and their potential therapeutic implications. Fibroblasts derived from skin HTS were cultured and characterized in vitro. The fibroblasts were synchronized and randomly assigned to two groups: cyclic stretch and cyclic stretch pre-treated with SB203580 (a p38MAPK inhibitor. Cyclic stretch at 10% strain was applied at a loading frequency of 10 cycles per minute (i.e. 5 seconds of tension and 5 seconds of relaxation for 0 h, 6 h and 12 h. Cyclic stretch on HTS fibroblasts led to an increase in the expression of α-SMA and TGF-β1 mRNA and protein and the phosphorylation of p38MAPK. SB203580 reversed these effects and caused a decrease in matrix contraction. Furthermore, HTS fibroblast growth was partially blocked by p38MAPK inhibition. Therefore, the mechanism of cyclic stretch involves p38 MAPK, and its inhibition is suggested as a novel therapeutic strategy for HTS.

  18. The effect of p38MAPK on cyclic stretch in human facial hypertrophic scar fibroblast differentiation.

    Science.gov (United States)

    Du, Qi-cui; Zhang, Dai-zun; Chen, Xiu-juan; Lan-Sun, Gui; Wu, Min; Xiao, Wen-lin

    2013-01-01

    Hypertrophic scars (HTS), the excessive deposition of scar tissue by fibroblasts, is one of the most common skin disorders. Fibroblasts derived from surgical scar tissue produce high levels of α-smooth muscle actin (α-SMA) and transforming growth factor-β1 (TGF-β1). However, the molecular mechanisms for this phenomenon is poorly understood. Thus, the purpose of this study was to evaluate the molecular mechanisms of HTS and their potential therapeutic implications. Fibroblasts derived from skin HTS were cultured and characterized in vitro. The fibroblasts were synchronized and randomly assigned to two groups: cyclic stretch and cyclic stretch pre-treated with SB203580 (a p38MAPK inhibitor). Cyclic stretch at 10% strain was applied at a loading frequency of 10 cycles per minute (i.e. 5 seconds of tension and 5 seconds of relaxation) for 0 h, 6 h and 12 h. Cyclic stretch on HTS fibroblasts led to an increase in the expression of α-SMA and TGF-β1 mRNA and protein and the phosphorylation of p38MAPK. SB203580 reversed these effects and caused a decrease in matrix contraction. Furthermore, HTS fibroblast growth was partially blocked by p38MAPK inhibition. Therefore, the mechanism of cyclic stretch involves p38 MAPK, and its inhibition is suggested as a novel therapeutic strategy for HTS.

  19. Regulation of mat responses by a differentiation MAPK pathway in Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Sheelarani Karunanithi

    Full Text Available Fungal species exhibit diverse behaviors when presented with extracellular challenges. Pathogenic fungi can undergo cell differentiation and biofilm formation in response to fluctuating nutrient levels, and these responses are required for virulence. In the model fungal eukaryote Saccharomyces cerevisiae, nutrient limitation induces filamentous growth and biofilm/mat formation. Both responses require the same signal transduction (MAPK pathway and the same cell adhesion molecule (Flo11 but have been studied under different conditions. We found that filamentous growth and mat formation are aspects of a related response that is regulated by the MAPK pathway. Cells in yeast-form mats differentiated into pseudohyphae in response to nutrient limitation. The MAPK pathway regulated mat expansion (in the plane of the XY-axis and substrate invasion (downward in the plane of the Z-axis, which optimized the mat's response to extracellular nutrient levels. The MAPK pathway also regulated an upward growth pattern (in the plane of the Z-axis in response to nutrient limitation and changes in surface rigidity. Upward growth allowed for another level of mat responsiveness and resembled a type of colonial chemorepulsion. Together our results show that signaling pathways play critical roles in regulating social behaviors in which fungal cells participate. Signaling pathways may regulate similar processes in pathogens, whose highly nuanced responses are required for virulence.

  20. Regulation of the expression of the whole genome of Ustilago maydis by a MAPK pathway.

    Science.gov (United States)

    Martínez-Soto, Domingo; Ruiz-Herrera, José

    2015-05-01

    The operation of mitogen-activated protein kinase (MAPK) signal transduction pathways is one of the most important mechanisms for the transfer of extracellular information into the cell. These pathways are highly conserved in eukaryotic organisms. In fungi, MAPK pathways are involved in the regulation of a number of cellular processes such as metabolism, homeostasis, pathogenesis and cell differentiation and morphogenesis. Considering the importance of pathways, in the present work we proceeded to identify all the genes that are regulated by the signal transduction pathway involved in mating, pathogenesis and morphogenesis of Ustilago maydis. Accordingly we made a comparison between the transcriptomes from a wild-type strain and an Ubc2 mutant affected in the interacting protein of this pathway by use of microarrays. By this methodology, we identified 939 genes regulated directly or indirectly by the MAPK pathway. Of them, 432 were positively, and 507 were negatively found regulated. By functional grouping, genes encoding cyclin-dependent kinases, transcription factors, proteins involved in signal transduction, in synthesis of wall and cell membrane, and involved in dimorphism were identified as differentially regulated. These data reveal the importance of these global studies, and the large (and unsuspected) number of functions of the fungus under the control of this MAPK, providing clues to the possible mechanisms involved.

  1. In Vitro Manganese Exposure Disrupts MAPK Signaling Pathways in Striatal and Hippocampal Slices from Immature Rats

    Science.gov (United States)

    Peres, Tanara Vieira; Pedro, Daniela Zótico; de Cordova, Fabiano Mendes; Lopes, Mark William; Gonçalves, Filipe Marques; Mendes-de-Aguiar, Cláudia Beatriz Nedel; Walz, Roger; Farina, Marcelo; Aschner, Michael; Leal, Rodrigo Bainy

    2013-01-01

    The molecular mechanisms mediating manganese (Mn)-induced neurotoxicity, particularly in the immature central nervous system, have yet to be completely understood. In this study, we investigated whether mitogen-activated protein kinases (MAPKs) and tyrosine hydroxylase (TH) could represent potential targets of Mn in striatal and hippocampal slices obtained from immature rats (14 days old). The aim of this study was to evaluate if the MAPK pathways are modulated after subtoxic Mn exposure, which do not significantly affect cell viability. The concentrations of manganese chloride (MnCl2; 10–1,000 μM) caused no change in cell viability in slices exposed for 3 or 6 hours. However, Mn exposure significantly increased extracellular signal-regulated kinase (ERK) 1/2, as well as c-Jun N-terminal kinase (JNK) 1/2/3 phosphorylation at both 3 and 6 hours incubations, in both brain structures. Furthermore, Mn exposure did not change the total content or phosphorylation of TH at the serine 40 site in striatal slices. Thus, Mn at concentrations that do not disrupt cell viability causes activation of MAPKs (ERK1/2 and JNK1/2/3) in immature hippocampal and striatal slices. These findings suggest that altered intracellular MAPKs signaling pathways may represent an early event concerning the effects of Mn in the immature brain. PMID:24324973

  2. PTK, MAPK, and NOC/oFQ impair hypercapnic cerebrovasodilation after hypoxia/ischemia.

    Science.gov (United States)

    Jagolino, Amanda L; Armstead, William M

    2003-01-01

    This study characterized the contributions of protein tyrosine kinase (PTK) and mitogen-activated protein kinase (MAPK) in nociceptin/orphanin FQ (NOC/oFQ)-induced impairment of hypercapnic pial artery dilation (PAD) after hypoxia/ischemia (H/I) in piglets equipped with a closed cranial window. NOC/oFQ (10(-10) M cerebrospinal fluid H/I concentration) impaired hypercapnic PAD (21 +/- 2% vs. 13 +/- 1%). Coadministration of either of the PTK inhibitors genistein or tyrphostin A23 or the MAPK inhibitors U-0126 or PD-98059 with NOC/oFQ (10(-10) M) partially prevented the inhibition of hypercapnic PAD compared with that observed in their absence (21 +/- 2% vs. 17 +/- 1% for genistein). After exposure to H/I, PAD in response to hypercapnia was impaired, but pretreatment with either genistein, tyrphostin A23, U-0126, or PD-98059 partially protected such impairment (17 +/- 1% vs. 4 +/- 1% vs. 9 +/- 1% for sham control, H/I, and H/I + genistein pretreatment, respectively). These data show that PTK and MAPK activation contribute to NOC/oFQ-induced impairment of hypercapnic PAD. These data suggest that activation of PTK and MAPK is also involved in the mechanism by which NOC/oFQ impairs hypercapnic PAD after H/I.

  3. Telmisartan ameliorates cisplatin-induced nephrotoxicity by inhibiting MAPK mediated inflammation and apoptosis.

    Science.gov (United States)

    Malik, Salma; Suchal, Kapil; Gamad, Nanda; Dinda, Amit Kumar; Arya, Dharamvir Singh; Bhatia, Jagriti

    2015-02-05

    Nephrotoxicity is a major adverse effect of the widely used anticancer drug cisplatin. Oxidative stress, inflammation and apoptosis are implicated in the pathophysiology of cisplatin-induced acute renal injury. Moreover, cisplatin activates many signal transduction pathways involved in cell injury and death, particularly mitogen activated protein kinase (MAPK) pathway. With this background, we aimed to investigate the protective effect of telmisartan, a widely used antihypertensive drug, in cisplatin-induced nephrotoxicity model in rats. To accomplish this, male albino wistar rats (150-200 g) were divided into 6 groups: Normal, cisplatin-control, telmisartan (2.5, 5 and 10 mg/kg) and telmisartan per se treatment groups. Normal saline or telmisartan was administered orally to rats for 10 days and cisplatin was given on 7th day (8 mg/kg; i.p.) to induce nephrotoxicity. On 10th day, rats were killed and both the kidneys were harvested for biochemical, histopathological and molecular studies. Cisplatin injected rats showed depressed renal function, altered proxidant-antioxidant balance and acute tubular necrosis which was significantly normalized by telmisartan co-treatment. Furthermore, cisplatin administration activated MAPK pathway that caused tubular inflammation and apoptosis in rats. Telmisartan treatment significantly prevented MAPK mediated inflammation and apoptosis. Among the three doses studied telmisartan at 10 mg/kg dose showed maximum nephroprotective effect which could be due to maintenance of cellular redox status and inhibition of MAPK activation. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Proteomic and functional analyses reveal MAPK1 regulates milk protein synthesis.

    Science.gov (United States)

    Lu, Li-Min; Li, Qing-Zhang; Huang, Jian-Guo; Gao, Xue-Jun

    2012-12-27

    L-Lysine (L-Lys) is an essential amino acid that plays fundamental roles in protein synthesis. Many nuclear phosphorylated proteins such as Stat5 and mTOR regulate milk protein synthesis. However, the details of milk protein synthesis control at the transcript and translational levels are not well known. In this current study, a two-dimensional gel electrophoresis (2-DE)/MS-based proteomic technology was used to identify phosphoproteins responsible for milk protein synthesis in dairy cow mammary epithelial cells (DCMECs). The effect of L-Lys on DCMECs was analyzed by CASY technology and reversed phase high performance liquid chromatography (RP-HPLC). The results showed that cell proliferation ability and β-casein expression were enhanced in DCMECs treated with L-Lys. By phosphoproteomics analysis, six proteins, including MAPK1, were identified up-expressed in DCMECs treated with 1.2 mM L-Lys for 24 h, and were verified by quantitative real-time PCR (qRT-PCR) and western blot. Overexpression and siRNA inhibition of MAPK1 experiments showed that MAPK1 upregulated milk protein synthesis through Stat5 and mTOR pathway. These findings that MAPK1 involves in regulation of milk synthesis shed new insights for understanding the mechanisms of milk protein synthesis.

  5. Syntheses and structure-activity relationships for some triazolyl p38 alpha MAPK inhibitors

    NARCIS (Netherlands)

    Seerden, Jean-Paul G.; Leusink-Ionescu, Gabriela; Leguijt, Robin; Saccavini, Catherine; Gelens, Edith; Dros, Bas; Woudenberg-Vrenken, Titia; Molema, Grietje; Kamps, Jan A. A. M.; Kellogg, Richard M.

    2014-01-01

    The design, synthesis and biological evaluation of novel triazolyl p38 alpha MAPK inhibitors with improved water solubility for formulation in cationic liposomes (SAINT-O-Somes) targeted at diseased endothelial cells is described. Water-solubilizing groups were introduced via a 'click' reaction of f

  6. PKG-Mediated MAPK Signaling Is Necessary for Long-Term Operant Memory in "Aplysia"

    Science.gov (United States)

    Michel, Maximilian; Green, Charity L.; Eskin, Arnold; Lyons, Lisa C.

    2011-01-01

    Signaling pathways necessary for memory formation, such as the mitogen-activated protein kinase (MAPK) pathway, appear highly conserved across species and paradigms. Learning that food is inedible (LFI) represents a robust form of associative, operant learning that induces short- (STM) and long-term memory (LTM) in "Aplysia." We investigated the…

  7. Formation of Subnuclear Foci Is a Unique Spatial Behavior of Mating MAPKs during Hyperosmotic Stress

    Directory of Open Access Journals (Sweden)

    Simon E. Vidal

    2013-02-01

    Full Text Available The assembly of signaling components and transcription factors in ordered subcellular structures is increasingly implicated as an important regulatory strategy for modulating the activity of cellular pathways. Here, we document the inducible formation of subnuclear foci formed by two mitogen-activated protein kinases (MAPKs in Saccharomyces cerevisiae upon hyperosmotic stress. Specifically, we demonstrate that activation of the hyperosmotic stress response pathway induces the mating pathway MAPK Fus3 and the filamentation pathway MAPK Kss1 to form foci in the nucleus that are organized by their shared downstream transcription factor Ste12. Foci formation of colocalized Ste12, Fus3, and Kss1 requires the kinase activity of the hyperosmotic response MAPK Hog1 and correlates with attenuated signaling in the mating pathway. Conversely, activation of the mating pathway prevents foci formation upon subsequent hyperosmotic stress. These results suggest that Hog1-mediated spatial localization of Fus3 and Ste12 into subnuclear foci could contribute to uncoupling the pheromone and osmolarity pathways, which share signaling components, under high-osmolarity conditions.

  8. Down-regulation of HIV-1 Infection by Inhibition of the MAPK Signaling Pathway

    Institute of Scientific and Technical Information of China (English)

    Jian Gong; Xi-hui Shen; Chao Chen; Hui Qiu; Rong-ge Yang

    2011-01-01

    The human immunodeficiency virus type 1(HIV-1)can interact with and exploit the host cellular machinery to replicate and propagate itself.Numerous studies have shown that the Mitogen-activated protein kinase(MAPK)signal pathway can positively regulate the replication of HIV-1,but exactly how each MAPK pathway affects HIV-1 infection and replication is not understood.In this study,we used the Extracellular signal-regulated kinase(ERK)pathway inhibitor,PD98059,the Jun N-terminal kinase(JNK)pathway inhibitor,SP600125,and the p38 pathway inhibitor,SB203580,to investigate the roles of these pathways in HIV-1replication.We found that application of PD98059 results in a strong VSV-G pseudotyped HIV-1NL4-3 luciferase reporter virus and HIV-1NL4-3 virus inhibition activity.In addition,SB203580 and SP600125 also elicited marked VSV-G pseudotyped HIV-1NL4-3 luciferase reporter virus inhibition activity but no HIV-1NL4-3 virus inhibition activity.We also found that SB203580 and SP600125 can enhance the HIV-1 inhibition activity of PD98059when cells were treated with all three MAPK pathway inhibitors in combination.Finally,we show that HIV-1virus inhibition activity of the MAPK pathway inhibitors was the result of the negative regulation of HIV-1 LTR promoter activity.

  9. p38-MAPK inhibition and endotoxin induced tubular dysfunction in men

    NARCIS (Netherlands)

    Zijlstra, JG; Tulleken, JE; Ligtenberg, JJM; de Boer, P; van der Werf, TS

    2004-01-01

    Background: To evaluate the possibility of preventing endotoxin induced renal damage by p38-MAPK inhibition in a human model. Design and Methods: Twenty-one healthy young male volunteers received 4 ng/kg Escherichia coli endotoxin as a single dose. Four groups of volunteers received an oral dose of

  10. Over-Expression of BnMAPK1 in Brassica napus Enhances Tolerance to Drought Stress

    Institute of Scientific and Technical Information of China (English)

    WENG Chang-mei; LU Jun-xing; WAN Hua-fang; WANG Shu-wen; WANG Zhen; LU Kun; LIANG Ying

    2014-01-01

    Mitogen-activated protein kinases (MAPKs) are a family of Ser/Thr protein kinases widely conserved in all eukaryotes and involved in responses to biotic and abiotic stresses. In this study, two over-expressing BnMAPK1 oilseed rape lines, ov3 and ov11, were used to study the drought-resistant mechanism of BnMAPK1 under natural drought and simulation drought through spraying 10%PEG 8000 in seedlings. Zhongyou 821 (WT) was used as control. Compared with wild type, transgenic seedlings had higher leaf water content, higher root activity, slightly higher peroxidase (POD) and superoxide dismutase (SOD) activity, higher proline content and lower malondialdehyde (MDA) content. The expression of drought-resistant related genes, including P5CSB, PLC, LEA4 and SCE1, have been up-regulated in some degree and the expressed time of transgenic lines were earlier than that of wild type. These results suggested that over-expression of BnMAPK1 can enhance the resistance to drought in oilseed rape (Brassica napus).

  11. Pheromone-induced morphogenesis improves osmoadaptation capacity by activating the HOG MAPK pathway.

    Science.gov (United States)

    Baltanás, Rodrigo; Bush, Alan; Couto, Alicia; Durrieu, Lucía; Hohmann, Stefan; Colman-Lerner, Alejandro

    2013-04-23

    Environmental and internal conditions expose cells to a multiplicity of stimuli whose consequences are difficult to predict. We investigate the response to mating pheromone of yeast cells adapted to high osmolarity. Events downstream of pheromone binding involve two mitogen-activated protein kinase (MAPK) cascades: the pheromone response (PR) and the cell wall integrity (CWI) response. Although the PR MAPK pathway shares components with a third MAPK pathway, the high osmolarity (HOG) response, each one is normally only activated by its cognate stimulus, a phenomenon called insulation. We found that in cells adapted to high osmolarity, PR activated the HOG pathway in a pheromone- and osmolarity-dependent manner. Activation of HOG by the PR was not due to loss of insulation, but rather a response to a reduction in internal osmolarity, which resulted from an increase in glycerol release caused by the PR. By analyzing single-cell time courses, we found that stimulation of HOG occurred in discrete bursts that coincided with the "shmooing" morphogenetic process. Activation required the polarisome, the CWI MAPK Slt2, and the aquaglyceroporin Fps1. HOG activation resulted in high glycerol turnover, which improved adaptability to rapid changes in osmolarity. Our work shows how a differentiation signal can recruit a second, unrelated sensory pathway to fine-tune yeast response in a complex environment.

  12. Role of ERK/MAPK in endothelin receptor signaling in human aortic smooth muscle cells

    DEFF Research Database (Denmark)

    Chen, Qing-wen; Edvinsson, Lars; Xu, Cang-Bao

    2009-01-01

    muscle cells (VSMCs) through activation of endothelin type A (ETA) and type B (ETB) receptors. The extracellular signal-regulated kinase 1 and 2 (ERK1/2) mitogen-activated protein kinases (MAPK) are involved in ET-1-induced VSMC contraction and proliferation. This study was designed to investigate...

  13. SlMAPK3 enhances tolerance to tomato yellow leaf curl virus (TYLCV) by regulating salicylic acid and jasmonic acid signaling in tomato (Solanum lycopersicum).

    Science.gov (United States)

    Li, Yunzhou; Qin, Lei; Zhao, Jingjing; Muhammad, Tayeb; Cao, Hehe; Li, Hailiang; Zhang, Yan; Liang, Yan

    2017-01-01

    Several recent studies have reported on the role of mitogen-activated protein kinase (MAPK3) in plant immune responses. However, little is known about how MAPK3 functions in tomato (Solanum lycopersicum L.) infected with tomato yellow leaf curl virus (TYLCV). There is also uncertainty about the connection between plant MAPK3 and the salicylic acid (SA) and jasmonic acid (JA) defense-signaling pathways. The results of this study indicated that SlMAPK3 participates in the antiviral response against TYLCV. Tomato seedlings were inoculated with TYLCV to investigate the possible roles of SlMAPK1, SlMAPK2, and SlMAPK3 against this virus. Inoculation with TYLCV strongly induced the expression and the activity of all three genes. Silencing of SlMAPK1, SlMAPK2, and SlMAPK3 reduced tolerance to TYLCV, increased leaf H2O2 concentrations, and attenuated expression of defense-related genes after TYLCV infection, especially in SlMAPK3-silenced plants. Exogenous SA and methyl jasmonic acid (MeJA) both significantly induced SlMAPK3 expression in tomato leaves. Over-expression of SlMAPK3 increased the transcript levels of SA/JA-mediated defense-related genes (PR1, PR1b/SlLapA, SlPI-I, and SlPI-II) and enhanced tolerance to TYLCV. After TYLCV inoculation, the leaves of SlMAPK3 over-expressed plants compared with wild type plants showed less H2O2 accumulation and greater superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX) activity. Overall, the results suggested that SlMAPK3 participates in the antiviral response of tomato to TYLCV, and that this process may be through either the SA or JA defense-signaling pathways.

  14. SlMAPK3 enhances tolerance to tomato yellow leaf curl virus (TYLCV) by regulating salicylic acid and jasmonic acid signaling in tomato (Solanum lycopersicum)

    Science.gov (United States)

    Li, Yunzhou; Qin, Lei; Zhao, Jingjing; Muhammad, Tayeb; Cao, Hehe; Li, Hailiang; Zhang, Yan; Liang, Yan

    2017-01-01

    Several recent studies have reported on the role of mitogen-activated protein kinase (MAPK3) in plant immune responses. However, little is known about how MAPK3 functions in tomato (Solanum lycopersicum L.) infected with tomato yellow leaf curl virus (TYLCV). There is also uncertainty about the connection between plant MAPK3 and the salicylic acid (SA) and jasmonic acid (JA) defense-signaling pathways. The results of this study indicated that SlMAPK3 participates in the antiviral response against TYLCV. Tomato seedlings were inoculated with TYLCV to investigate the possible roles of SlMAPK1, SlMAPK2, and SlMAPK3 against this virus. Inoculation with TYLCV strongly induced the expression and the activity of all three genes. Silencing of SlMAPK1, SlMAPK2, and SlMAPK3 reduced tolerance to TYLCV, increased leaf H2O2 concentrations, and attenuated expression of defense-related genes after TYLCV infection, especially in SlMAPK3-silenced plants. Exogenous SA and methyl jasmonic acid (MeJA) both significantly induced SlMAPK3 expression in tomato leaves. Over-expression of SlMAPK3 increased the transcript levels of SA/JA-mediated defense-related genes (PR1, PR1b/SlLapA, SlPI-I, and SlPI-II) and enhanced tolerance to TYLCV. After TYLCV inoculation, the leaves of SlMAPK3 over-expressed plants compared with wild type plants showed less H2O2 accumulation and greater superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX) activity. Overall, the results suggested that SlMAPK3 participates in the antiviral response of tomato to TYLCV, and that this process may be through either the SA or JA defense-signaling pathways. PMID:28222174

  15. Angiotensin II upregulates endothelial lipase expression via the NF-kappa B and MAPK signaling pathways.

    Directory of Open Access Journals (Sweden)

    Xiaoli Zhang

    Full Text Available BACKGROUND: Angiotensin II (AngII participates in endothelial damage and inflammation, and accelerates atherosclerosis. Endothelial lipase (EL is involved in the metabolism and clearance of high density lipoproteins (HDL, the serum levels of which correlate negatively with the onset of cardiovascular diseases including atherosclerosis. However, the relationship between AngII and EL is not yet fully understood. In this study, we investigated the effects of AngII on the expression of EL and the signaling pathways that mediate its effects in human umbilical vein endothelial cells (HUVECs. METHODS AND FINDINGS: HUVECs were cultured in vitro with different treatments as follows: 1 The control group without any treatment; 2 AngII treatment for 0 h, 4 h, 8 h, 12 h and 24 h; 3 NF-κB activation inhibitor pyrrolidine dithiocarbamate (PDTC pretreatment for 1 h before AngII treatment; and 4 mitogen-activated protein kinase (MAPK p38 inhibitor (SB203580 pretreatment for 1 h before AngII treatment. EL levels in each group were detected by immunocytochemical staining and western blotting. HUVECs proliferation was detected by MTT and proliferating cell nuclear antigen (PCNA immunofluorescence staining. NF-kappa B (NF-κB p65, MAPK p38, c-Jun N-terminal kinase (JNK, extracellular signal-regulated kinase (ERK and phosphorylated extracellular signal-regulated kinase (p-ERK expression levels were assayed by western blotting. The results showed that the protein levels of EL, NF-κB p65, MAPK p38, JNK, and p-ERK protein levels, in addition to the proliferation of HUVECs, were increased by AngII. Both the NF-kB inhibitor (PDTC and the MAPK p38 inhibitor (SB203580 partially inhibited the effects of AngII on EL expression. CONCLUSION: AngII may upregulate EL protein expression via the NF-κB and MAPK signaling pathways.

  16. The Role of MAPK Modules and ABA during Abiotic Stress Signaling

    KAUST Repository

    Zélicourt, Axel de

    2016-05-01

    To respond to abiotic stresses, plants have developed specific mechanisms that allow them to rapidly perceive and respond to environmental changes. The phytohormone abscisic acid (ABA) was shown to be a pivotal regulator of abiotic stress responses in plants, triggering major changes in plant physiology. The ABA core signaling pathway largely relies on the activation of SnRK2 kinases to mediate several rapid responses, including gene regulation, stomatal closure, and plant growth modulation. Mitogen-activated protein kinases (MAPKs) have also been implicated in ABA signaling, but an entire ABA-activated MAPK module was uncovered only recently. In this review, we discuss the evidence for a role of MAPK modules in the context of different plant ABA signaling pathways. Abiotic stresses impact average yield in agriculture by more than 50% globally.Since ABA is a key regulator of abiotic stress responses, an understanding of its functioning at the molecular level is essential for plant breeding. Although the ABA core signaling pathway has been unraveled, several downstream events are still unclear.MAPKs are involved in most plant developmental stages and in response to stresses. Several members of the MAPK family were shown to be directly or indirectly activated by the ABA core signaling pathway.Recent evidence shows that the complete MAP3K17/18-MKK3-MPK1/2/7/14 module is under the control of ABA, whose members are under the transcriptional and post-translational control of the ABA core signaling pathway. © 2016 Elsevier Ltd.

  17. Salidroside Protects against Cadmium-Induced Hepatotoxicity in Rats via GJIC and MAPK Pathways.

    Science.gov (United States)

    Zou, Hui; Liu, Xuezhong; Han, Tao; Hu, Di; Wang, Yi; Yuan, Yan; Gu, Jianhong; Bian, Jianchun; Zhu, Jiaqiao; Liu, Zong-ping

    2015-01-01

    It is known that cadmium (Cd) induces cytotoxicity in hepatocytes; however, the underlying mechanism is unclear. Here, we studied the molecular mechanisms of Cd-induced hepatotoxicity in rat liver cells (BRL 3A) and in vivo. We observed that Cd treatment was associated with a time- and concentration-dependent decrease in the cell index (CI) of BRL 3A cells and cellular organelle ultrastructure injury in the rat liver. Meanwhile, Cd treatment resulted in the inhibition of gap junction intercellular communication (GJIC) and activation of mitogen-activated protein kinase (MAPK) pathways. Gap junction blocker 18-β-glycyrrhetinic acid (GA), administered in combination with Cd, exacerbated cytotoxic injury in BRL 3A cells; however, GA had a protective effect on healthy cells co-cultured with Cd-exposed cells in a co-culture system. Cd-induced cytotoxic injury could be attenuated by co-treatment with an extracellular signal-regulated kinase (ERK) inhibitor (U0126) and a p38 inhibitor (SB202190) but was not affected by co-treatment with a c-Jun N-terminal kinase (JNK) inhibitor (SP600125). These results indicate that ERK and p38 play critical roles in Cd-induced hepatotoxicity and mediate the function of gap junctions. Moreover, MAPKs induce changes in GJIC by controlling connexin gene expression, while GJIC has little effect on the Cd-induced activation of MAPK pathways. Collectively, our study has identified a possible mechanistic pathway of Cd-induced hepatotoxicity in vitro and in vivo, and identified the participation of GJIC and MAPK-mediated pathways in Cd-induced hepatotoxicity. Furthermore, we have shown that salidroside may be a functional chemopreventative agent that ameliorates the negative effects of Cd via GJIC and MAPK pathways.

  18. Differential contribution of hypothalamic MAPK activity to anxiety-like behaviour in virgin and lactating rats.

    Directory of Open Access Journals (Sweden)

    Benjamin Jurek

    Full Text Available The c-Raf - MEK1/2 - ERK1/2 mitogen-activated protein kinase (MAPK intracellular signalling cascade in neurons plays important roles in the control of a variety of behaviours, including social behaviours and anxiety. These roles partially overlap with those described for oxytocin (OXT, and it has been shown that OXT activates the MAPK pathway in the hypothalamus (of male, and hippocampus (of female rats. Here, by combining behavioural (light/dark box and biochemical analyses (western blotting, we tested two hypotheses: (i that OXT is anxiolytic within the hypothalamus of females, and (ii that this effect, as well as that of lactation-associated anxiolysis, depends on the recruitment of the MAPK pathway. We found that, when injected bilaterally into the hypothalamic paraventricular nucleus (PVN, OXT decreased anxiety-like behaviour in virgins, and that this effect depended on phosphorylation of MEK1/2. MAPK pathway activation in lactation was evident by high phosphorylated (p MEK1/2 levels, and nuclear translocation of ERK1. The high pMEK1/2 levels were necessary for the anxiolytic phenotype typically observed during lactation. Interestingly, exogenous OXT in lactating rats reduced pMEK1/2 levels without a concomitant effect on anxiety, indicating that OXT receptor activation can lead to recruitment of additional intracellular pathways to modulate MEK activity. Still other pathways could include MEK, but without subsequent activation of ERK, as we did not observe any increase in OXT-induced ERK phosphorylation. Together the results demonstrate that the MAPK pathway, especially MEK1/2, is critically involved in the regulation of anxiety-like behaviour in female rats.

  19. Salidroside Protects against Cadmium-Induced Hepatotoxicity in Rats via GJIC and MAPK Pathways.

    Directory of Open Access Journals (Sweden)

    Hui Zou

    Full Text Available It is known that cadmium (Cd induces cytotoxicity in hepatocytes; however, the underlying mechanism is unclear. Here, we studied the molecular mechanisms of Cd-induced hepatotoxicity in rat liver cells (BRL 3A and in vivo. We observed that Cd treatment was associated with a time- and concentration-dependent decrease in the cell index (CI of BRL 3A cells and cellular organelle ultrastructure injury in the rat liver. Meanwhile, Cd treatment resulted in the inhibition of gap junction intercellular communication (GJIC and activation of mitogen-activated protein kinase (MAPK pathways. Gap junction blocker 18-β-glycyrrhetinic acid (GA, administered in combination with Cd, exacerbated cytotoxic injury in BRL 3A cells; however, GA had a protective effect on healthy cells co-cultured with Cd-exposed cells in a co-culture system. Cd-induced cytotoxic injury could be attenuated by co-treatment with an extracellular signal-regulated kinase (ERK inhibitor (U0126 and a p38 inhibitor (SB202190 but was not affected by co-treatment with a c-Jun N-terminal kinase (JNK inhibitor (SP600125. These results indicate that ERK and p38 play critical roles in Cd-induced hepatotoxicity and mediate the function of gap junctions. Moreover, MAPKs induce changes in GJIC by controlling connexin gene expression, while GJIC has little effect on the Cd-induced activation of MAPK pathways. Collectively, our study has identified a possible mechanistic pathway of Cd-induced hepatotoxicity in vitro and in vivo, and identified the participation of GJIC and MAPK-mediated pathways in Cd-induced hepatotoxicity. Furthermore, we have shown that salidroside may be a functional chemopreventative agent that ameliorates the negative effects of Cd via GJIC and MAPK pathways.

  20. Salidroside Protects against Cadmium-Induced Hepatotoxicity in Rats via GJIC and MAPK Pathways

    Science.gov (United States)

    Han, Tao; Hu, Di; Wang, Yi; Yuan, Yan; Gu, Jianhong; Bian, Jianchun; Zhu, Jiaqiao; Liu, Zong-ping

    2015-01-01

    It is known that cadmium (Cd) induces cytotoxicity in hepatocytes; however, the underlying mechanism is unclear. Here, we studied the molecular mechanisms of Cd-induced hepatotoxicity in rat liver cells (BRL 3A) and in vivo. We observed that Cd treatment was associated with a time- and concentration-dependent decrease in the cell index (CI) of BRL 3A cells and cellular organelle ultrastructure injury in the rat liver. Meanwhile, Cd treatment resulted in the inhibition of gap junction intercellular communication (GJIC) and activation of mitogen-activated protein kinase (MAPK) pathways. Gap junction blocker 18-β-glycyrrhetinic acid (GA), administered in combination with Cd, exacerbated cytotoxic injury in BRL 3A cells; however, GA had a protective effect on healthy cells co-cultured with Cd-exposed cells in a co-culture system. Cd-induced cytotoxic injury could be attenuated by co-treatment with an extracellular signal-regulated kinase (ERK) inhibitor (U0126) and a p38 inhibitor (SB202190) but was not affected by co-treatment with a c-Jun N-terminal kinase (JNK) inhibitor (SP600125). These results indicate that ERK and p38 play critical roles in Cd-induced hepatotoxicity and mediate the function of gap junctions. Moreover, MAPKs induce changes in GJIC by controlling connexin gene expression, while GJIC has little effect on the Cd-induced activation of MAPK pathways. Collectively, our study has identified a possible mechanistic pathway of Cd-induced hepatotoxicity in vitro and in vivo, and identified the participation of GJIC and MAPK-mediated pathways in Cd-induced hepatotoxicity. Furthermore, we have shown that salidroside may be a functional chemopreventative agent that ameliorates the negative effects of Cd via GJIC and MAPK pathways. PMID:26070151

  1. Tight interconnection and multi-level control of Arabidopsis MYB44 in MAPK cascade signalling.

    Science.gov (United States)

    Persak, Helene; Pitzschke, Andrea

    2013-01-01

    Abiotic stress poses a huge, ever-increasing problem to plants and agriculture. The dissection of signalling pathways mediating stress tolerance is a prerequisite to develop more resistant plant species. Mitogen-activated protein kinase (MAPK) cascades are universal signalling modules. In Arabidopsis, the MAPK MPK3 and its upstream regulator MAPK kinase MKK4 initiate the adaptation response to numerous abiotic and biotic stresses. Yet, molecular steps directly linked with MKK4-MPK3 activation are largely unknown. Starting with a yeast-two-hybrid screen for interacting partners of MKK4, we identified a transcription factor, MYB44. MYB44 is controlled at multiple levels by and strongly inter-connected with MAPK signalling. As we had shown earlier, stress-induced expression of the MYB44 gene is regulated by a MPK3-targeted bZIP transcription factor VIP1. At the protein level, MYB44 interacts with MPK3 in vivo. MYB44 is phosphorylated by MPK3 in vitro at a single residue, Ser145. Although replacement of Ser145 by a non-phosphorylatable (S145A) or phosphomimetic (S145D) residue did not alter MYB44 subcellular localisation, dimerization behaviour nor DNA-binding characteristics, abiotic stress tolerance tests in stable transgenic Arabidopsis plants clearly related S145 phosphorylation to MYB44 function: Compared to Arabidopsis wild type plants, MYB44 overexpressing lines exhibit an enhanced tolerance to osmotic stress and are slightly more sensitive to abscisic acid. Interestingly, overexpression of the S145A variant revealed that impaired phosphorylation does not render the MYB44 protein non-functional. Instead, S145A lines are highly sensitive to abiotic stress, and thereby remarkably similar to mpk3-deficient plants. Its in vivo interaction with the nuclear sub-pools of both MPK3 and MKK4 renders MYB44 the first plant transcription factor to have a second function as putative MAPK cascade scaffolding protein.

  2. Mouse preimplantation embryo responses to culture medium osmolarity include increased expression of CCM2 and p38 MAPK activation

    Directory of Open Access Journals (Sweden)

    Watson Andrew J

    2007-01-01

    Full Text Available Abstract Background Mechanisms that confer an ability to respond positively to environmental osmolarity are fundamental to ensuring embryo survival during the preimplantation period. Activation of p38 mitogen-activated protein kinase (MAPK occurs following exposure to hyperosmotic treatment. Recently, a novel scaffolding protein called Osmosensing Scaffold for MEKK3 (OSM was linked to p38 MAPK activation in response to sorbitol-induced hypertonicity. The human ortholog of OSM is cerebral cavernous malformation 2 (CCM2. The present study was conducted to investigate whether CCM2 is expressed during mouse preimplantation development and to determine whether this scaffolding protein is associated with p38 MAPK activation following exposure of preimplantation embryos to hyperosmotic environments. Results Our results indicate that Ccm2 along with upstream p38 MAPK pathway constituents (Map3k3, Map2k3, Map2k6, and Map2k4 are expressed throughout mouse preimplantation development. CCM2, MAP3K3 and the phosphorylated forms of MAP2K3/MAP2K6 and MAP2K4 were also detected throughout preimplantation development. Embryo culture in hyperosmotic media increased p38 MAPK activity in conjunction with elevated CCM2 levels. Conclusion These results define the expression of upstream activators of p38 MAPK during preimplantation development and indicate that embryo responses to hyperosmotic environments include elevation of CCM2 and activation of p38 MAPK.

  3. Impact of MAPK Pathway Activation in BRAF(V600) Melanoma on T Cell and Dendritic Cell Function.

    Science.gov (United States)

    Ott, Patrick A; Bhardwaj, Nina

    2013-10-28

    Constitutive upregulation of the MAPK pathway by a BRAF(V600) mutation occurs in about half of melanomas. This leads to increased oncogenic properties such as tumor cell invasion, metastatic potential, and resistance to apoptosis. Blockade of the MAPK pathway with highly specific kinase inhibitors induces unprecedented tumor response rates in patients with advanced BRAF(V600) mutant melanoma. Immune checkpoint blockade with monoclonal antibodies targeting cytotoxic T-lymphocyte antigen 4 and programed death-1/PD-L1 has also demonstrated striking anti-tumor activity in patients with advanced melanoma. Tumor responses are likely limited by multiple additional layers of immune suppression in the tumor microenvironment. There is emerging preclinical and clinical evidence suggesting that MAPK inhibition has a beneficial effect on the immunosuppressive tumor microenvironment, providing a strong rationale for combined immunotherapy and MAPK pathway inhibition in melanoma. The T cell response has been the main focus in the studies reported to date. Since dendritic cells (DCs) are important in the induction of tumor-specific T cell responses, the impact of MAPK pathway activation in melanoma on DC function is critical for the melanoma directed immune response. BRAF(V600E) melanoma cells modulate DCs through the MAPK pathway because its blockade in melanoma cells can reverse suppression of DC function. As both MEK/BRAF inhibition and immune checkpoint blockade have recently taken center stage in the treatment of melanoma, a deeper understanding of how MAPK pathway inhibition affects the tumor immune response is needed.

  4. A functional screen reveals an extensive layer of transcriptional and splicing control underlying RAS/MAPK signaling in Drosophila.

    Directory of Open Access Journals (Sweden)

    Dariel Ashton-Beaucage

    2014-03-01

    Full Text Available The small GTPase RAS is among the most prevalent oncogenes. The evolutionarily conserved RAF-MEK-MAPK module that lies downstream of RAS is one of the main conduits through which RAS transmits proliferative signals in normal and cancer cells. Genetic and biochemical studies conducted over the last two decades uncovered a small set of factors regulating RAS/MAPK signaling. Interestingly, most of these were found to control RAF activation, thus suggesting a central regulatory role for this event. Whether additional factors are required at this level or further downstream remains an open question. To obtain a comprehensive view of the elements functionally linked to the RAS/MAPK cascade, we used a quantitative assay in Drosophila S2 cells to conduct a genome-wide RNAi screen for factors impacting RAS-mediated MAPK activation. The screen led to the identification of 101 validated hits, including most of the previously known factors associated to this pathway. Epistasis experiments were then carried out on individual candidates to determine their position relative to core pathway components. While this revealed several new factors acting at different steps along the pathway--including a new protein complex modulating RAF activation--we found that most hits unexpectedly work downstream of MEK and specifically influence MAPK expression. These hits mainly consist of constitutive splicing factors and thereby suggest that splicing plays a specific role in establishing MAPK levels. We further characterized two representative members of this group and surprisingly found that they act by regulating mapk alternative splicing. This study provides an unprecedented assessment of the factors modulating RAS/MAPK signaling in Drosophila. In addition, it suggests that pathway output does not solely rely on classical signaling events, such as those controlling RAF activation, but also on the regulation of MAPK levels. Finally, it indicates that core splicing

  5. A Functional Screen Reveals an Extensive Layer of Transcriptional and Splicing Control Underlying RAS/MAPK Signaling in Drosophila

    Science.gov (United States)

    Ashton-Beaucage, Dariel; Udell, Christian M.; Gendron, Patrick; Sahmi, Malha; Lefrançois, Martin; Baril, Caroline; Guenier, Anne-Sophie; Duchaine, Jean; Lamarre, Daniel; Lemieux, Sébastien; Therrien, Marc

    2014-01-01

    The small GTPase RAS is among the most prevalent oncogenes. The evolutionarily conserved RAF-MEK-MAPK module that lies downstream of RAS is one of the main conduits through which RAS transmits proliferative signals in normal and cancer cells. Genetic and biochemical studies conducted over the last two decades uncovered a small set of factors regulating RAS/MAPK signaling. Interestingly, most of these were found to control RAF activation, thus suggesting a central regulatory role for this event. Whether additional factors are required at this level or further downstream remains an open question. To obtain a comprehensive view of the elements functionally linked to the RAS/MAPK cascade, we used a quantitative assay in Drosophila S2 cells to conduct a genome-wide RNAi screen for factors impacting RAS-mediated MAPK activation. The screen led to the identification of 101 validated hits, including most of the previously known factors associated to this pathway. Epistasis experiments were then carried out on individual candidates to determine their position relative to core pathway components. While this revealed several new factors acting at different steps along the pathway—including a new protein complex modulating RAF activation—we found that most hits unexpectedly work downstream of MEK and specifically influence MAPK expression. These hits mainly consist of constitutive splicing factors and thereby suggest that splicing plays a specific role in establishing MAPK levels. We further characterized two representative members of this group and surprisingly found that they act by regulating mapk alternative splicing. This study provides an unprecedented assessment of the factors modulating RAS/MAPK signaling in Drosophila. In addition, it suggests that pathway output does not solely rely on classical signaling events, such as those controlling RAF activation, but also on the regulation of MAPK levels. Finally, it indicates that core splicing components can also

  6. Activation of Erk and JNK MAPK pathways by acute swim stress in rat brain regions

    Directory of Open Access Journals (Sweden)

    Salvadore Christopher

    2004-09-01

    Full Text Available Abstract Background The mitogen-activated protein kinases (MAPKs have been shown to participate in a wide array of cellular functions. A role for some MAPKs (e.g., extracellular signal-regulated kinase, Erk1/2 has been documented in response to certain physiological stimuli, such as ischemia, visceral pain and electroconvulsive shock. We recently demonstrated that restraint stress activates the Erk MAPK pathway, but not c-Jun-N-terminal kinase/stress-activated protein kinase (JNK/SAPK or p38MAPK, in several rat brain regions. In the present study, we investigated the effects of a different stressor, acute forced swim stress, on the phosphorylation (P state of these MAPKs in the hippocampus, neocortex, prefrontal cortex, amygdala and striatum. In addition, effects on the phosphorylation state of the upstream activators of the MAPKs, their respective MAPK kinases (MAPKKs; P-MEK1/2, P-MKK4 and P-MKK3/6, were determined. Finally, because the Erk pathway can activate c-AMP response element (CRE binding (CREB protein, and swim stress has recently been reported to enhance CREB phosphorylation, changes in P-CREB were also examined. Results A single 15 min session of forced swimming increased P-Erk2 levels 2–3-fold in the neocortex, prefrontal cortex and striatum, but not in the hippocampus or amygdala. P-JNK levels (P-JNK1 and/or P-JNK2/3 were increased in all brain regions about 2–5-fold, whereas P-p38MAPK levels remained essentially unchanged. Surprisingly, levels of the phosphorylated MAPKKs, P-MEK1/2 and P-MKK4 (activators of the Erk and JNK pathways, respectively were increased in all five brain regions, and much more dramatically (P-MEK1/2, 4.5 to > 100-fold; P-MKK4, 12 to ~300-fold. Consistent with the lack of forced swim on phosphorylation of p38MAPK, there appeared to be no change in levels of its activator, P-MKK3/6. P-CREB was increased in all but cortical (prefrontal, neocortex areas. Conclusions Swim stress specifically and markedly

  7. Atmospheric mercury concentrations at two sites in the Kyushu Islands, Japan, and evidence of long-range transport from East Asia

    Science.gov (United States)

    Marumoto, Kohji; Hayashi, Masahiko; Takami, Akinori

    2015-09-01

    Continuous monitoring of atmospheric gaseous mercury at Fukuoka and Minamata in the Kyushu Islands, western Japan, was carried out from June 2012 to May 2013 to investigate the influence of long-range transport of mercury in the Asian region. Speciation data at Fukuoka indicated that approximately 99% of the atmospheric mercury was in the gaseous elemental form. The average concentration of gaseous elemental mercury (GEM) at Fukuoka was slightly higher than that of total gaseous mercury (TGM) at Minamata. Synchronous pollution events of higher concentrations of both GEM at Fukuoka and TGM at Minamata were frequently observed from late fall to early spring. We infer that these events occurred due to long-range transport of mercury rather than local, domestic emission sources because the two sites are far apart (about 150 km), and local sources would be unlikely to synchronously influence concentrations at both sites over such a long distance. The results of backward trajectory analyses indicated that these events occurred when air masses came from the Asian continent. In addition, the pollution events were often the result of cold fronts or migratory anticyclones that passed over the Kyushu Islands, often accompanied by descending cool and heavy air currents. Thus, these results indicate that, under specific climate conditions, higher concentrations of atmospheric mercury are transported to the Kyushu Islands from the Asian continent, and are evident in ground-based observations there.

  8. Making space for expressive and creative writing in African primary Schools: a two-site action research study in Kenya and South Africa

    Directory of Open Access Journals (Sweden)

    Paula Gains

    2011-05-01

    Full Text Available Similar concerns about the development of children’s creative writing abilities in Kenya and South Africa prompted two Mother Tongue (MT education practitioners in Summer Institute of Linguistics (SIL and Molteno Institute for Language and Literacy of Linguistics (MILL to undertake parallel intervention studies to increase teachers’ competence in writing pedagogy and improve the quantity and quality of learners’writing. Most early literacy teachers have had no experience themselves of expressive writing, so it is not surprising that this activity rarely, if ever, features in public school early literacy classrooms. The hypothesis which formed the basis for this action research study was that educators, exposed to extensive and expressive writing themselves, will be more skilled in the generation of such activities with learners. This paper reports on the workshop processes in the two sites, identifying similarities and di"erences in the experience. Whilst the hypothesis, though tested, remains unproven,this paper presents findings that are of relevance to further study in the area of writing pedagogy research and also to teachers and teacher educators involved in writing in the primary school.

  9. An ecological and comparative analysis of parasites in juvenile Mugil liza (Pisces, Mugilidae from two sites in Samborombón bay, Argentina

    Directory of Open Access Journals (Sweden)

    Martin M. Montes

    2015-12-01

    Full Text Available ABSTRACT Mugil liza Valenciennes, 1836 is an economically important food fish and has been recommended for aquaculture in South America. A total of 278 fishes were collected in the spring and summer of 2009 and 2010. These fish were sorted into sample groups according to their size class. We used Bayesian statistics and 95% credible intervals for each parameter tested were calculated. Fish studied harbored a total of 15 different species of parasites. Diversity of parasite species found on Mugil liza was greatest at the S.R.C. collection site, but evidenced a lower species richness than at A.R. site. The 1st size fishes of both sites evidenced greater parasite diversity than either 2nd or 3rd size fish. Differences observed could be explained by the different use of habitat types at the two sites or differential susceptibility to infection by parasites. The dominance of D. fastigatainfluenced observed results of lower community diversity indexes. New works elucidating different parasite life cycles within juvenile and adults ofM. liza in Argentina, promise to be important for determining the risk of the parasitism by zoonotic metacercariae A. (P. longa and use of this fish as food and an economic resource, and the possible use of mullet parasites in other promising fields as indicators of biodiversity, and/ or water contamination.

  10. Phage Anti-Immunocomplex Assay (PHAIA) for clomazone: Two-site recognition increases assay specificity and facilitates adaptation into a rapid on-site format

    Science.gov (United States)

    Rossotti, M.A.; Carlomagno, M.; González-Techera, A.; Hammock, B.D.; Last, J.; González-Sapienza, G.

    2010-01-01

    The impact of the use of herbicides in agriculture can be minimized by compliance with good management practices that reduce the amount used and their release into the environment. Simple tests that provide real time on-site information about these chemicals are a major aid for these programs. In this work we show that PHAIA, a method that uses phage-borne peptides to detect the formation of antibody-analyte immunocomplexes, is an advantageous technology to produce such field tests. A monoclonal antibody to the herbicide clomazone was raised and used in the development of conventional competitive and noncompetitive PHAIA immunoassays. The sensitivity attained with the PHAIA format was over ten times higher than that of the competitive format. The cross-reactivity of the two methods was also compared by using structurally related compounds, and we observed that the two-site binding of PHAIA “double-checks” the recognition of the analyte, thereby increasing the assay specificity. The positive readout of the noncompetitive PHAIA method allowed adaptation of the assay into a rapid and simple format where as little as 0.4 ng/ml of clomazone (more than 10-fold lower than the proposed standard) in water samples from a rice field could be easily detected by simple visual inspection. PMID:20886819

  11. Phage anti-immunocomplex assay for clomazone: two-site recognition increasing assay specificity and facilitating adaptation into an on-site format.

    Science.gov (United States)

    Rossotti, M A; Carlomagno, M; González-Techera, A; Hammock, B D; Last, J; González-Sapienza, G

    2010-11-01

    The impact of the use of herbicides in agriculture can be minimized by compliance with good management practices that reduce the amount used and their release into the environment. Simple tests that provide real time on-site information about these chemicals are a major aid for these programs. In this work, we show that phage anti-immunocomplex assay (PHAIA), a method that uses phage-borne peptides to detect the formation of antibody-analyte immunocomplexes, is an advantageous technology to produce such field tests. A monoclonal antibody to the herbicide clomazone was raised and used in the development of conventional competitive and noncompetitive PHAIA immunoassays. The sensitivity attained with the PHAIA format was over 10 times higher than that of the competitive format. The cross-reactivity of the two methods was also compared using structurally related compounds, and we observed that the two-site binding of PHAIA "double-checks" the recognition of the analyte, thereby increasing the assay specificity. The positive readout of the noncompetitive PHAIA method allowed adaptation of the assay into a rapid and simple format where as little as 0.4 ng/mL clomazone (more than 10-fold lower than the proposed standard) in water samples from a rice field could be easily detected by simple visual inspection.

  12. Parasites and hystopathology of Mullus barbatus and Citharus linguatula (Pisces from two sites in the NW Mediterranean with different degrees of pollution

    Directory of Open Access Journals (Sweden)

    Marta Carreras-Aubets

    2011-06-01

    Full Text Available The usefulness of fish parasite communities as bioindicators of environmental stress was tested on two benthic fish species, the red mullet (Mullus barbatus and the spotted flounder (Citharus linguatula, during the spring of 2006 at two sites of the Catalan coast (northwestern Mediterranean: an anthropogenic-impacted area located close to the city of Barcelona, and a less polluted area close to Blanes (Girona. Gonadosomatic and hepatosomatic indices and condition factor were determined for the fishes caught. Prevalence, mean intensity, mean abundance and species richness of the parasites found in the survey were calculated for both species and locations, and the main histological alterations were recorded. Cysts of unknown aetiology and intestinal coccidians were reported only in red mullets from the area close to Barcelona, which were highly parasitized by the digenean Opecoeloides furcatus and the nematode Capillaria sp. However, a higher prevalence of Ichthyophonus sp. was reported in the spotted flounder from Blanes. Cysts of unknown aetiology, some nematodes and Ichthyophonus sp. may be associated with pollution.

  13. Genome-Wide Survey and Expression Profile Analysis of the Mitogen-Activated Protein Kinase (MAPK Gene Family in Brassica rapa.

    Directory of Open Access Journals (Sweden)

    Kun Lu

    Full Text Available Mitogen-activated protein kinase (MAPK cascades are fundamental signal transduction modules in plants, controlling cell division, development, hormone signaling, and biotic and abiotic stress responses. Although MAPKs have been investigated in several plant species, a comprehensive analysis of the MAPK gene family has hitherto not been performed in Brassica rapa. In this study, we identified 32 MAPKs in the B. rapa genome by conducting BLASTP and syntenic block analyses, and screening for the essential signature motif (TDY or TEY of plant MAPK proteins. Of the 32 BraMAPK genes retrieved from the Brassica Database, 13 exhibited exon splicing errors, excessive splicing of the 5' sequence, excessive retention of the 5' sequence, and sequencing errors of the 3' end. Phylogenetic trees of the 32 corrected MAPKs from B. rapa and of MAPKs from other plants generated by the neighbor-joining and maximum likelihood methods suggested that BraMAPKs could be divided into four groups (groups A, B, C, and D. Gene number expansion was observed for BraMAPK genes in groups A and D, which may have been caused by the tandem duplication and genome triplication of the ancestral genome of the Brassica progenitor. Except for five members of the BraMAPK10 subfamily, the identified BraMAPKs were expressed in most of the tissues examined, including callus, root, stem, leaf, flower, and silique. Quantitative real-time PCR demonstrated that at least six and five BraMAPKs were induced or repressed by various abiotic stresses and hormone treatments, respectively, suggesting their potential roles in the abiotic stress response and various hormone signal transduction pathways in B. rapa. This study provides valuable insight into the putative physiological and biochemical functions of MAPK genes in B. rapa.

  14. Genome-Wide Survey and Expression Profile Analysis of the Mitogen-Activated Protein Kinase (MAPK) Gene Family in Brassica rapa.

    Science.gov (United States)

    Lu, Kun; Guo, Wenjin; Lu, Junxing; Yu, Hao; Qu, Cunmin; Tang, Zhanglin; Li, Jiana; Chai, Yourong; Liang, Ying

    2015-01-01

    Mitogen-activated protein kinase (MAPK) cascades are fundamental signal transduction modules in plants, controlling cell division, development, hormone signaling, and biotic and abiotic stress responses. Although MAPKs have been investigated in several plant species, a comprehensive analysis of the MAPK gene family has hitherto not been performed in Brassica rapa. In this study, we identified 32 MAPKs in the B. rapa genome by conducting BLASTP and syntenic block analyses, and screening for the essential signature motif (TDY or TEY) of plant MAPK proteins. Of the 32 BraMAPK genes retrieved from the Brassica Database, 13 exhibited exon splicing errors, excessive splicing of the 5' sequence, excessive retention of the 5' sequence, and sequencing errors of the 3' end. Phylogenetic trees of the 32 corrected MAPKs from B. rapa and of MAPKs from other plants generated by the neighbor-joining and maximum likelihood methods suggested that BraMAPKs could be divided into four groups (groups A, B, C, and D). Gene number expansion was observed for BraMAPK genes in groups A and D, which may have been caused by the tandem duplication and genome triplication of the ancestral genome of the Brassica progenitor. Except for five members of the BraMAPK10 subfamily, the identified BraMAPKs were expressed in most of the tissues examined, including callus, root, stem, leaf, flower, and silique. Quantitative real-time PCR demonstrated that at least six and five BraMAPKs were induced or repressed by various abiotic stresses and hormone treatments, respectively, suggesting their potential roles in the abiotic stress response and various hormone signal transduction pathways in B. rapa. This study provides valuable insight into the putative physiological and biochemical functions of MAPK genes in B. rapa.

  15. Tumor cell phenotype is sustained by selective MAPK oxidation in mitochondria.

    Directory of Open Access Journals (Sweden)

    Soledad Galli

    Full Text Available Mitochondria are major cellular sources of hydrogen peroxide (H(2O(2, the production of which is modulated by oxygen availability and the mitochondrial energy state. An increase of steady-state cell H(2O(2 concentration is able to control the transition from proliferating to quiescent phenotypes and to signal the end of proliferation; in tumor cells thereby, low H(2O(2 due to defective mitochondrial metabolism can contribute to sustain proliferation. Mitogen-activated protein kinases (MAPKs orchestrate signal transduction and recent data indicate that are present in mitochondria and regulated by the redox state. On these bases, we investigated the mechanistic connection of tumor mitochondrial dysfunction, H(2O(2 yield, and activation of MAPKs in LP07 murine tumor cells with confocal microscopy, in vivo imaging and directed mutagenesis. Two redox conditions were examined: low 1 microM H(2O(2 increased cell proliferation in ERK1/2-dependent manner whereas high 50 microM H(2O(2 arrested cell cycle by p38 and JNK1/2 activation. Regarding the experimental conditions as a three-compartment model (mitochondria, cytosol, and nuclei, the different responses depended on MAPKs preferential traffic to mitochondria, where a selective activation of either ERK1/2 or p38-JNK1/2 by co-localized upstream kinases (MAPKKs facilitated their further passage to nuclei. As assessed by mass spectra, MAPKs activation and efficient binding to cognate MAPKKs resulted from oxidation of conserved ERK1/2 or p38-JNK1/2 cysteine domains to sulfinic and sulfonic acids at a definite H(2O(2 level. Like this, high H(2O(2 or directed mutation of redox-sensitive ERK2 Cys(214 impeded binding to MEK1/2, caused ERK2 retention in mitochondria and restricted shuttle to nuclei. It is surmised that selective cysteine oxidations adjust the electrostatic forces that participate in a particular MAPK-MAPKK interaction. Considering that tumor mitochondria are dysfunctional, their inability to

  16. Specific MAPK-Associated MicroRNAs in Serum Differentiate Pancreatic Cancer from Autoimmune Pancreatitis.

    Directory of Open Access Journals (Sweden)

    Manabu Akamatsu

    Full Text Available Pancreatic ductal adenocarcinoma (PDAC is difficult to distinguish from autoimmune pancreatitis (AIP because of their clinical and radiological similarities, and therefore simple and minimally invasive surrogate markers for differential diagnosis would be useful. In our previous studies, we identified four microRNAs (miRNAs-miR-7, miR-34a, miR-181d, and miR-193b -as MAPK-associated microRNAs whose expression was altered significantly with upregulation of the MAPK signaling pathway. Recently it has been reported that these miRNAs could be used as biomarkers in serum samples for accurate diagnosis of pancreatic lesions. The aim of the present study was to evaluate whether these MAPK-associated miRNAs in serum could be used as biomarkers for differentiating PDAC from AIP. We enrolled 69 patients with PDAC, 26 with intraductal papillary mucinous neoplasm (IPMN and 15 with AIP. The expression of MAPK-associated miRNAs in serum was measured by quantitative real-time PCR. The 2-ΔCT method was used to quantify the expression of miRNAs, and the data were normalized using spiked-in synthetic cel-miR-39. Patients with PDAC or IPMN showed significantly higher amounts of serum MAPK-associated miRNAs than those with AIP (p<0.009 for miR-7, p<0.002 for miR-34a, p<0.001 for miR-181d, p<0.002 for miR-193b. ROC curve analysis demonstrated that these miRNAs had an area under the ROC curve (AUC of 0.723-0.882 for differentiation between PDAC or IPMN from AIP. Furthermore, serum miR-181d was significantly associated with the presence of metastasis in patients with PDA (p = 0.014. Serum MAPK-associated miRNAs could be novel noninvasive biomarkers for differentiation between PDAC or IPMN and AIP.

  17. Convergence of Multiple MAP3Ks on MKK3 Identifies a Set of Novel Stress MAPK Modules

    KAUST Repository

    Colcombet, Jean

    2016-12-22

    Since its first description in 1995 and functional characterization 12 years later, plant MKK3-type MAP2Ks have emerged as important integrators in plant signaling. Although they have received less attention than the canonical stress-activated mitogen-activated protein kinases (MAPKs), several recent publications shed light on their important roles in plant adaptation to environmental conditions. Nevertheless, the MKK3-related literature is complicated. This review summarizes the current knowledge and discrepancies on MKK3 MAPK modules in plants and highlights the singular role of MKK3 in green plants. In the light of the latest data, we hypothesize a general model that all clade-III MAP3Ks converge on MKK3 and C-group MAPKs, thereby defining a set of novel MAPK modules which are activated by stresses and internal signals through the transcriptional regulation of MAP3K genes.

  18. Ganoderma lucidum polysaccharide exerts anti-tumor activity via MAPK pathways in HL-60 acute leukemia cells.

    Science.gov (United States)

    Yang, Guohua; Yang, Lei; Zhuang, Yun; Qian, Xifeng; Shen, Yunfeng

    2016-01-01

    In this study, we investigated the anti-tumor activity both in vitro and in vivo of a polysaccharide obtained from Ganoderma lucidum on HL-60 acute myeloid leukemia cells, and focused on its targeting effect on mitogen-activated protein kinase (MAPK) pathways. It was found by the methods such as western blot and flow cytometry (FCM), that G. lucidum polysaccharide (GLP) blocked the extracellular signal-regulated kinase/MAPK signaling pathway, simultaneously activated p38 and JNK MAPK pathways, and therefore regulated their downstream genes and proteins, including p53, c-myc, c-fos, c-jun, Bcl-2, Bax, cleaved caspase-3 and cyclin D1. As a result, cycle arrest and apoptosis of HL-60 cells were induced. Therefore, GLP exerted anti-tumor activity via MAPK pathways in HL-60 acute leukemia cells.

  19. Development of an efficient signal amplification strategy for label-free enzyme immunoassay using two site-specific biotinylated recombinant proteins

    Energy Technology Data Exchange (ETDEWEB)

    Tang, Jin-Bao [School of Pharmacy, Weifang Medical University, Weifang 261053 (China); Tang, Ying [Affiliated Hospital of Weifang Medical University, Weifang 261041 (China); Yang, Hong-Ming, E-mail: yanghongming2006@sohu.com [School of Pharmacy, Weifang Medical University, Weifang 261053 (China)

    2015-02-15

    Highlights: • An efficient signal amplification strategy for label-free EIA is proposed. • Divalent biotinylated AP and monovalent biotinylated ZZ were prepared via Avitag–BirA system. • The above site-specific biotinylated fusion proteins form complex via SA–biotin interaction. • The mechanism relies on the ZZ–Avi-B/SA/AP–(Avi-B){sub 2} complex. • The analytical signals are enhanced (32-fold) by the proposed strategy. - Abstract: Constructing a recombinant protein between a reporter enzyme and a detector protein to produce a homogeneous immunological reagent is advantageous over random chemical conjugation. However, the approach hardly recombines multiple enzymes in a difunctional fusion protein, which results in insufficient amplification of the enzymatic signal, thereby limiting its application in further enhancement of analytical signal. In this study, two site-specific biotinylated recombinant proteins, namely, divalent biotinylated alkaline phosphatase (AP) and monovalent biotinylated ZZ domain, were produced by employing the Avitag–BirA system. Through the high streptavidin (SA)–biotin interaction, the divalent biotinylated APs were clustered in the SA–biotin complex and then incorporated with the biotinylated ZZ. This incorporation results in the formation of a functional macromolecule that involves numerous APs, thereby enhancing the enzymatic signal, and in the production of several ZZ molecules for the interaction with immunoglobulin G (IgG) antibody. The advantage of this signal amplification strategy is demonstrated through ELISA, in which the analytical signal was substantially enhanced, with a 32-fold increase in the detection sensitivity compared with the ZZ–AP fusion protein approach. The proposed immunoassay without chemical modification can be an alternative strategy to enhance the analytical signals in various applications involving immunosensors and diagnostic chips, given that the label-free IgG antibody is suitable

  20. PM chemical composition and oxidative potential of the soluble fraction of particles at two sites in the urban area of Milan, Northern Italy

    Science.gov (United States)

    Perrone, Maria Grazia; Zhou, Jun; Malandrino, Mery; Sangiorgi, Giorgia; Rizzi, Cristiana; Ferrero, Luca; Dommen, Josef; Bolzacchini, Ezio

    2016-03-01

    Recent epidemiological evidence support the hypothesis that health effects from inhalation of air particles are governed by more than just particle mass, since specific chemical components have been identified as important contributors to mortality and hospitality admissions. We studied the chemical composition and the oxidative potential (OP) of total suspended particle (TSP) samples from Milan at two sites with different traffic loads: a site in the low emission zone (LEZ) and a traffic site (TR) outside. Two a-cellular assays; dithiothreitol (OPDTT) and 2‧,7' dichlorofluorescin (OPDCFH) were used to characterize the OP of the soluble fraction of particles. TSP samples from LEZ showed significantly lower concentrations of traffic-related chemical components compared to TR. The decrease in the concentrations from TR to LEZ was maximum for EC, with a LEZ/TR ratio of 0.64 (±0.18), and a significant reduction (p cellular assays gave complementary information on the OP of particles in Milan. The two OP assays resulted to be sensitive to different chemical properties of PM samples. OPDTT correlated positively only with Global Radiation (Spearman's rs = 0.38, p < 0.05), which could be considered as a proxy for high concentrations of secondary oxidizing organics, while OPDCFH was related to various PM chemical species, mainly correlated with total mass (rs = 0.65; p < 0.01), elements (e.g. Zn, rs = 0.67; As, rs = 0.65; p < 0.01) and the sum of sulfate and nitrate (rs = 0.63; p < 0.01), a proxy for secondary aerosol.

  1. Two Drosophila model neurons can regenerate axons from the stump or from a converted dendrite, with feedback between the two sites.

    Science.gov (United States)

    Rao, Kavitha S; Rolls, Melissa M

    2017-08-17

    After axon severing, neurons recover function by reinitiating axon outgrowth. New outgrowth often originates from the remaining axon stump. However, in many mammalian neurons, new axons initiate from a dendritic site when the axon is injured close to the cell body. Drosophila sensory neurons are ideal for studying neuronal injury responses because they can be injured reproducibly in a variety of genetic backgrounds. In Drosophila, it has been shown that a complex sensory neuron, ddaC, can regenerate an axon from a stump, and a simple sensory neuron, ddaE, can regenerate an axon from a dendrite. To provide a more complete picture of axon regeneration in these cell types, we performed additional injury types. We found that ddaE neurons can initiate regeneration from an axon stump when a stump remains. We also showed that ddaC neurons regenerate from the dendrite when the axon is severed close to the cell body. We next demonstrated if a stump remains, new axons can originate from this site and a dendrite at the same time. Because cutting the axon close to the cell body results in growth of the new axon from a dendrite, and cutting further out may not, we asked whether the initial response in the cell body was similar after both types of injury. A transcriptional reporter for axon injury signaling, puc-GFP, increased with similar timing and levels after proximal and distal axotomy. However, changes in dendritic microtubule polarity differed in response to the two types of injury, and were influenced by the presence of a scar at the distal axotomy site. We conclude that both ddaE and ddaC can regenerate axons either from the stump or a dendrite, and that there is some feedback between the two sites that modulates dendritic microtubule polarity.

  2. Amarogentin, a Secoiridoid Glycoside, Abrogates Platelet Activation through PLCγ2-PKC and MAPK Pathways

    Directory of Open Access Journals (Sweden)

    Ting-Lin Yen

    2014-01-01

    Full Text Available Amarogentin, an active principle of Gentiana lutea, possess antitumorigenic, antidiabetic, and antioxidative properties. Activation of platelets is associated with intravascular thrombosis and cardiovascular diseases. The present study examined the effects of amarogentin on platelet activation. Amarogentin treatment (15~60 μM inhibited platelet aggregation induced by collagen, but not thrombin, arachidonic acid, and U46619. Amarogentin inhibited collagen-induced phosphorylation of phospholipase C (PLCγ2, protein kinase C (PKC, and mitogen-activated protein kinases (MAPKs. It also inhibits in vivo thrombus formation in mice. In addition, neither the guanylate cyclase inhibitor ODQ nor the adenylate cyclase inhibitor SQ22536 affected the amarogentin-mediated inhibition of platelet aggregation, which suggests that amarogentin does not regulate the levels of cyclic AMP and cyclic GMP. In conclusion, amarogentin prevents platelet activation through the inhibition of PLCγ2-PKC cascade and MAPK pathway. Our findings suggest that amarogentin may offer therapeutic potential for preventing or treating thromboembolic disorders.

  3. Resistin increases platelet P-selectin levels via p38 MAPK signal pathway.

    Science.gov (United States)

    Qiu, Wenbing; Chen, Naping; Zhang, Qin; Zhuo, Liyuan; Wang, Xihong; Wang, Dongming; Jin, Hong

    2014-03-01

    Resistin, an adipokine associated with the metabolic syndrome, is believed to have a role in thrombotic conditions. This work analyses the effects of resistin on P-selectin expression using a combination of ex vivo human studies, in vivo animal models and in vitro cell cultures. Human platelets and vascular endothelial cells were incubated with resistin, with or without anti-Toll-like receptor 4 (TLR-4) or mitogen-activated protein kinases (MAPK) pathway inhibitors, whereas mice were treated with resistin infusion followed by analysis of P-selectin expression. Resistin increased both human and murine platelet P-selectin expression compared with controls (human: 48.02% ± 7.6% vs 35.12% ± 2.62%, p P-selectin production. We conclude that resistin induces platelet activation by increasing P-selectin expression through the p38 MAPK-dependent pathway. These data provide one mechanism for the prothrombotic state in individuals with the metabolic syndrome.

  4. The Mitogen-Activated Protein Kinase (MAPK) Pathway: Role in Immune Evasion by Trypanosomatids

    Science.gov (United States)

    Soares-Silva, Mercedes; Diniz, Flavia F.; Gomes, Gabriela N.; Bahia, Diana

    2016-01-01

    Leishmania spp. and Trypanosoma cruzi are the causative agents of leishmaniasis and Chagas disease, respectively, two neglected tropical diseases that affect about 25 million people worldwide. These parasites belong to the family Trypanosomatidae, and are both obligate intracellular parasites that manipulate host signaling pathways and the innate immune system to establish infection. Mitogen-activated protein kinases (MAPKs) are serine and threonine protein kinases that are highly conserved in eukaryotes, and are involved in signal transduction pathways that modulate physiological and pathophysiological cell responses. This mini-review highlights existing knowledge concerning the mechanisms that Leishmania spp. and T. cruzi have evolved to target the host’s MAPK signaling pathways and highjack the immune response, and, in this manner, promote parasite maintenance in the host. PMID:26941717

  5. Caspases and p38 MAPK regulate endothelial cell adhesiveness for mesenchymal stem cells.

    Directory of Open Access Journals (Sweden)

    Irina A Potapova

    Full Text Available Mesenchymal stem cells natively circulating or delivered into the blood stream home to sites of injury. The mechanism of mesenchymal stem cell homing to sites of injury is poorly understood. We have shown that the development of apoptosis in endothelial cells stimulates endothelial cell adhesiveness for mesenchymal stem cells. Adhesion of mesenchymal stem cells to apoptotic endothelial cells depends on the activation of endothelial caspases and p38 MAPK. Activation of p38 MAPK in endothelial cells has a primary effect while the activation of caspases potentiates the mesenchymal stem cell adhesion. Overall, our study of the mesenchymal stem cell interaction with endothelial cells indicates that mesenchymal stem cells recognize and specifically adhere to distressed/apoptotic endothelial cells.

  6. IL-13–induced airway mucus production is attenuated by MAPK13 inhibition

    Science.gov (United States)

    Alevy, Yael G.; Patel, Anand C.; Romero, Arthur G.; Patel, Dhara A.; Tucker, Jennifer; Roswit, William T.; Miller, Chantel A.; Heier, Richard F.; Byers, Derek E.; Brett, Tom J.; Holtzman, Michael J.

    2012-01-01

    Increased mucus production is a common cause of morbidity and mortality in inflammatory airway diseases, including asthma, chronic obstructive pulmonary disease (COPD), and cystic fibrosis. However, the precise molecular mechanisms for pathogenic mucus production are largely undetermined. Accordingly, there are no specific and effective anti-mucus therapeutics. Here, we define a signaling pathway from chloride channel calcium-activated 1 (CLCA1) to MAPK13 that is responsible for IL-13–driven mucus production in human airway epithelial cells. The same pathway was also highly activated in the lungs of humans with excess mucus production due to COPD. We further validated the pathway by using structure-based drug design to develop a series of novel MAPK13 inhibitors with nanomolar potency that effectively reduced mucus production in human airway epithelial cells. These results uncover and validate a new pathway for regulating mucus production as well as a corresponding therapeutic approach to mucus overproduction in inflammatory airway diseases. PMID:23187130

  7. MAPK-Mediated YAP Activation Controls Mechanical-Tension-Induced Pulmonary Alveolar Regeneration

    Directory of Open Access Journals (Sweden)

    Zhe Liu

    2016-08-01

    Full Text Available The pulmonary alveolar epithelium undergoes extensive regeneration in response to lung injuries, including lung resection. In recent years, our understanding of cell lineage relationships in the pulmonary alveolar epithelium has improved significantly. However, the molecular and cellular mechanisms that regulate pneumonectomy (PNX-induced alveolar regeneration remain largely unknown. In this study, we demonstrate that mechanical-tension-induced YAP activation in alveolar stem cells plays a major role in promoting post-PNX alveolar regeneration. Our results indicate that JNK and p38 MAPK signaling is critical for mediating actin-cytoskeleton-remodeling-induced nuclear YAP expression in alveolar stem cells. Moreover, we show that Cdc42-controlled actin remodeling is required for the activation of JNK, p38, and YAP in post-PNX lungs. Our findings together establish that the Cdc42/F-actin/MAPK/YAP signaling cascade is essential for promoting alveolar regeneration in response to mechanical tension in the lung.

  8. Functional analysis of Arabidopsis immune-related MAPKs uncovers a role for MPK3 as negative regulator of inducible defences

    KAUST Repository

    Frei dit Frey, Nicolas

    2014-06-30

    Background Mitogen-activated protein kinases (MAPKs) are key regulators of immune responses in animals and plants. In Arabidopsis, perception of microbe-associated molecular patterns (MAMPs) activates the MAPKs MPK3, MPK4 and MPK6. Increasing information depicts the molecular events activated by MAMPs in plants, but the specific and cooperative contributions of the MAPKs in these signalling events are largely unclear. Results In this work, we analyse the behaviour of MPK3, MPK4 and MPK6 mutants in early and late immune responses triggered by the MAMP flg22 from bacterial flagellin. A genome-wide transcriptome analysis reveals that 36% of the flg22-upregulated genes and 68% of the flg22-downregulated genes are affected in at least one MAPK mutant. So far MPK4 was considered as a negative regulator of immunity, whereas MPK3 and MPK6 were believed to play partially redundant positive functions in defence. Our work reveals that MPK4 is required for the regulation of approximately 50% of flg22-induced genes and we identify a negative role for MPK3 in regulating defence gene expression, flg22-induced salicylic acid accumulation and disease resistance to Pseudomonas syringae. Among the MAPK-dependent genes, 27% of flg22-upregulated genes and 76% of flg22-downregulated genes require two or three MAPKs for their regulation. The flg22-induced MAPK activities are differentially regulated in MPK3 and MPK6 mutants, both in amplitude and duration, revealing a highly interdependent network. Conclusions These data reveal a new set of distinct functions for MPK3, MPK4 and MPK6 and indicate that the plant immune signalling network is choreographed through the interplay of these three interwoven MAPK pathways.

  9. Increases in cAMP, MAPK Activity and CREB Phosphorylation during REM Sleep: Implications for REM Sleep and Memory Consolidation

    OpenAIRE

    Luo, Jie; Phan, Trongha X.; Yang, Yimei; Garelick, Michael G.; Storm, Daniel R.

    2013-01-01

    The cyclic adenosine monophosphate (cAMP), mitogen-activated protein kinase (MAPK) and cAMP response element-binding protein (CREB) transcriptional pathway is required for consolidation of hippocampus-dependent memory. In mice, this pathway undergoes a circadian oscillation required for memory persistence that reaches a peak during the daytime. Since mice exhibit polyphasic sleep patterns during the day, this suggested the interesting possibility that cAMP, MAPK activity and CREB phosphorylat...

  10. 弓形虫MAPK1基因的克隆与原核表达∗%Cloning and Prokaryotic Expression of MAPK1 Gene of Toxoplasma gondii

    Institute of Scientific and Technical Information of China (English)

    宫上舒; 赵权; 刘全

    2015-01-01

    To clone the MAPK1 gene and construct the prokaryotic expression vector PET⁃28a⁃MAPK1, to transform the expression vector into E. coli BL21, to induce and identify recombinant protein, and to lay experimental foundation for further analysis of biological characteristics and im⁃mune protection of MAPK1 gene,the study was conducted. A pair of primers were designed accord⁃ing to the sequence of MAPK1 from GenBank. The MAPK1 gene was amplified by PCR. Using NCoⅠ and HindⅢ double enzyme, the primers were connected to the prokaryotic expression vector of PET⁃28a with the same double enzyme cut. Recombinant expression plasmid PET⁃28a⁃MAPK1 was constructed and transformed to the fabrication E.coli BL21 competent expressed by IPTG induc⁃tion. Expression product was purified with Ni2+ chelating affinity column, and the purified recombi⁃nant plasmid was analyzed by SDS-PAGE and Western blotting. Results showed that the cloned MAPK1 gene had 99�9% homology with gene sequence included in GenBank, The recombinantplasmids was identified by restriction enzyme and the desired size of 1 599 bp fragment was ob⁃tained, which was consistent with the expected results, indicating that recombinant plasmid PET⁃28a⁃MAPK1 was successfully constructed. Protein concentration of Ni2+ chelate column after purifi⁃cation was 0�26 mg/mL, indicating that the Ni2+ chelate column could be used as an immunogen. SDS-PAGE electrophoresis showed that molecular weight of fusion protein of PET⁃28a⁃MAPK1 was about 58 ku. Western⁃blotting showed that the fusion protein had good antigenicity. The expression plasmid was expressed in E.coli BL21, and the study lays foundation for further exploration of toxo⁃plasmosis diagnosis.%为获取具有生物学活性的弓形虫MAPK1蛋白,根据GenBank中编码MAPK1的已知序列设计并合成1对引物,用NCoⅠ和HindⅢ双酶切后,连接到同样双酶切的原核表达载体PET⁃28a

  11. Correlation between Expression of P38 MAPK-Signaling and uPA in Breast Cancer

    Institute of Scientific and Technical Information of China (English)

    Yanchun Han; Luying Liu; Dongxia Yan; Guihua Wang

    2008-01-01

    OBJECTIVE To study the expression of phosphorylated p38 mitogen.Activated protein kinase(p-p38)and uPA and the correlation of their expression with breast cancer Clinic.patholodiCal characteristics,and to investigate the role of the p38MAPK-signaling pathway in regulating uPA expression in breast cancer cells.METHODS Immunohistochemistry(S-P)was used to test the expression of P-p38 and uPA in 60 specimens of breast cancer tissues.Western blots were adopted to detect expression of the p-p38 and uPA proteins in MDA-MB-231 and MCF-7 breast cancer cells.And uPA expression after treatment with SB203580,a specific inhibitor of p38 MAPK.RESULTS The positive rate of the P.P38 protein and uPA protein expression in the breast cancer tissues was 56.7% and 60.0%.Respectively.The expression of P.P38 was positively related to the expression of uPA(r=0.316,P0.05).The expression of p-p38 and uPA in MDA. MB-231 cells was higher than that in MCF.7 cells.SB203580 inhibited the p38 MAPK pathway and reduced uPA protein expression.CONCLUSI0N The p38 MAPK-signaling pathway promotes breast cancer malignant progression by up.Regulating uPA expression,and it may be an important process in breast cancer invasion and metastasis.

  12. Increased p38-MAPK is responsible for chemotherapy resistance in human gastric cancer cells

    Directory of Open Access Journals (Sweden)

    Jiang Guocheng

    2008-12-01

    Full Text Available Abstract Background Chemoresistance is one of the main obstacles to successful cancer therapy and is frequently associated with Multidrug resistance (MDR. Many different mechanisms have been suggested to explain the development of an MDR phenotype in cancer cells. One of the most studied mechanisms is the overexpression of P-glycoprotein (P-gp, which is a product of the MDR1 gene. Tumor cells often acquire the drug-resistance phenotype due to upregulation of the MDR1 gene. Overexpression of MDR1 gene has often been reported in primary gastric adenocarcinoma. Methods This study investigated the role of p38-MAPK signal pathway in vincristine-resistant SGC7901/VCR cells. P-gp and MDR1 RNA were detected by Western blot analysis and RT-PCR amplification. Mitgen-activated protein kinases and function of P-gp were demonstrated by Western blot and FACS Aria cytometer analysis. Ap-1 activity and cell apoptosis were detected by Dual-Luciferase Reporter Assay and annexin V-PI dual staining. Results The vincristine-resistant SGC7901/VCR cells with increased expression of the multidrug-resistance 1 (MDR1 gene were resistant to P-gp-related drug and P-gp-unrelated drugs. Constitutive increases of phosphorylated p38-MAPK and AP-1 activities were also found in the drug-resistant cells. Inhibition of p38-MAPK by SB202190 reduced activator protein-1 (AP-1 activity and MDR1 expression levels and increased the sensitivity of SGC7901/VCR cells to chemotherapy. Conclusion Activation of the p38-MAPK pathway might be responsible for the modulation of P-glycoprotein-mediated and P-glycoprotein-unmediated multidrug resistance in the SGC7901/VCR cell line.

  13. Silica nanoparticles induce endoplasmic reticulum stress response and activate mitogen activated kinase (MAPK signalling

    Directory of Open Access Journals (Sweden)

    Verena Christen

    2016-01-01

    Full Text Available Humans may be exposed to engineered silica nanoparticles (SiO2-NPs but potential adverse effects are poorly understood, in particular in relation to cellular effects and modes of action. Here we studied effects of SiO2-NPs on cellular function in human hepatoma cells (Huh7. Exposure for 24 h to 10 and 50 μg/ml SiO2-NPs led to induction of endoplasmic reticulum (ER stress as demonstrated by transcriptional induction of DNAJB9, GADD34, CHOP, as well as CHOP target genes BIM, CHAC-1, NOXA and PUMA. In addition, CHOP protein was induced. In addition, SiO2-NPs induced an inflammatory response as demonstrated by induction of TNF-α and IL-8. Activation of MAPK signalling was investigated employing a PCR array upon exposure of Huh7 cells to SiO2-NPs. Five of 84 analysed genes, including P21, P19, CFOS, CJUN and KSR1 exhibited significant transcriptional up-regulation, and 18 genes a significant down-regulation. Strongest down-regulation occurred for the proto-oncogene BRAF, MAPK11, one of the four p38 MAPK genes, and for NFATC4. Strong induction of CFOS, CJUN, FRA1 and CMYC was found after exposure to 50 μg/ml SiO2-NPs for 24 h. To analyse for effects derived from up-regulation of TNF-α, Huh7 cells were exposed to SiO2-NPs in the presence of the TNF-α inhibitor sauchinone, which reduced the induction of the TNF-α transcript by about 50%. These data demonstrate that SiO2-NPs induce ER stress, MAPK pathway and lead to inflammatory reaction in human hepatoma cells. Health implications of SiO2-NPs exposure should further be investigated for a risk assessment of these frequently used nanoparticles.

  14. Musashi2 modulates K562 leukemic cell proliferation and apoptosis involving the MAPK pathway

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Huijuan; Tan, Shi; Wang, Juan; Chen, Shana; Quan, Jing; Xian, Jingrong; Zhang, Shuai shuai; He, Jingang; Zhang, Ling, E-mail: lingzhang@cqmu.edu.cn

    2014-01-01

    The RNA-binding protein Musashi2 (Msi2) has been identified as a master regulator within a variety of stem cell populations via the regulation of translational gene expression. A recent study has suggested that Msi2 is strongly expressed in leukemic cells of acute myeloid leukemia patients, and elevated Msi2 is associated with poor prognosis. However, the potential role of Msi2 in leukemogenesis is still not well understood. Here, we investigated the effect of Msi2 knockdown on the biological properties of leukemic cells. High expression of Msi2 was found in K562 and KG-1a leukemic cell lines, and low expression was observed in the U937 cell line. We transduced K562 cells with two independent adenoviral shRNA vectors targeting Msi2 and confirmed knockdown of Msi2 at the mRNA and protein levels. Msi2 silencing inhibited cell growth and caused cell cycle arrest by increasing the expression of p21 and decreasing the expression of cyclin D1 and cdk2. In addition, knockdown of Msi2 promoted cellular apoptosis via the upregulation of Bax and downregulation of Bcl-2 expression. Furthermore, Msi2 knockdown resulted in the inactivation of the ERK/MAPK and p38/MAPK pathways, but no remarkable change in p-AKT was observed. These data provide evidence that Msi2 plays an important role in leukemogenesis involving the MAPK signaling pathway, which indicates that Msi2 may be a novel target for leukemia treatment. - Highlights: • Knockdown of Msi2 inhibited K562 cell growth and arrested cell cycle progression. • Knockdown of Msi2 induced K562 cell apoptosis via the regulation of Bax and Bcl-2. • The MAPK pathway was involved in the process of Msi2-mediated leukemogenesis. • Our data indicate that Msi2 is a potential new target for leukemia treatment.

  15. FGFR3 promotes synchondrosis closure and fusion of ossification centers through the MAPK pathway

    OpenAIRE

    Matsushita, Takehiko; Wilcox, William R.; Chan, Yuk Yu; Kawanami, Aya; Bükülmez, Hülya; Balmes, Gener; Krejci, Pavel; Mekikian, Pertchoui B.; Otani, Kazuyuki; Yamaura, Isakichi; Warman, Matthew L.; Givol, David; Murakami, Shunichi

    2008-01-01

    Activating mutations in FGFR3 cause achondroplasia and thanatophoric dysplasia, the most common human skeletal dysplasias. In these disorders, spinal canal and foramen magnum stenosis can cause serious neurologic complications. Here, we provide evidence that FGFR3 and MAPK signaling in chondrocytes promote synchondrosis closure and fusion of ossification centers. We observed premature synchondrosis closure in the spine and cranial base in human cases of homozygous achondroplasia and thanatoph...

  16. MAPK signaling pathways regulate mitochondrial-mediated apoptosis induced by isoorientin in human hepatoblastoma cancer cells.

    Science.gov (United States)

    Yuan, Li; Wang, Jing; Xiao, Haifang; Wu, Wanqiang; Wang, Yutang; Liu, Xuebo

    2013-03-01

    Isoorientin (ISO) (CAS RN: 4261-42-1) is a flavonoid compound that can be extracted from several plant species, such as Phyllostachys pubescens, Patrinia, and Drosophyllum lusitanicum. ISO is able to induce apoptosis through mitochondrial dysfunction and inhibition of PI3K/Akt signaling pathway in HepG2 cells, however, the effects of ISO on MAPK signaling pathways remain unknown. The present study investigated the effects of ISO on this pathway, and the roles of MAPK kinases on mitochondrial-mediated apoptosis in HepG2 cells. The results showed that ISO induced cell death in a dose- and time-dependent manner, and induction apoptosis is main cause for ISO-induced cytotoxicity in HepG2 cells. ISO significantly inhibited the levels of ERK1/2 kinase and increased the expression of JNK and p38 kinases. Furthermore, U0126 (an ERK1/2 inhibitor) significantly enhanced the ISO-induced the Bax/Bcl-2 ratio, the release of cytochrome c to the cytosol fraction, and the levels of cleaved caspase-3. While SP600125 (a JNK inhibitor) and SB203580 (a p38 inhibitor) markedly prevented the expression of these proteins induced by ISO. Furthermore, the ROS inhibitor (NAC) notably promoted the inhibited effect of ISO on the ERK1/2 kinase. NAC also suppressed the p-JNK and p-p38, but failed to reverse the effects of ISO. These results demonstrated for the first time that ISO induces apoptosis in HepG2 cells through inactivating ERK1/2 kinase and activating JNK and p38 kinases, and ROS stimulated by ISO is able to activate the MAPK singaling pathway as the upstream signaling molecules. Initiating event of the mitochondrial-mediated apoptosis induced by ISO is MAPK signals.

  17. Cadmium induces vascular permeability via activation of the p38 MAPK pathway

    Energy Technology Data Exchange (ETDEWEB)

    Dong, Fengyun [Laboratory of Microvascular Medicine, Medical Research Center, Shandong Provincial Qianfoshan Hospital, Shandong University, 16766 Jingshi Road, Jinan, Shandong 250014 (China); Guo, Fang [Department of Cardiology, Provincial Hospital Affiliated to Shandong University, 324 Jingwu Road, Jinan, Shandong 250021 (China); Li, Liqun [Laboratory of Microvascular Medicine, Medical Research Center, Shandong Provincial Qianfoshan Hospital, Shandong University, 16766 Jingshi Road, Jinan, Shandong 250014 (China); Guo, Ling; Hou, Yinglong; Hao, Enkui; Yan, Suhua [Department of Cardiology, Shandong Provincial Qianfoshan Hospital, Shandong University, 16766 Jingshi Road, Jinan, Shandong 250014 (China); Allen, Thaddeus D. [G.W. Hooper Research Foundation, University of California at San Francisco, 513 Parnassus Ave., HSW1501, San Francisco, CA 94143-0552 (United States); Liu, Ju, E-mail: ju.liu@sdu.edu.cn [Laboratory of Microvascular Medicine, Medical Research Center, Shandong Provincial Qianfoshan Hospital, Shandong University, 16766 Jingshi Road, Jinan, Shandong 250014 (China)

    2014-07-18

    Highlights: • Low-dose cadmium (Cd) induces vascular hyper-permeability. • p38 MAPK mediates Cd-induced disruption of endothelial cell barrier function. • SB203850 inhibits Cd-induced membrane dissociation of VE-cadherin and β-catenin. • SB203850 reduces Cd-induced expression and secretion of TNF-α. - Abstract: The vasculature of various organs is a targeted by the environmental toxin, cadmium (Cd). However, mechanisms leading to pathological conditions are poorly understood. In the present study, we examined the effect of cadmium chloride (CdCl{sub 2}) on human umbilical vein endothelial cells (HUVECs). At 4 μM, CdCl{sub 2} induced a hyper-permeability defect in HUVECs, but not the inhibition of cell growth up to 24 h. This effect of CdCl{sub 2} was dependent on the activation of the p38 mitogen-activated protein kinase (MAPK) pathway. The p38 MAPK inhibitor SB203850 suppressed the CdCl{sub 2}-induced alteration in trans-endothelial electrical resistance in HUVEC monolayers, a model measurement of vascular endothelial barrier integrity. SB203850 also inhibited the Cd-induced membrane dissociation of vascular endothelial (VE) cadherin and β-catenin, the important components of the adherens junctional complex. In addition, SB203850 reduces the Cd-induced expression and secretion of tumor necrosis factor α (TNF-α). Taken together, our findings suggest that Cd induces vascular hyper-permeability and disruption of endothelial barrier integrity through stimulation of p38 MAPK signaling.

  18. Norcantharidin induces apoptosis in HeLa cells through caspase, MAPK, and mitochondrial pathways

    Institute of Scientific and Technical Information of China (English)

    Wei-weiAN; Xian-fengGONG; Min-weiWANG; Shin-ichiTASHIRO; SatoshiONODERA; TakashiIKEJIMA

    2004-01-01

    AIM: To investigate the mechanism of norcantharidin (NCTD)-induced HeLa cell apoptosis. METHODS: HeLa cell growth inhibition was measured by MTT method. Apoptosis was detected by Hoechst 33258 staining and agarose gel electrophoresis. Caspase activities were assayed using caspase apoptosis detection kit. Western blot analysis was used to evaluate the level of ICAD, ERK/p-ERK, JNK/p-JNK, and Bcl-X.L/Bax expression. RESULTS: Norcantharidin inhibited HeLa cell growth in a time- and dose-dependent manner. HeLa cells treated with norcantharidin showed typical characteristics of apoptosis including the morphological changes and DNA fragmentation. Caspase family inhibitor (z-VAD-fmk), caspase-8, -9 inhibitor (z-IETD-fmk, Ac-LEHD-CHO, respectively) and caspase-3 inhibitor (z-DEVD-fmk) partially prevent norcantharidin-induced apoptosis, but initiator caspase-1 inhibitor (Ac-YVAD-fmk) did not. The activities of caspase-3, -8, and -9 were up-regulated after norcantharidin treatment. Furthermore, NCTD-induced activation of caspase-3 resulted in the degradation of the inhibitor of caspase-activated DNase (ICAD). Up-regulation of mitochondrial Bax expression and down-regulation of Bcl-xLexpression also participated in the apoptosis induced by NCTD. Although p38 MAPK inhibitor (SB203580) failed to block cell death, ERK MAPK inhibitor (PD98059) and JNK MAPK inhibitor (SP600125) had marked inhibitory effects on norcantharidin-induced apoptosis. Moreover, the phosphorylation of JNK were up-regulated followed by delayed ERK phosphorylation after treatment with NCTD, suggesting that ERK and JNK were both responsible for NCTD-induced apoptosis in HeLa cells and worked at different stages. CONCLUSION: The cytotoxic effect of NCTD on HeLa cells was mainly due to apoptosis. The anti-tumor mechanism of NCTD might involve caspses, mitochondrial, and MAPKs pathways.

  19. Norcantharidin induces apoptosis in HeLa cells through caspase,MAPK,and mitochondrial pathways

    Institute of Scientific and Technical Information of China (English)

    Wei-wei AN; Xian-feng GONG; Min-wei WANG; Shin-ichi TASHIRO; Satoshi ONODERA; Takashi IKEJIMA

    2004-01-01

    AIM: To investigate the mechanism of norcantharidin (NCTD)-induced HeLa cell apoptosis. METHODS: HeLa cell growth inhibition was measured by MTT method. Apoptosis was detected by Hoechst 33258 staining and agarose gel electrophoresis. Caspase activities were assayed using caspase apoptosis detection kit. Western blot analysis was used to evaluate the level of ICAD, ERK/p-ERK, JNK/p-JNK, and Bcl-XL/Bax expression. RESULTS:Norcantharidin inhibited HeLa cell growth in a time- and dose-dependent manner. HeLa cells treated with norcantharidin showed typical characteristics of apoptosis including the morphological changes and DNA fragmentation. Caspase family inhibitor (z-VAD-fmk), caspase-8, -9 inhibitor (z-IETD-fmk, Ac-LEHD-CHO,respectively) and caspase-3 inhibitor (z-DEVD-fmk) partially prevent norcantharidin-induced apoptosis, but initiator caspase-1 inhibitor (Ac-YVAD-fmk) did not. The activities of caspase-3, -8, and -9 were up-regulated after norcantharidin treatment. Furthermore, NCTD-induced activation of caspase-3 resulted in the degradation of the inhibitor of caspase-activated DNase (ICAD). Up-regulation of mitochondrial Bax expression and down-regulation of Bcl-xL expression also participated in the apoptosis induced by NCTD. Although p38 MAPK inhibitor (SB203580)failed to block cell death, ERK MAPK inhibitor (PD98059) and JNK MAPK inhibitor (SP600125) had marked inhibitory effects on norcantharidin-induced apoptosis. Moreover, the phosphorylation of JNK were up-regulated followed by delayed ERK phosphorylation after treatment with NCTD, suggesting that ERK and JNK were both responsible for NCTD-induced apoptosis in HeLa cells and worked at different stages. CONCLUSION: The cytotoxic effect of NCTD on HeLa cells was mainly due to apoptosis. The anti-tumor mechanism of NCTD might involve caspses, mitochondrial, and MAPKs pathways.

  20. Loss of MLK3 signaling impedes ulcer healing by modulating MAPK signaling in mouse intestinal mucosa

    OpenAIRE

    Kovalenko, Pavlo L.; KUNOVSKA, LYUDMYLA; Chen, Jian; Kathleen A Gallo; Basson, Marc D.

    2012-01-01

    Mixed-lineage kinase 3 (MLK3) activates multiple MAPK pathways and can initiate apoptosis, proliferation, migration, or differentiation in different cell types. However, whether MLK3 signaling regulates intestinal epithelial cell sheet migration in vivo is not known. We sought to investigate whether MLK3 signaling is important in intestinal mucosal healing and epithelial cell motility in vivo and in vitro. In vivo, we compared the healing of jejunal mucosal ulcers induced in MLK3 knockout (KO...

  1. Integrative modelling of the influence of MAPK network on cancer cell fate decision.

    Directory of Open Access Journals (Sweden)

    Luca Grieco

    2013-10-01

    Full Text Available The Mitogen-Activated Protein Kinase (MAPK network consists of tightly interconnected signalling pathways involved in diverse cellular processes, such as cell cycle, survival, apoptosis and differentiation. Although several studies reported the involvement of these signalling cascades in cancer deregulations, the precise mechanisms underlying their influence on the balance between cell proliferation and cell death (cell fate decision in pathological circumstances remain elusive. Based on an extensive analysis of published data, we have built a comprehensive and generic reaction map for the MAPK signalling network, using CellDesigner software. In order to explore the MAPK responses to different stimuli and better understand their contributions to cell fate decision, we have considered the most crucial components and interactions and encoded them into a logical model, using the software GINsim. Our logical model analysis particularly focuses on urinary bladder cancer, where MAPK network deregulations have often been associated with specific phenotypes. To cope with the combinatorial explosion of the number of states, we have applied novel algorithms for model reduction and for the compression of state transition graphs, both implemented into the software GINsim. The results of systematic simulations for different signal combinations and network perturbations were found globally coherent with published data. In silico experiments further enabled us to delineate the roles of specific components, cross-talks and regulatory feedbacks in cell fate decision. Finally, tentative proliferative or anti-proliferative mechanisms can be connected with established bladder cancer deregulations, namely Epidermal Growth Factor Receptor (EGFR over-expression and Fibroblast Growth Factor Receptor 3 (FGFR3 activating mutations.

  2. Formaldehyde induces apoptosis through decreased Prx 2 via p38 MAPK in lung epithelial cells.

    Science.gov (United States)

    Lim, Seul Ki; Kim, Jong Chun; Moon, Chang Jong; Kim, Gye Yeop; Han, Ho Jae; Park, Soo Hyun

    2010-05-27

    Formaldehyde (FA) is an important substance that induces sick house syndrome and diseases, such as asthma and allergies. Oxidative stress is involved in the development of respiratory disease, and diverse antioxidants may protect respiratory tract cells from apoptosis. Peroxiredoxin is a pivotal endogenous antioxidant. In the present study, FA induced death in A549 cells, a lung epithelial cell line, in a dose-dependent manner. FA also increased lipid peroxide formation (LPO) in A549 cells, suggesting a role for oxidative stress. Additionally, FA decreased peroxiredoxin 2 (Prx 2) protein levels after a 24 or 48h exposure to FA. We also examined whether the FA-induced decrease in Prx 2 was associated with apoptosis. Prx 2 overexpression protected against FA-induced cell apoptosis but not necrosis. Prx 2 overexpression blocked FA-induced increase in Bax, a pro-apoptotic molecule, and a decrease in Bcl-2, an anti-apoptotic molecule. Prx 2 overexpression also protected against FA-induced activation of some special apoptosis-associated proteins [caspase-3, caspase-9, and polypeptide poly (ADP-ribose) polymerase (PARP)]. Furthermore, we examined the signaling molecules involved in the FA-induced decrease in Prx 2 expression. The FA-induced decrease in Prx 2 and increase in cell apoptosis was restored by treatment with SB203580 [a p38 mitogen activated protein kinase (MAPK) inhibitor], but not by SP600125 [a c-jun-N-terminal kinase (JNK) inhibitor]. Also, FA-induced events were blocked by treatment with p38 siRNA, but not by scrambled siRNA. Indeed, FA increased p38 MAPK activation, suggesting a role for p38 MAPK in FA action. In conclusion, FA mediated apoptosis in lung epithelial cells by decreasing Prx 2 via p38 MAPK.

  3. Nitric oxide enhances increase in cytosolic Ca(2+) and promotes nicotine-triggered MAPK pathway in PC12 cells.

    Science.gov (United States)

    Kajiwara, Aya; Tsuchiya, Yukihiro; Takata, Tsuyoshi; Nyunoya, Mayumi; Nozaki, Naohito; Ihara, Hideshi; Watanabe, Yasuo

    2013-11-01

    The purpose of this study was to investigate the roles of neuronal nitric oxide synthase (nNOS), Ca(2+)/calmodulin (CaM)-dependent protein kinases (CaMKs), and protein kinase C (PKC) in nicotine-induced extracellular signal-regulated kinases 1 and 2 (ERK1/2) and p38 mitogen-activated protein kinase (MAPK) activation. Treatment with nicotine stimulated ERK1/2 and p38 MAPK phosphorylation in the PC12 cells expressing nNOS (NPC12 cells) as compared with that in control PC12 cells. An inhibitor of L-type voltage-sensitive Ca(2+) channel suppressed the nicotine-induced phosphorylation of p38 MAPK. The inhibition of CaMK-kinase, the upstream activator of CaMKI and CaMKIV, did not inhibit the enhanced their phosphorylation. ERK1/2 phosphorylation was attenuated by inhibitors of p38 MAPK, PKC, and MAPK-kinase 1/2, indicating the involvement of these protein kinases upstream of ERK1/2. Furthermore, we found that nNOS expression enhances the nicotine-induced increase in the intracellular concentration of Ca(2+), using the Ca(2+)-sensitive fluorescent probe Fura2. These data suggest that NO promotes nicotine-triggered Ca(2+) transient in PC12 cells to activate possibly CaMKII, leading to sequential phosphorylation of p38 MAPK and ERK1/2.

  4. Oncogenic ETS proteins mimic activated RAS/MAPK signaling in prostate cells

    Science.gov (United States)

    Hollenhorst, Peter C.; Ferris, Mary W.; Hull, Megan A.; Chae, Heejoon; Kim, Sun; Graves, Barbara J.

    2011-01-01

    The aberrant expression of an oncogenic ETS transcription factor is implicated in the progression of the majority of prostate cancers, 40% of melanomas, and most cases of gastrointestinal stromal tumor and Ewing's sarcoma. Chromosomal rearrangements in prostate cancer result in overexpression of any one of four ETS transcription factors. How these four oncogenic ETS genes differ from the numerous other ETS genes expressed in normal prostate and contribute to tumor progression is not understood. We report that these oncogenic ETS proteins, but not other ETS factors, enhance prostate cell migration. Genome-wide binding analysis matched this specific biological function to occupancy of a unique set of genomic sites highlighted by the presence of ETS- and AP-1-binding sequences. ETS/AP-1-binding sequences are prototypical RAS-responsive elements, but oncogenic ETS proteins activated a RAS/MAPK transcriptional program in the absence of MAPK activation. Thus, overexpression of oncogenic ETS proteins can replace RAS/MAPK pathway activation in prostate cells. The genomic description of this ETS/AP-1-regulated, RAS-responsive, gene expression program provides a resource for understanding the role of these ETS factors in both an oncogenic setting and the developmental processes where these genes normally function. PMID:22012618

  5. RUNX1 Regulates Migration, Invasion, and Angiogenesis via p38 MAPK Pathway in Human Glioblastoma.

    Science.gov (United States)

    Sangpairoj, Kant; Vivithanaporn, Pornpun; Apisawetakan, Somjai; Chongthammakun, Sukumal; Sobhon, Prasert; Chaithirayanon, Kulathida

    2016-12-24

    Runt-related transcription factor 1 (RUNX1) is essential for the establishment of fetal and adult hematopoiesis and neuronal development. Aberrant expression of RUNX1 led to proliferation and metastasis of several cancers. The aim of the present study was to investigate the role of RUNX1 in migration, invasion, and angiogenesis of human glioblastoma using IL-1β-treated U-87 MG human glioblastoma cells as a model. IL-1β at 10 ng/ml stimulated translocation of RUNX1 into the nucleus with increased expressions of RUNX1, MMP-1, MMP-2, MMP-9, MMP-19, and VEGFA in U-87 MG cells. In addition, silencing of RUNX1 gene significantly suppressed U-87 MG cell migration and invasion abilities. Moreover, knockdown of RUNX1 mRNA in U-87 MG cells reduced the tube formation of human umbilical vein endothelial cells. Further investigation revealed that IL-1β-induced RUNX1 expression might be mediated via the p38 mitogen-activated protein kinase (MAPK) signaling molecule for the expression of these invasion- and angiogenic-related molecules. Together with an inhibitor of p38 MAPK (SB203580) could decrease RUNX1 mRNA expression. Thus, RUNX1 may be one of the putative molecular targeted therapies against glioma metastasis and angiogenesis through the activation of p38 MAPK signaling pathway.

  6. Modulation of inflammation and pathology during dengue virus infection by p38 MAPK inhibitor SB203580.

    Science.gov (United States)

    Fu, Yilong; Yip, Andy; Seah, Peck Gee; Blasco, Francesca; Shi, Pei-Yong; Hervé, Maxime

    2014-10-01

    Dengue virus (DENV) infection could lead to dengue fever (DF), dengue hemorrhagic fever (DHF) or dengue shock syndrome (DSS). The disease outcome is controlled by both viral and host factors. Inflammation mediators from DENV-infected cells could contribute to increased vascular permeability, leading to severe DHF/DSS. Therefore, suppression of inflammation could be a potential therapeutic approach for treatment of dengue patients. In this context, p38 MAPK (mitogen-activated protein kinase) is a key enzyme that modulates the initiation of stress and inflammatory responses. Here we show that SB203580, a p38 MAPK inhibitor, suppressed the over production of DENV-induced pro-inflammatory mediators such as TNF-α, IL-8, and RANTES from human PBMCs, monocytic THP-1, and granulocyte KU812 cell lines. Oral administration of SB203580 in DENV-infected AG129 mice prevented hematocrit rise and lymphopenia, limited the development of inflammation and pathology (including intestine leakage), and significantly improved survival. These results, for the first time, have provided experimental evidence to imply that a short term inhibition of p38 MAPK may be beneficial to reduce disease symptoms in dengue patients.

  7. Conservation of protein abundance patterns reveals the regulatory architecture of the EGFR-MAPK pathway

    Energy Technology Data Exchange (ETDEWEB)

    Shi, T.; Niepel, M.; McDermott, J. E.; Gao, Y.; Nicora, C. D.; Chrisler, W. B.; Markillie, L. M.; Petyuk, V. A.; Smith, R. D.; Rodland, K. D.; Sorger, P. K.; Qian, W. -J.; Wiley, H. S.

    2016-07-12

    It is not known whether cancer cells generally show quantitative differences in the expression of signaling pathway proteins that could dysregulate signal transduction. To explore this issue, we first defined the primary components of the EGF-MAPK pathway in normal human mammary epithelial cells, identifying 16 core proteins and 10 feedback regulators. We then quantified their absolute abundance across a panel of normal and cancer cell lines. We found that core pathway proteins were expressed at very similar levels across all cell types. In contrast, the EGFR and transcriptionally controlled feedback regulators were expressed at highly variable levels. The absolute abundance of most core pathway proteins was between 50,000- 70,000 copies per cell, but the adaptors SOS1, SOS2, and GAB1 were found at far lower levels (2,000-5,000 per cell). MAPK signaling showed saturation in all cells between 3,000-10,000 occupied EGFR, consistent with the idea that low adaptor levels limit signaling. Our results suggest that the core MAPK pathway is essentially invariant across different cell types, with cell- specific differences in signaling likely due to variable levels of feedback regulators. The low abundance of adaptors relative to the EGFR could be responsible for previous observation of saturable signaling, endocytosis, and high affinity EGFR.

  8. Copper oxide nanoparticles aggravate airway inflammation and mucus production in asthmatic mice via MAPK signaling.

    Science.gov (United States)

    Park, Ji-Won; Lee, In-Chul; Shin, Na-Rae; Jeon, Chan-Mi; Kwon, Ok-Kyoung; Ko, Je-Won; Kim, Jong-Choon; Oh, Sei-Ryang; Shin, In-Sik; Ahn, Kyung-Seop

    2016-01-01

    Copper oxide nanoparticles (CuONPs), metal oxide nanoparticles were used in multiple applications including wood preservation, antimicrobial textiles, catalysts for carbon monoxide oxidation and heat transfer fluid in machines. We investigated the effects of CuONPs on the respiratory system in Balb/c mice. In addition, to investigate the effects of CuONPs on asthma development, we used a murine model of ovalbumin (OVA)-induced asthma. CuONPs markedly increased airway hyper-responsiveness (AHR), inflammatory cell counts, proinflammatory cytokines and reactive oxygen species (ROS). CuONPs induced airway inflammation and mucus secretion with increases in phosphorylation of the MAPKs (Erk, JNK and p38). In the OVA-induced asthma model, CuONPs aggravated the increased AHR, inflammatory cell count, proinflammatory cytokines, ROS and immunoglobulin E induced by OVA exposure. In addition, CuONPs markedly increased inflammatory cell infiltration into the lung and mucus secretions, and MAPK phosphorylation was elevated compared to OVA-induced asthmatic mice. Taken together, CuONPs exhibited toxicity on the respiratory system, which was associated with the MAPK phosphorylation. In addition, CuONPs exposure aggravated the development of asthma. We conclude that CuONPs exposure has a potential toxicity in humans with respiratory disease.

  9. Atractylodin Inhibits Interleukin-6 by Blocking NPM-ALK Activation and MAPKs in HMC-1.

    Science.gov (United States)

    Chae, Hee-Sung; Kim, Young-Mi; Chin, Young-Won

    2016-09-02

    Atractylodin is one of the major constituents of the rhizome of Atractylodes lancea, which is widely used in Korean traditional medicine as a remedy for the treatment of gastritis and gastric ulcers. Despite of a major constituent of widely used botanical to treat inflammatory responses little is known about anti-inflammatory effect of atractylodin in the human mast cell (HMC-1). Hence, we evaluated the effect of atractylodin on the release of IL-6, the involvement of nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) and mitogen-activated protein kinases (MAPKs) in phorbol-12-myristate-13-acetate and A23187-induced HMC-1. In addition, Janus kinase 2 (JAK2), signal transducer and activator of transcription 3 (STAT3), phospholipase C (PLC) gamma 1, and AKT phosphorylation relevant to NPM-ALK signal pathway were assessed. IL-6 levels in the HMC-1 stimulated by phorbol-12-myristate-13-acetate and A23187 were apparently decreased by the treatment of atractylodin. Concurrently, atractylodin not only inhibited the phosphorylation of NPM-ALK, but also suppressed the phosphorylation of JAK2, STAT3, PLC gamma 1, and AKT. Furthermore, the activated mitogen-activated protein kinases (MAPKs) by phorbol-12-myristate-13-acetate and A23187 were inhibited by atractylodin. These results suggested that atractylodin might have a potential regulatory effect on inflammatory mediator expression through blockade of both the phosphorylation of MAPKs and the NPM-ALK signaling pathway.

  10. Atractylodin Inhibits Interleukin-6 by Blocking NPM-ALK Activation and MAPKs in HMC-1

    Directory of Open Access Journals (Sweden)

    Hee-Sung Chae

    2016-09-01

    Full Text Available Atractylodin is one of the major constituents of the rhizome of Atractylodes lancea, which is widely used in Korean traditional medicine as a remedy for the treatment of gastritis and gastric ulcers. Despite of a major constituent of widely used botanical to treat inflammatory responses little is known about anti-inflammatory effect of atractylodin in the human mast cell (HMC-1. Hence, we evaluated the effect of atractylodin on the release of IL-6, the involvement of nucleophosmin-anaplastic lymphoma kinase (NPM-ALK and mitogen-activated protein kinases (MAPKs in phorbol-12-myristate-13-acetate and A23187-induced HMC-1. In addition, Janus kinase 2 (JAK2, signal transducer and activator of transcription 3 (STAT3, phospholipase C (PLC gamma 1, and AKT phosphorylation relevant to NPM-ALK signal pathway were assessed. IL-6 levels in the HMC-1 stimulated by phorbol-12-myristate-13-acetate and A23187 were apparently decreased by the treatment of atractylodin. Concurrently, atractylodin not only inhibited the phosphorylation of NPM-ALK, but also suppressed the phosphorylation of JAK2, STAT3, PLC gamma 1, and AKT. Furthermore, the activated mitogen-activated protein kinases (MAPKs by phorbol-12-myristate-13-acetate and A23187 were inhibited by atractylodin. These results suggested that atractylodin might have a potential regulatory effect on inflammatory mediator expression through blockade of both the phosphorylation of MAPKs and the NPM-ALK signaling pathway.

  11. Oxidant stress and skeletal muscle glucose transport: roles of insulin signaling and p38 MAPK.

    Science.gov (United States)

    Kim, John S; Saengsirisuwan, Vitoon; Sloniger, Julie A; Teachey, Mary K; Henriksen, Erik J

    2006-09-01

    Oxidative stress can impact the regulation of glucose transport activity in a variety of cell lines. In the present study, we assessed the direct effects of an oxidant stress on the glucose transport system in intact mammalian skeletal muscle preparations. Type IIb (epitrochlearis) and type I (soleus) muscles from insulin-sensitive lean Zucker rats were incubated in 8 mM glucose for 2 h in the absence or presence of 100 mU/ml glucose oxidase to produce the oxidant hydrogen peroxide (H(2)O(2)) (60-90 microM). Glucose transport, glycogen synthase activity, and metabolic signaling factors were then assessed. H(2)O(2) significantly (p oxidant stress was prevented by the PI3-kinase inhibitor wortmannin. The oxidant stress also significantly increased phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK) and 5'-AMP-dependent protein kinase. Interestingly, selective inhibition of p38 MAPK using A304000 substantially reduced the activation of glucose transport induced by the oxidant stress. These results support a direct role for oxidative stress in the activation of the glucose transport system in mammalian skeletal muscle and indicate that this process involves engagement of and possible interactions between the PI3-kinase-dependent signaling pathway and activation of p38 MAPK.

  12. FGFR1-induced epithelial to mesenchymal transition through MAPK/PLCγ/COX-2-mediated mechanisms.

    Directory of Open Access Journals (Sweden)

    Darren C Tomlinson

    Full Text Available Tumour invasion and metastasis is the most common cause of death from cancer. For epithelial cells to invade surrounding tissues and metastasise, an epithelial-mesenchymal transition (EMT is required. We have demonstrated that FGFR1 expression is increased in bladder cancer and that activation of FGFR1 induces an EMT in urothelial carcinoma (UC cell lines. Here, we created an in vitro FGFR1-inducible model of EMT, and used this model to identify regulators of urothelial EMT. FGFR1 activation promoted EMT over a period of 72 hours. Initially a rapid increase in actin stress fibres occurred, followed by an increase in cell size, altered morphology and increased migration and invasion. By using site-directed mutagenesis and small molecule inhibitors we demonstrated that combined activation of the mitogen activated protein kinase (MAPK and phospholipase C gamma (PLCγ pathways regulated this EMT. Actin stress fibre formation was regulated by PLCγ activation, and was also important for the increase in cell size, migration and altered morphology. MAPK activation regulated migration and E-cadherin expression, indicating that combined activation of PLCγ and MAPK is required for a full EMT. We used expression microarrays to assess changes in gene expression downstream of these signalling cascades. COX-2 was transcriptionally upregulated by FGFR1 and caused increased intracellular prostaglandin E(2 levels, which promoted migration. In conclusion, we have demonstrated that FGFR1 activation in UC cells lines promotes EMT via coordinated activation of multiple signalling pathways and by promoting activation of prostaglandin synthesis.

  13. MLK4β functions as a negative regulator of MAPK signaling and cell invasion.

    Science.gov (United States)

    Abi Saab, W F; Brown, M S; Chadee, D N

    2012-03-26

    Mixed lineage kinase (MLK) 4, or MLK4, is a member of the MLK family of mitogen-activated protein kinase kinase kinases (MAP3Ks). Typically, MAP3Ks function to activate the mitogen-activated protein kinase (MAPK)-signaling pathways and regulate different cellular responses. However, here we report that MLK4β, unlike the other MLKs, negatively regulates the activities of the MAPKs, p38, c-Jun N-terminal kinase and extracellular signal-regulated kinase, and the MAP2Ks, MEK3 and 6. Our results show that MLK4β inhibits sorbitol- and tumor necrosis factor-induced activation of p38. Furthermore, MLK4β interacts with another MLK family member, MLK3, in HCT116 cells. Exogenous expression of MLK4β inhibits activation of MLK3 and also blocks matrix metalloproteinase-9 gelatinase activity and invasion in SKOV3 ovarian cancer cells. Collectively, our data establish MLK4β as a novel suppressor of MLK3 activation, MAPK signaling and cell invasion.

  14. Rice GDP dissociation inhibitor 3 inhibits OsMAPK2 activity through physical interaction.

    Science.gov (United States)

    Heo, Jae Bok; Yi, Young Byung; Bahk, Jeong Dong

    2011-11-04

    GDP dissociation inhibitor (GDI) plays an essential role in regulating the state of bound nucleotides and subcellular localizations of Rab proteins. In our previous study, we showed that OsGDI3 facilitates the recycling of OsRab11 with a help of OsGAP1. In this study, we show that OsGDI3 complement the yeast sec19-1 mutant, a temperature-sensitive allele of the yeast GDI gene, suggesting that OsGDI3 is a functional ortholog of yeast GDI. To obtain further knowledge on the function of OsGDI3, candidate OsGDI3-interacting proteins were identified by yeast two-hybrid screens. OsMAPK2 is one of OsGDI3 interacting proteins from yeast two-hybrid screens and subject to further analysis. A kinase assay showed that the autophosphorylation activity of OsMAPK2 is inhibited by OsGDI3 in vitro. In addition, ectopic expressions of OsGDI3-in Arabidopsis cause reductions at the level of phosphorylated AtMPK in phosphorylation activity. Taken together, OsGDI3 functions as a negative regulator of OsMAPK2 through modulating its kinase activity.

  15. Slt2 MAPK pathway is essential for cell integrity in the presence of arsenate.

    Science.gov (United States)

    Matia-González, Ana M; Rodríguez-Gabriel, Miguel A

    2011-01-01

    Arsenate is a common toxic metalloid found in drinking water worldwide that causes several human diseases. The biochemical action underlying cellular response to arsenate, however, is not yet completely understood. Here we used Saccharomyces cerevisiae as an eukaryotic model system to identify proteins essential for adaptation to arsenate treatment. Previous studies have demonstrated a function for Hog1 MAPK in modulating the cellular response to arsenite. Our results, however, showed that cells deficient in Hog1 did not show increased sensitivity to arsenate, suggesting that perhaps other MAPKs may be involved in the response to this particular arsenic species. Here, we found that Slt2 MAPK and several of its upstream regulators are essential in modulating the response to arsenate, and that Slt2 is phosphorylated after arsenate treatment. Furthermore, whole-genome transcriptional analysis showed that Slt2 is required for the induction of several genes in response to arsenate exposure. Many of these genes are involved in the cellular response to heat, suggesting an overlap between these two stress response pathways, and pointing toward a common response to both arsenate and heat exposure in Saccharomyces cerevisiae. Furthermore, our results support the idea that cellular exposure to arsenate results in induction of cellular signalling pathways different from those induced under arsenite treatment.

  16. A new mathematical approach for qualitative modeling of the insulin-TOR-MAPK network

    Directory of Open Access Journals (Sweden)

    H. Frederik Nijhout

    2013-09-01

    Full Text Available In this paper we develop a novel mathematical model of the insulin-TOR-MAPK signaling network that controls growth. Most data on the properties of the insulin and MAPK signaling networks are static and the responses to experimental interventions, such as knockouts, overexpression, and hormonal input are typically reported as scaled quantities. The modeling paradigm we develop here uses scaled variables and is ideally suited to simulate systems in which much of the available data are scaled. Our mathematical representation of signaling networks provides a way to reconcile theory and experiments, thus leading to a better understanding of the properties and function of these signaling networks. We test the performance of the model against a broad diversity of experimental data. The model correctly reproduces experimental insulin dose-response relationships. We study the interaction between insulin and MAPK signaling in the control of protein synthesis, and the interactions between amino acids, insulin and TOR signaling. We study the effects of variation in FOXO expression on protein synthesis and glucose transport capacity, and show that a FOXO knockout can partially rescue protein synthesis capacity of an insulin receptor knockout. We conclude that the modeling paradigm we develop provides a simple tool to investigate the qualitative properties of signaling networks.

  17. Arctigenin induces apoptosis in colon cancer cells through ROS/p38MAPK pathway.

    Science.gov (United States)

    Li, Qing-chun; Liang, Yun; Tian, Yuan; Hu, Guang-rui

    2016-01-01

    In the current study the antiproliferative effect of arctigenin, plant lignin, was evaluated on human colon cancer cell line HT-29. Furthermore, attempts were made to explore the signaling mechanism which may be responsible for its effect. Cell growth inhibition was assessed by MTT and LDH assays. Flow cytometric analysis was performed to determine cell arrest in the cell cycle phase and apoptosis. Furthermore, to confirm the apoptotic activity of arctigenin, caspase-9 and -3 activities analysis was performed. The levels of reactive oxygen species (ROS) and p38 mitogen activated protein kinase (MAPK) were investigated to determine their role in inducing apoptosis in arctigenin-treated HT-29 colon cancer cell line. MTT and LDH results demonstrated significant cell growth inhibitory effect of arctigenin on HT-29 cells in a dose-dependent manner. Furthermore, increase in cell number arrested at G2/M phase was observed in flow cytometric analysis upon arctigenin treatment. In addition, arctigenin increased the apoptotic ratio in a dose-dependent manner. The involvement of intrinsic apoptotic pathway was indicated by the activation of caspase-9 and -3. Moreover, increased ROS production, activation of p38 MAPK and changes in mitochondrial membrane potential (ΔΨm) also revealed the role of intrinsic apoptotic signaling pathway in cell growth inhibition after arctigenin exposure. Arctigenin induces apoptosis in HT-29 colon cancer cells by regulating ROS and p38 MAPK pathways.

  18. Involvement of MAPKs in ICAM-1 Expression in Glomerular Endothelial Cells in Diabetic Nephropathy

    Directory of Open Access Journals (Sweden)

    Watanabe,Naomi

    2011-08-01

    Full Text Available Inflammatory processes are involved in the pathogenesis of diabetic nephropathy. The aim of this study was to clarify the role of mitogen-activated protein kinase (MAPK pathways for induction of intercellular adhesion molecule-1 (ICAM-1 expression in glomerular endothelial cells under diabetic conditions. We examined the expression of ICAM-1 in the kidneys of experimental diabetic rats. Human glomerular endothelial cells (GE cells were exposed to normal glucose concentration, high glucose concentration (HG, or high mannitol concentration (HM, and then the expression of the ICAM-1 protein and the phosphorylation of the 3 subfamilies of mitogen-activated protein kinase (MAPK were determined using Western blot analysis. Next, to evaluate the involvement of MAPKs in HG- or HM-induced ICAM-1 expression, we preincubated GE cells with the inhibitors for ERK, p38 or JNK 1h prior to the application of glucose or mannitol. Expression of ICAM-1 was increased in the glomeruli of diabetic rats. Both HG and HM induced ICAM-1 expression and phosphorylation of ERK1/2, p38 and JNK in GE cells. Expression of ICAM-1 was significantly attenuated by inhibitors of ERK, p38 and JNK. We conclude that activation of ERK1/2, p38 and JNK cascades may be involved in ICAM-1 expression in glomerular endothelial cells under diabetic conditions.

  19. Use of p38 MAPK Inhibitors for the Treatment of Werner Syndrome

    Directory of Open Access Journals (Sweden)

    Mark C. Bagley

    2010-06-01

    Full Text Available Werner syndrome provides a convincing model for aspects of the normal ageing phenotype and may provide a suitable model for therapeutic interventions designed to combat the ageing process. Cultured primary fibroblast cells from Werner syndrome patients provide a powerful model system to study the link between replicative senescence in vitro and in vivo pathophysiology. Genome instability, together with an increased pro-oxidant state, and frequent replication fork stalling, all provide plausible triggers for intracellular stress in Werner syndrome cells, and implicates p38 MAPK signaling in their shortened replicative lifespan. A number of different p38 MAPK inhibitor chemotypes have been prepared rapidly and efficiently using microwave heating techniques for biological study in Werner syndrome cells, including SB203580, VX-745, RO3201195, UR-13756 and BIRB 796, and their selectivity and potency evaluated in this cellular context. Werner syndrome fibroblasts treated with a p38 MAPK inhibitor reveal an unexpected reversal of the accelerated ageing phenotype. Thus the study of p38 inhibition and its effect upon Werner pathophysiology is likely to provide new revelations into the biological mechanisms operating in cellular senescence and human ageing in the future.

  20. BMP9-Induced Survival Effect in Liver Tumor Cells Requires p38MAPK Activation

    Directory of Open Access Journals (Sweden)

    María García-Álvaro

    2015-08-01

    Full Text Available The study of bone morphogenetic proteins (BMPs role in tumorigenic processes, and specifically in the liver, has gathered importance in the last few years. Previous studies have shown that BMP9 is overexpressed in about 40% of hepatocellular carcinoma (HCC patients. In vitro data have also shown evidence that BMP9 has a pro-tumorigenic action, not only by inducing epithelial to mesenchymal transition (EMT and migration, but also by promoting proliferation and survival in liver cancer cells. However, the precise mechanisms driving these effects have not yet been established. In the present work, we deepened our studies into the intracellular mechanisms implicated in the BMP9 proliferative and pro-survival effect on liver tumor cells. In HepG2 cells, BMP9 induces both Smad and non-Smad signaling cascades, specifically PI3K/AKT and p38MAPK. However, only the p38MAPK pathway contributes to the BMP9 growth-promoting effect on these cells. Using genetic and pharmacological approaches, we demonstrate that p38MAPK activation, although dispensable for the BMP9 proliferative activity, is required for the BMP9 protective effect on serum withdrawal-induced apoptosis. These findings contribute to a better understanding of the signaling pathways involved in the BMP9 pro-tumorigenic role in liver tumor cells.

  1. Ras-induced and extracellular signal-regulated kinase 1 and 2 phosphorylation-dependent isomerization of protein tyrosine phosphatase (PTP)-PEST by PIN1 promotes FAK dephosphorylation by PTP-PEST.

    Science.gov (United States)

    Zheng, Yanhua; Yang, Weiwei; Xia, Yan; Hawke, David; Liu, David X; Lu, Zhimin

    2011-11-01

    Protein tyrosine phosphatase (PTP)-PEST is a critical regulator of cell adhesion and migration. However, the mechanism by which PTP-PEST is regulated in response to oncogenic signaling to dephosphorylate its substrates remains unclear. Here, we demonstrate that activated Ras induces extracellular signal-regulated kinase 1 and 2-dependent phosphorylation of PTP-PEST at S571, which recruits PIN1 to bind to PTP-PEST. Isomerization of the phosphorylated PTP-PEST by PIN1 increases the interaction between PTP-PEST and FAK, which leads to the dephosphorylation of FAK Y397 and the promotion of migration, invasion, and metastasis of v-H-Ras-transformed cells. These findings uncover an important mechanism for the regulation of PTP-PEST in activated Ras-induced tumor progression.

  2. cis-Proline-mediated Ser(P)[superscript 5] Dephosphorylation by the RNA Polymerase II C-terminal Domain Phosphatase Ssu72

    Energy Technology Data Exchange (ETDEWEB)

    Werner-Allen, Jon W.; Lee, Chul-Jin; Liu, Pengda; Nicely, Nathan I.; Wang, Su; Greenleaf, Arno L.; Zhou, Pei (Duke)

    2012-05-16

    RNA polymerase II coordinates co-transcriptional events by recruiting distinct sets of nuclear factors to specific stages of transcription via changes of phosphorylation patterns along its C-terminal domain (CTD). Although it has become increasingly clear that proline isomerization also helps regulate CTD-associated processes, the molecular basis of its role is unknown. Here, we report the structure of the Ser(P){sup 5} CTD phosphatase Ssu72 in complex with substrate, revealing a remarkable CTD conformation with the Ser(P){sup 5}-Pro{sup 6} motif in the cis configuration. We show that the cis-Ser(P){sup 5}-Pro{sup 6} isomer is the minor population in solution and that Ess1-catalyzed cis-trans-proline isomerization facilitates rapid dephosphorylation by Ssu72, providing an explanation for recently discovered in vivo connections between these enzymes and a revised model for CTD-mediated small nuclear RNA termination. This work presents the first structural evidence of a cis-proline-specific enzyme and an unexpected mechanism of isomer-based regulation of phosphorylation, with broad implications for CTD biology

  3. The nonhistone, N-terminal tail of an essential, chimeric H2A variant regulates mitotic H3-S10 dephosphorylation.

    Science.gov (United States)

    Song, Xiaoyuan; Bowen, Josephine; Miao, Wei; Liu, Yifan; Gorovsky, Martin A

    2012-03-15

    H2A.Y is an essential, divergent Tetrahymena thermophila histone variant. It has a long nonhistone N terminus that contains leucine-rich repeats (LRR) and an LRR cap domain with similarity to Sds22p, a regulator of yeast protein phosphatase 1 (PP1) activity in the nucleus. In growing cells, H2A.Y is incorporated into micronuclei only during S phase, which occurs immediately after micronuclear mitosis. Depletion of H2A.Y causes prolonged retention of mitosis-associated histone H3-S10 phosphorylation and mitotic abnormalities that mimic S10E mutation. In cells where H2A.Y is depleted, an inducible chimeric gene, in which the H2A.Y N terminus is attached to H2A.X, is shown to regulate micronuclear H3-S10 phosphorylation. H2A.Y can also be specifically coimmunoprecipitated with a Tetrahymena PP1 ortholog (Ppo1p). Taken together, these results argue that the N terminus of H2A.Y functions to regulate H3-S10 dephosphorylation. This striking in vivo case of "cross-talk" between a H2A variant and a specific post-translational modification of another histone demonstrates a novel function for a histone variant.

  4. LoVo colon cancer cells resistant to oxaliplatin overexpress c-MET and VEGFR-1 and respond to VEGF with dephosphorylation of c-MET.

    Science.gov (United States)

    Mezquita, Belén; Pineda, Estela; Mezquita, Jovita; Mezquita, Pau; Pau, Montserrat; Codony-Servat, Jordi; Martínez-Balibrea, Eva; Mora, Conchi; Maurel, Joan; Mezquita, Cristóbal

    2016-05-01

    Oxaliplatin-resistant LoVo colon cancer cells overexpressing c-MET and VEGFR-1 were selected to study several signaling pathways involved in chemoresistance, as well as the effect of increasing amounts of VEGF in the regulation of c-MET. In comparison with chemosensitive LoVo colon cancer cells, oxaliplatin-resistant cells (LoVoR) overexpress and phosphorylate c-MET, upregulate the expression of transmembrane and soluble VEGFR-1 and, unexpectedly, downregulate VEGF. In addition, LoVoR cells activate other transduction pathways involved in chemoresistance such as Akt, β-catenin-TCF4 and E-cadherin. While c-MET is phosphorylated in LoVoR cells expressing low levels of VEGF, c-MET phosphorylation decreases when recombinant VEGF is added into the culture medium. Inhibition of c-MET by VEGF is mediated by VEGFR-1, since phosphorylation of c-MET in the presence of VEGF is restored after silencing VEGFR-1. Dephosphorylation of c-MET by VEGF suggests that tumors coexpressing VEGFR-1 and c-MET may activate c-MET as a result of anti-VEGF therapy. © 2015 Wiley Periodicals, Inc.

  5. Differential gene expressions of the MAPK signaling pathway in enterovirus 71-infected rhabdomyosarcoma cells

    Directory of Open Access Journals (Sweden)

    Weifeng Shi

    2013-08-01

    Full Text Available BACKGROUND: Mitogen-activated protein kinase (MAPK signaling pathway plays an important role in response to viral infection. The aim of this study was to explore the function and mechanism of MAPK signaling pathway in enterovirus 71 (EV71 infection of human rhabdomyosarcoma (RD cells. METHODS: Apoptosis of RD cells was observed using annexin V-FITC/PI binding assay under a fluorescence microscope. Cellular RNA was extracted and transcribed to cDNA. The expressions of 56 genes of MAPK signaling pathway in EV71-infected RD cells at 8 h and 20 h after infection were analyzed by PCR array. The levels of IL-2, IL-4, IL-10, and TNF-α in the supernatant of RD cells infected with EV71 at different time points were measured by ELISA. RESULTS: The viability of RD cells decreased obviously within 48 h after EV71 infection. Compared with the control group, EV71 infection resulted in the significantly enhanced releases of IL-2, IL-4, IL-10 and TNF-α from infected RD cells (p < 0.05. At 8 h after infection, the expressions of c-Jun, c-Fos, IFN-i, MEKK1, MLK3 and NIK genes in EV71-infected RD cells were up-regulated by 2.08-6.12-fold, whereas other 19 genes (e.g. AKT1, AKT2, E2F1, IKK and NF-κB1 exhibited down-regulation. However, at 20 h after infection, those MAPK signaling molecules including MEKK1, ASK1, MLK2, MLK3, NIK, MEK1, MEK2, MEK4, MEK7, ERK1, JNK1 and JNK2 were up-regulated. In addition, the expressions of AKT2, ELK1, c-Jun, c-Fos, NF-κB p65, PI3K and STAT1 were also increased. CONCLUSION: EV71 infection induces the differential gene expressions of MAPK signaling pathway such as ERK, JNK and PI3K/AKT in RD cells, which may be associated with the secretions of inflammatory cytokines and host cell apoptosis.

  6. Renoprotective Effect of Benazepril on Diabetic Nephropathy Mediated by P42/44MAPK

    Institute of Scientific and Technical Information of China (English)

    LV Yongman; DONG Junwu; NIU Xiaochun; LIU Xiaocheng

    2005-01-01

    The effects of benazepril on P42/44MAPK, angiotensin Ⅱ expression in renal tissue and renal pathological change of the experimental diabetic rats were assessed and the possible mechanism of benazepril's renoprotective effect was explored. Adult male Wistar rats, 11-12 weeks age,weighing initially 160 to 200 g were randomly allocated into 2 groups: control group (A, n=6) and experimental group (n= 12). Diabetic rats in experimental group were rendered diabetic by intraperitoneal injection of Streptozotocin (60 mg/kg body weight), and randomly subdivided into B group (diabetic control) and C group (diabetic rats treated with benazepril, 6 mg/kg every day).Studies were performed 8 weeks after induction of diabetes. Twenty-four h urine of every rat was collected to detect urine creatinine. Serum glucose concentration and serum creatinine were determined by collecting blood samples from the inferior vena cava. Body and kidney weight were recorded. Creatinine clearance (Ccr) and ratio of kidney weight to body weight were calculated. Plasma and renal tissue angiotensin Ⅱ concentration was assayed by radioimmunoassay (RIA). The phospo-p44/42MAPK protein expression was detected by Western-blot. The results showed that benazepril had no significant effect on the blood glucose level in diabetic rats in two experimental groups.Ccr and ratio of kidney weight to body weight were increased in group B (P<0. 01) as coapared with normal rats at the end of the 8th week. At the end of the 8th week, Ccr in group C was lower than that in group B (P<0.01). The ratio of kidney weight to body weight in group C was lower than that in group B at the 8th week. There were glomeruli hypertrophy and slight or moderate mesangium proliferation in diabetic rats, while there was fragmentally proliferative mesangium in group C at the end of the 8th week. Renal tissue angiotensin Ⅱ concentration was significantly increased in group B, while benazepril could significantly decrease the

  7. Apoptosis induction by the satratoxins and other trichothecene mycotoxins: relationship to ERK, p38 MAPK, and SAPK/JNK activation.

    Science.gov (United States)

    Yang, G H; Jarvis, B B; Chung, Y J; Pestka, J J

    2000-04-15

    The satratoxins are members of the trichothecene mycotoxin family that are produced by the fungus Stachybotrys and that have been etiologically associated with building-related health problems. The purpose of this study was to relate cytotoxic and apoptotic capacities of satratoxins and other trichothecenes to the activation of three groups of mitogen-activated protein kinases (MAPKs) (extracellular signal-regulated protein kinase (ERK), p38 MAPK, and stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK)). Two myeloid models, RAW 264.7 murine macrophage and U937 human leukemic cells were used. Upon evaluating representative trichothecenes in the 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyl tetrazolium bromide (MTT) cleavage assay, cytotoxicity was evident according to the following rank order: satratoxin G, roridin A, and verrucarin A > T-2 toxin, satratoxin F, H > nivalenol, and vomitoxin. Comparable results were found when measuring trichothecene-mediated apoptosis using DNA fragmentation and fluorescence microscopy assays, thus suggesting that cytotoxicity was mediated through an apoptotic process. Assessment of MAPK activation using Western blot analysis revealed that trichothecenes activated not only SAPK/JNK and p38 MAPK but also ERK. Activation of MAPKs by satratoxins and other trichothecenes correlated with and preceded apoptosis. The concentration of satratoxin G sufficient for protein synthesis inhibition correlated with that required for apoptosis and activation of all three MAPKs. Cycloheximide had similar effects to trichothecenes, suggesting that ribosome binding or protein synthesis inhibition may play roles in MAPK activation and apoptosis induction. Apoptosis induction by satratoxin G and vomitoxin was markedly enhanced when ERK activation was selectively inhibited by ERK-specific inhibitor PD98059, thus indicating a negative role for ERK. Inhibition of p38 MAPK activity with the p38-specific inhibitor SB203580 had no effect on apoptosis

  8. Zinc rescues obesity-induced cardiac hypertrophy via stimulating metallothionein to suppress oxidative stress-activated BCL10/CARD9/p38 MAPK pathway.

    Science.gov (United States)

    Wang, Shudong; Gu, Junlian; Xu, Zheng; Zhang, Zhiguo; Bai, Tao; Xu, Jianxiang; Cai, Jun; Barnes, Gregory; Liu, Qiu-Ju; Freedman, Jonathan H; Wang, Yonggang; Liu, Quan; Zheng, Yang; Cai, Lu

    2017-02-03

    Obesity often leads to obesity-related cardiac hypertrophy (ORCH), which is suppressed by zinc-induced inactivation of p38 mitogen-activated protein kinase (p38 MAPK). In this study, we investigated the mechanisms by which zinc inactivates p38 MAPK to prevent ORCH. Mice (4-week old) were fed either high fat diet (HFD, 60% kcal fat) or normal diet (ND, 10% kcal fat) containing variable amounts of zinc (deficiency, normal and supplement) for 3 and 6 months. P38 MAPK siRNA and the p38 MAPK inhibitor SB203580 were used to suppress p38 MAPK activity in vitro and in vivo, respectively. HFD activated p38 MAPK and increased expression of B-cell lymphoma/CLL 10 (BCL10) and caspase recruitment domain family member 9 (CARD9). These responses were enhanced by zinc deficiency and attenuated by zinc supplement. Administration of SB203580 to HFD mice or specific siRNA in palmitate-treated cardiomyocytes eliminated the HFD and zinc deficiency activation of p38 MAPK, but did not significantly impact the expression of BCL10 and CARD9. In cultured cardiomyocytes, inhibition of BCL10 expression by siRNA prevented palmitate-induced increased p38 MAPK activation and atrial natriuretic peptide (ANP) expression. In contrast, inhibition of p38 MAPK prevented ANP expression, but did not affect BCL10 expression. Deletion of metallothionein abolished the protective effect of zinc on palmitate-induced up-regulation of BCL10 and phospho-p38 MAPK. HFD and zinc deficiency synergistically induce ORCH by increasing oxidative stress-mediated activation of BCL10/CARD9/p38 MAPK signalling. Zinc supplement ameliorates ORCH through activation of metallothionein to repress oxidative stress-activated BCL10 expression and p38 MAPK activation.

  9. The Protective Effect of Beraprost Sodium on Diabetic Nephropathy by Inhibiting Inflammation and p38 MAPK Signaling Pathway in High-Fat Diet/Streptozotocin-Induced Diabetic Rats

    OpenAIRE

    Li Peng; Jie Li; Yixing Xu; Yangtian Wang; Hong Du; Jiaqing Shao; Zhimin Liu

    2016-01-01

    Background. p38 mitogen-activated protein kinase (MAPK) plays a crucial role in regulating signaling pathways implicated in inflammatory processes leading to diabetic nephropathy (DN). This study aimed to examine p38 MAPK activation in DN and determine whether beraprost sodium (BPS) ameliorates DN by inhibiting inflammation and p38 MAPK signaling pathway in diabetic rats. Methods. Forty male Sprague Dawley (SD) rats were randomly divided into the normal control group, type 2 diabetic group, a...

  10. Sesamin suppresses activation of microglia and p44/42 MAPK pathway, which confers neuroprotection in rat intracerebral hemorrhage.

    Science.gov (United States)

    Ohnishi, M; Monda, A; Takemoto, R; Matsuoka, Y; Kitamura, C; Ohashi, K; Shibuya, H; Inoue, A

    2013-03-01

    Thrombin plays important roles in the pathology of intracerebral hemorrhage (ICH). The recruitment of activated microglia, accompanied by thrombin-induced phosphorylation of the mitogen-activated protein kinase (MAPK) family, contributes to ICH-associated neuron loss. Here we investigated the possibility that sesamin, a lignan of sesame seed oil, is a natural candidate as an inhibitor of microglial activation and MAPK pathways under ICH insults. Sesamin (30-100 μM) suppressed thrombin-induced nitric oxide (NO) production by primary-cultured rat microglia via inhibition of inducible NO synthase (iNOS) protein expression, independently of the antioxidative effect. Sesamin selectively inhibited p44/42 MAPK phosphorylation in the MAPK family (p38 and p44/42) involved in iNOS protein expression in primary-cultured rat microglia. An in vivo rat ICH model was prepared by intrastriatal injection of 0.20U collagenase type IV unilaterally. ICH evoked the phosphorylation of p44/42 MAPK, microglial proliferation with morphological change into the activated ameboid form, and neuron loss. The phosphorylation of p44/42 MAPK was inhibited by intracerebroventricular administration of 30-nmol sesamin. Sesamin prevented ICH-induced increase of microglial cells in the perihematomal area. Notably, ramified microglia, the resting morphology, were observed in brain sections of the animals administrated sesamin. Sesamin furthermore achieved neuroprotection in the perihematomal area but not in the hematomal center. These results suggest that sesamin is a promising natural product as a novel therapeutic strategy based on the regulation of microglial activities accompanied by the activated p44/42 MAPK pathway in ICH.

  11. Exocyst Sec10 protects renal tubule cells from injury by EGFR/MAPK activation and effects on endocytosis.

    Science.gov (United States)

    Fogelgren, Ben; Zuo, Xiaofeng; Buonato, Janine M; Vasilyev, Aleksandr; Baek, Jeong-In; Choi, Soo Young; Chacon-Heszele, Maria F; Palmyre, Aurélien; Polgar, Noemi; Drummond, Iain; Park, Kwon Moo; Lazzara, Matthew J; Lipschutz, Joshua H

    2014-12-15

    Acute kidney injury is common and has a high mortality rate, and no effective treatment exists other than supportive care. Using cell culture models, we previously demonstrated that exocyst Sec10 overexpression reduced damage to renal tubule cells and speeded recovery and that the protective effect was mediated by higher basal levels of mitogen-activated protein kinase (MAPK) signaling. The exocyst, a highly-conserved eight-protein complex, is known for regulating protein trafficking. Here we show that the exocyst biochemically interacts with the epidermal growth factor receptor (EGFR), which is upstream of MAPK, and Sec10-overexpressing cells express greater levels of phosphorylated (active) ERK, the final step in the MAPK pathway, in response to EGF stimulation. EGFR endocytosis, which has been linked to activation of the MAPK pathway, increases in Sec10-overexpressing cells, and gefitinib, a specific EGFR inhibitor, and Dynasore, a dynamin inhibitor, both reduce EGFR endocytosis. In turn, inhibition of the MAPK pathway reduces ligand-mediated EGFR endocytosis, suggesting a potential feedback of elevated ERK activity on EGFR endocytosis. Gefitinib also decreases MAPK signaling in Sec10-overexpressing cells to levels seen in control cells and, demonstrating a causal role for EGFR, reverses the protective effect of Sec10 overexpression following cell injury in vitro. Finally, using an in vivo zebrafish model of acute kidney injury, morpholino-induced knockdown of sec10 increases renal tubule cell susceptibility to injury. Taken together, these results suggest that the exocyst, acting through EGFR, endocytosis, and the MAPK pathway is a candidate therapeutic target for acute kidney injury.

  12. N-methyl-D-aspartate (NMDA) and the regulation of mitogen-activated protein kinase (MAPK) signaling pathways: a revolving neurochemical axis for therapeutic intervention?

    Science.gov (United States)

    Haddad, John J

    2005-11-01

    Excitatory synaptic transmission in the central nervous system (CNS) is mediated by the release of glutamate from presynaptic terminals onto postsynaptic channels gated by N-methyl-D-aspartate (NMDA) and non-NMDA (AMPA and KA) receptors. Extracellular signals control diverse neuronal functions and are responsible for mediating activity-dependent changes in synaptic strength and neuronal survival. Influx of extracellular calcium ([Ca(2+)](e)) through the NMDA receptor (NMDAR) is required for neuronal activity to change the strength of many synapses. At the molecular level, the NMDAR interacts with signaling modules, which, like the mitogen-activated protein kinase (MAPK) superfamily, transduce excitatory signals across neurons. Recent burgeoning evidence points to the fact that MAPKs play a crucial role in regulating the neurochemistry of NMDARs, their physiologic and biochemical/biophysical properties, and their potential role in pathophysiology. It is the purpose of this review to discuss: (i) the MAPKs and their role in a plethora of cellular functions; (ii) the role of MAPKs in regulating the biochemistry and physiology of NMDA receptors; (iii) the kinetics of MAPK-NMDA interactions and their biologic and neurochemical properties; (iv) how cellular signaling pathways, related cofactors and intracellular conditions affect NMDA-MAPK interactions and (v) the role of NMDA-MAPK pathways in pathophysiology and the evolution of disease conditions. Given the versatility of the NMDA-MAPK interactions, the NMDA-MAPK axis will likely form a neurochemical target for therapeutic interventions.

  13. Model simulations of cooking organic aerosol (COA) over the UK using estimates of emissions based on measurements at two sites in London

    Science.gov (United States)

    Ots, Riinu; Vieno, Massimo; Allan, James D.; Reis, Stefan; Nemitz, Eiko; Young, Dominique E.; Coe, Hugh; Di Marco, Chiara; Detournay, Anais; Mackenzie, Ian A.; Green, David C.; Heal, Mathew R.

    2016-11-01

    Cooking organic aerosol (COA) is currently not included in European emission inventories. However, recent positive matrix factorization (PMF) analyses of aerosol mass spectrometer (AMS) measurements have suggested important contributions of COA in several European cities. In this study, emissions of COA were estimated for the UK, based on hourly AMS measurements of COA made at two sites in London (a kerbside site in central London and an urban background site in a residential area close to central London) for the full calendar year of 2012 during the Clean Air for London (ClearfLo) campaign. Iteration of COA emissions estimates and subsequent evaluation and sensitivity experiments were conducted with the EMEP4UK atmospheric chemistry transport modelling system with a horizontal resolution of 5 km × 5 km. The spatial distribution of these emissions was based on workday population density derived from the 2011 census data. The estimated UK annual COA emission was 7.4 Gg per year, which is an almost 10 % addition to the officially reported UK national total anthropogenic emissions of PM2.5 (82 Gg in 2012), corresponding to 320 mg person-1 day-1 on average. Weekday and weekend diurnal variation in COA emissions were also based on the AMS measurements. Modelled concentrations of COA were then independently evaluated against AMS-derived COA measurements from another city and time period (Manchester, January-February 2007), as well as with COA estimated by a chemical mass balance model of measurements for a 2-week period at the Harwell rural site (˜ 80 km west of central London). The modelled annual average contribution of COA to ambient particulate matter (PM) in central London was between 1 and 2 µg m-3 (˜ 20 % of total measured OA1) and between 0.5 and 0.7 µg m-3 in other major cities in England (Manchester, Birmingham, Leeds). It was also shown that cities smaller than London can have a central hotspot of population density of smaller area than the

  14. The role of the organic layer for phosphorus nutrition of young beech trees (Fagus sylvatica L.) at two sites differing in soil Phosphorus availability

    Science.gov (United States)

    Hauenstein, Simon

    2016-04-01

    Simon Hauenstein1, Thomas Pütz2, and Yvonne Oelmann1, 1 Geoecology, Department of Geosciences, University of Tübingen, Tübingen, Germany 2 Agrosphere (IBG-3), Forschungszentrum Jülich, Jülich, Germany The accumulation of an organic layer in forests is linked to the ratio between litterfall rates and decomposition rates with decomposition rates being decelerated due to acidification and associated nutrient depletion with proceeding ecosystem development. Nevertheless, the nutrient pool in the organic layer might still represent an important source for Phosphorus (P) nutrition of forests on nutrient-poor soils. Our objective was to assess the importance of the organic layer to P nutrition of young beech trees at two sites differing in soil P availability. We established a mesocosm experiment including plants and soil from a Phosphorus depleted forest site on a Haplic Podzol in Lüss and a Phosphorus rich forest site on a Eutric Cambisol in Bad Brückenau either with or without the organic layer. After 1 year under outdoor conditions, we applied 33P to the pots. After 0h, 24h, 48h, 96h, 192h, 528h we destructively harvested the young beech trees (separated into leaves, branches, stems) and sampled the organic layer and mineral soil of the pots. In each soil horizon we measured concentrations of resin-extractable P, plant available P fractions and total P. We extracted the xylem sap of the whole 2-year-old trees by means of scholander pressure bomb. 33P activity was measured for every compartment in soil and plant. The applied 33P was recovered mainly in the organic layer in Lüss, whereas it was evenly distributed among organic and mineral horizons in pots of Bad Brückenau soil. Comparing pots with and without an organic layer, the specific 33P activity differed by 323% between pots with and without an organic layer present in the Lüss soil. For both sites, the presence of the organic layer increased 33P activity in xylem sap compared to the treatment without

  15. REX-1 expression and p38 MAPK activation status can determine proliferation/differentiation fates in human mesenchymal stem cells.

    Directory of Open Access Journals (Sweden)

    Dilli Ram Bhandari

    Full Text Available BACKGROUND: REX1/ZFP42 is a well-known embryonic stem cell (ESC marker. However, the role of REX1, itself, is relatively unknown because the function of REX1 has only been reported in the differentiation of ESCs via STAT signaling pathways. Human mesenchymal stem cells (hMSCs isolated from young tissues and cancer cells express REX1. METHODOLOGY/PRINCIPAL FINDING: Human umbilical cord blood-derived MSCs (hUCB-MSCs and adipose tissue-derived MSCs (hAD-MSCs strongly express REX1 and have a lower activation status of p38 MAPK, but bone marrow-derived MSCs (hBM-MSCs have weak REX1 expression and higher activation of p38 MAPK. These results indicated that REX1 expression in hMSCs was positively correlated with proliferation rates but inversely correlated with the phosphorylation of p38 MAPK. In hUCB-MSCs, the roles of REX1 and p38 MAPK were investigated, and a knockdown study was performed using a lentiviral vector-based small hairpin RNA (shRNA. After REX1 knockdown, decreased cell proliferation was observed. In REX1 knocked-down hUCB-MSCs, the osteogenic differentiation ability deteriorated, but the adipogenic potential increased or was similar to that observed in the controls. The phosphorylation of p38 MAPK in hUCB-MSCs significantly increased after REX1 knockdown. After p38 MAPK inhibitor treatment, the cell growth in REX1 knocked-down hUCB-MSCs almost recovered, and the suppressed expression levels of CDK2 and CCND1 were also restored. The expression of MKK3, an upstream regulator of p38 MAPK, significantly increased in REX1 knocked-down hUCB-MSCs. The direct binding of REX1 to the MKK3 gene was confirmed by a chromatin immunoprecipitation (ChIP assay. CONCLUSIONS/SIGNIFICANCE: These findings showed that REX1 regulates the proliferation/differentiation of hMSCs through the suppression of p38 MAPK signaling via the direct suppression of MKK3. Therefore, p38 MAPK and REX-1 status can determine the cell fate of adult stem cells (ASCs. These

  16. Metabolic respiration induces AMPK- and Ire1p-dependent activation of the p38-Type HOG MAPK pathway.

    Science.gov (United States)

    Adhikari, Hema; Cullen, Paul J

    2014-10-01

    Evolutionarily conserved mitogen activated protein kinase (MAPK) pathways regulate the response to stress as well as cell differentiation. In Saccharomyces cerevisiae, growth in non-preferred carbon sources (like galactose) induces differentiation to the filamentous cell type through an extracellular-signal regulated kinase (ERK)-type MAPK pathway. The filamentous growth MAPK pathway shares components with a p38-type High Osmolarity Glycerol response (HOG) pathway, which regulates the response to changes in osmolarity. To determine the extent of functional overlap between the MAPK pathways, comparative RNA sequencing was performed, which uncovered an unexpected role for the HOG pathway in regulating the response to growth in galactose. The HOG pathway was induced during growth in galactose, which required the nutrient regulatory AMP-dependent protein kinase (AMPK) Snf1p, an intact respiratory chain, and a functional tricarboxylic acid (TCA) cycle. The unfolded protein response (UPR) kinase Ire1p was also required for HOG pathway activation in this context. Thus, the filamentous growth and HOG pathways are both active during growth in galactose. The two pathways redundantly promoted growth in galactose, but paradoxically, they also inhibited each other's activities. Such cross-modulation was critical to optimize the differentiation response. The human fungal pathogen Candida albicans showed a similar regulatory circuit. Thus, an evolutionarily conserved regulatory axis links metabolic respiration and AMPK to Ire1p, which regulates a differentiation response involving the modulated activity of ERK and p38 MAPK pathways.

  17. Effect of Repeated Electroacupuncture Intervention on Hippocampal ERK and p38MAPK Signaling in Neuropathic Pain Rats

    Directory of Open Access Journals (Sweden)

    Jun-ying Wang

    2015-01-01

    Full Text Available Results of our past studies showed that hippocampal muscarinic acetylcholine receptor (mAChR-1 mRNA and differentially expressed proteins participating in MAPK signaling were involved in electroacupuncture (EA induced cumulative analgesia in neuropathic pain rats, but the underlying intracellular mechanism remains unknown. The present study was designed to observe the effect of EA stimulation (EAS on hippocampal extracellular signal-regulated kinases (ERK and p38 MAPK signaling in rats with chronic constrictive injury (CCI of the sciatic nerve, so as to reveal its related intracellular targets in pain relief. After CCI, the thermal pain thresholds of the affected hind were significantly decreased compared with the control group (P<0.05. Following one and two weeks’ EAS of ST 36-GB34, the pain thresholds were significantly upregulated (P<0.05, and the effect of EA2W was remarkably superior to that of EA2D and EA1W (P<0.05. Correspondingly, CCI-induced decreased expression levels of Ras, c-Raf, ERK1 and p-ERK1/2 proteins, and p38 MAPK mRNA and p-p38MAPK protein in the hippocampus tissues were reversed by EA2W (P<0.05. The above mentioned results indicated that EA2W induced cumulative analgesic effect may be closely associated with its function in removing neuropathic pain induced suppression of intracellular ERK and p38MAPK signaling in the hippocampus.

  18. Deoxycholic acid mediates non-canonical EGFR-MAPK activation through the induction of calcium signaling in colon cancer cells.

    Science.gov (United States)

    Centuori, Sara M; Gomes, Cecil J; Trujillo, Jesse; Borg, Jamie; Brownlee, Joshua; Putnam, Charles W; Martinez, Jesse D

    2016-07-01

    Obesity and a western diet have been linked to high levels of bile acids and the development of colon cancer. Specifically, increased levels of the bile acid deoxycholic acid (DCA), an established tumor promoter, has been shown to correlate with increased development of colorectal adenomas and progression to carcinoma. Herein we investigate the mechanism by which DCA leads to EGFR-MAPK activation, a candidate mechanism by which DCA may promote colorectal tumorigenesis. DCA treated colon cancer cells exhibited strong and prolonged activation of ERK1/2 when compared to EGF treatment alone. We also showed that DCA treatment prevents EGFR degradation as opposed to the canonical EGFR recycling observed with EGF treatment. Moreover, the combination of DCA and EGF treatment displayed synergistic activity, suggesting DCA activates MAPK signaling in a non-canonical manner. Further evaluation showed that DCA treatment increased intracellular calcium levels and CAMKII phosphorylation, and that blocking calcium with BAPTA-AM abrogated MAPK activation induced by DCA, but not by EGF. Finally we showed that DCA-induced CAMKII leads to MAPK activation through the recruitment of c-Src. Taken together, we demonstrated that DCA regulates MAPK activation through calcium signaling, an alternative mechanism not previously recognized in human colon cancer cells. Importantly, this mechanism allows for EGFR to escape degradation and thus achieve a constitutively active state, which may explain its tumor promoting effects.

  19. Calcium paradox induces apoptosis in the isolated perfused Rana ridibunda heart: involvement of p38-MAPK and calpain.

    Science.gov (United States)

    Aggeli, Ioanna-Katerina; Zacharias, Triantafyllos; Papapavlou, Georgia; Gaitanaki, Catherine; Beis, Isidoros

    2013-12-01

    "Calcium paradox" as a term describes the deleterious effects conferred to a heart perfused with a calcium-free solution followed by repletion, including loss of mechanical activity and sarcomere disruption. Given that the signaling mechanisms triggered by calcium paradox remain elusive, in the present study, we tried to investigate them in the isolated perfused heart from Rana ridibunda. Calcium paradox was found to markedly activate members of the MAPKs (p43-ERK, JNKs, p38-MAPK). In addition to lactate dehydrogenase (LDH) release in the perfusate (indicative of necrosis), we also confirmed the occurrence of apoptosis by using the TUNEL assay and identifying poly(ADP-ribose) polymerase (PARP) fragmentation and upregulated Bax expression. Furthermore, using MDL28170 (a selective calpain inhibitor), a role for this protease was revealed. In addition, various divalent cations were shown to exert a protective effect against the calcium paradox. Interestingly, SB203580, a p38-MAPK inhibitor, alleviated calcium-paradox-conferred apoptosis. This result indicates that p38-MAPK plays a pro-apoptotic role, contributing to the resulting myocardial dysfunction and cell death. To our knowledge, this is the first time that the calcium paradox has been shown to induce apoptosis in amphibians, with p38-MAPK and calpain playing significant roles.

  20. MAPK13 is preferentially expressed in gynecological cancer stem cells and has a role in the tumor-initiation

    Energy Technology Data Exchange (ETDEWEB)

    Yasuda, Kazuyo [Department of Pathology, Sapporo Medical University School of Medicine, South-1 West-17, Chuo-Ku, Sapporo, 060-8556 (Japan); Hirohashi, Yoshihiko, E-mail: hirohash@sapmed.ac.jp [Department of Pathology, Sapporo Medical University School of Medicine, South-1 West-17, Chuo-Ku, Sapporo, 060-8556 (Japan); Kuroda, Takafumi [Department of Obstetrics and Gynecology, Sapporo Medical University School of Medicine, South-1 West-17, Chuo-Ku, Sapporo, 060-8556 (Japan); Takaya, Akari; Kubo, Terufumi; Kanaseki, Takayuki; Tsukahara, Tomohide [Department of Pathology, Sapporo Medical University School of Medicine, South-1 West-17, Chuo-Ku, Sapporo, 060-8556 (Japan); Hasegawa, Tadashi [Department of Surgical Pathology, Sapporo Medical University School of Medicine, South-1 West-17, Chuo-Ku, Sapporo, 060-8556 (Japan); Saito, Tsuyoshi [Department of Obstetrics and Gynecology, Sapporo Medical University School of Medicine, South-1 West-17, Chuo-Ku, Sapporo, 060-8556 (Japan); Sato, Noriyuki [Department of Pathology, Sapporo Medical University School of Medicine, South-1 West-17, Chuo-Ku, Sapporo, 060-8556 (Japan); Torigoe, Toshihiko, E-mail: torigoe@sapmed.ac.jp [Department of Pathology, Sapporo Medical University School of Medicine, South-1 West-17, Chuo-Ku, Sapporo, 060-8556 (Japan)

    2016-04-15

    Cancer stem-like cells (CSCs)/cancer-initiating cells (CICs) are defined as small subpopulation of cancer cells that are endowed with higher tumor-initiating ability. CSCs/CICs are resistant to standard cancer therapies including chemotherapy and radiotherapy, and they are thus thought to be responsible for cancer recurrence and metastasis. Therefore, elucidation of molecular mechanisms of CSCs/CICs is essential to cure cancer. In this study, we analyzed the gene expression profiles of gynecological CSCs/CICs isolated as aldehyde dehydrogenase high (ALDH{sup high}) cells, and found that MAPK13, PTTG1IP, CAPN1 and UBQLN2 were preferentially expressed in CSCs/CICs. MAPK13 is expressed in uterine, ovary, stomach, colon, liver and kidney cancer tissues at higher levels compared with adjacent normal tissues. MAPK13 gene knockdown using siRNA reduced the ALDH{sup high} population and abrogated the tumor-initiating ability. These results indicate that MAPK13 is expressed in gynecological CSCs/CICs and has roles in the maintenance of CSCs/CICs and tumor-initiating ability, and MAPK13 might be a novel molecular target for treatment-resistant CSCs/CICs.

  1. MAPK genes interact with diet and lifestyle factors to alter risk of breast cancer: the Breast Cancer Health Disparities Study.

    Science.gov (United States)

    Slattery, Martha L; Lundgreen, Abbie; John, Esther M; Torres-Mejia, Gabriela; Hines, Lisa; Giuliano, Anna R; Baumgartner, Kathy B; Stern, Mariana C; Wolff, Roger K

    2015-01-01

    Mitogen-activated protein kinases (MAPK) are integration points for multiple biochemical signals. We evaluated 13 MAPK genes with breast cancer risk and determined if diet and lifestyle factors mediated risk. Data from 3 population-based case-control studies conducted in Southwestern United States, California, and Mexico included 4183 controls and 3592 cases. Percent Indigenous American (IA) ancestry was determined from 104 ancestry informative markers. The adaptive rank truncated product (ARTP) was used to determine the significance of each gene and the pathway with breast cancer risk, by menopausal status, genetic ancestry level, and estrogen receptor (ER)/progesterone receptor (PR) strata. MAP3K9 was associated with breast cancer overall (P(ARTP) = 0.02) with strongest association among women with the highest IA ancestry (P(ARTP) = 0.04). Several SNPs in MAP3K9 were associated with ER+/PR+ tumors and interacted with dietary oxidative balance score (DOBS), dietary folate, body mass index (BMI), alcohol consumption, cigarette smoking, and a history of diabetes. DUSP4 and MAPK8 interacted with calories to alter breast cancer risk; MAPK1 interacted with DOBS, dietary fiber, folate, and BMI; MAP3K2 interacted with dietary fat; and MAPK14 interacted with dietary folate and BMI. The patterns of association across diet and lifestyle factors with similar biological properties for the same SNPs within genes provide support for associations.

  2. Single-cell analysis reveals that insulation maintains signaling specificity between two yeast MAPK pathways with common components.

    Science.gov (United States)

    Patterson, Jesse C; Klimenko, Evguenia S; Thorner, Jeremy

    2010-10-19

    Eukaryotic cells use multiple mitogen-activated protein kinase (MAPK) cascades to evoke appropriate responses to external stimuli. In Saccharomyces cerevisiae, the MAPK Fus3 is activated by pheromone-binding heterotrimeric guanosine triphosphate-binding protein (G protein)-coupled receptors to promote mating, whereas the MAPK Hog1 is activated by hyperosmotic stress to elicit the high-osmolarity glycerol (HOG) response. Although these MAPK pathways share several upstream components, exposure to either pheromone or osmolyte alone triggers only the appropriate response. We used fluorescence localization- and transcription-specific reporters to assess activation of these pathways in individual cells on the minute and hour time scale, respectively. Dual activation of these two MAPK pathways occurred over a broad range of stimulant concentrations and temporal regimes in wild-type cells subjected to costimulation. Thus, signaling specificity is achieved through an "insulation" mechanism, not a "cross-inhibition" mechanism. Furthermore, we showed that there was a critical period during which Hog1 activity had to occur for proper insulation of the HOG pathway.

  3. Astaxanthin Pretreatment Attenuates Hepatic Ischemia Reperfusion-Induced Apoptosis and Autophagy via the ROS/MAPK Pathway in Mice

    Directory of Open Access Journals (Sweden)

    Jingjing Li

    2015-05-01

    Full Text Available Background: Hepatic ischemia reperfusion (IR is an important issue in complex liver resection and liver transplantation. The aim of the present study was to determine the protective effect of astaxanthin (ASX, an antioxidant, on hepatic IR injury via the reactive oxygen species/mitogen-activated protein kinase (ROS/MAPK pathway. Methods: Mice were randomized into a sham, IR, ASX or IR + ASX group. The mice received ASX at different doses (30 mg/kg or 60 mg/kg for 14 days. Serum and tissue samples at 2 h, 8 h and 24 h after abdominal surgery were collected to assess alanine aminotransferase (ALT, aspartate aminotransferase (AST, inflammation factors, ROS, and key proteins in the MAPK family. Results: ASX reduced the release of ROS and cytokines leading to inhibition of apoptosis and autophagy via down-regulation of the activated phosphorylation of related proteins in the MAPK family, such as P38 MAPK, JNK and ERK in this model of hepatic IR injury. Conclusion: Apoptosis and autophagy caused by hepatic IR injury were inhibited by ASX following a reduction in the release of ROS and inflammatory cytokines, and the relationship between the two may be associated with the inactivation of the MAPK family.

  4. RAS-MAPK dependence underlies a rational polytherapy strategy in EML4-ALK–positive lung cancer

    Science.gov (United States)

    Hrustanovic, Gorjan; Olivas, Victor; Pazarentzos, Evangelos; Tulpule, Asmin; Asthana, Saurabh; Blakely, Collin M; Okimoto, Ross A; Lin, Luping; Neel, Dana S; Sabnis, Amit; Flanagan, Jennifer; Chan, Elton; Varella-Garcia, Marileila; Aisner, Dara L; Vaishnavi, Aria; Ou, Sai-Hong I; Collisson, Eric A; Ichihara, Eiki; Mack, Philip C; Lovly, Christine M; Karachaliou, Niki; Rosell, Rafael; Riess, Jonathan W; Doebele, Robert C; Bivona, Trever G

    2016-01-01

    One strategy for combating cancer-drug resistance is to deploy rational polytherapy up front that suppresses the survival and emergence of resistant tumor cells. Here we demonstrate in models of lung adenocarcinoma harboring the oncogenic fusion of ALK and EML4 that the GTPase RAS–mitogen-activated protein kinase (MAPK) pathway, but not other known ALK effectors, is required for tumor-cell survival. EML4-ALK activated RAS-MAPK signaling by engaging all three major RAS isoforms through the HELP domain of EML4. Reactivation of the MAPK pathway via either a gain in the number of copies of the gene encoding wild-type K-RAS (KRASWT) or decreased expression of the MAPK phosphatase DUSP6 promoted resistance to ALK inhibitors in vitro, and each was associated with resistance to ALK inhibitors in individuals with EML4-ALK–positive lung adenocarcinoma. Upfront inhibition of both ALK and the kinase MEK enhanced both the magnitude and duration of the initial response in preclinical models of EML4-ALK lung adenocarcinoma. Our findings identify RAS-MAPK dependence as a hallmark of EML4-ALK lung adenocarcinoma and provide a rationale for the upfront inhibition of both ALK and MEK to forestall resistance and improve patient outcomes. PMID:26301689

  5. Metabolic respiration induces AMPK- and Ire1p-dependent activation of the p38-Type HOG MAPK pathway.

    Directory of Open Access Journals (Sweden)

    Hema Adhikari

    2014-10-01

    Full Text Available Evolutionarily conserved mitogen activated protein kinase (MAPK pathways regulate the response to stress as well as cell differentiation. In Saccharomyces cerevisiae, growth in non-preferred carbon sources (like galactose induces differentiation to the filamentous cell type through an extracellular-signal regulated kinase (ERK-type MAPK pathway. The filamentous growth MAPK pathway shares components with a p38-type High Osmolarity Glycerol response (HOG pathway, which regulates the response to changes in osmolarity. To determine the extent of functional overlap between the MAPK pathways, comparative RNA sequencing was performed, which uncovered an unexpected role for the HOG pathway in regulating the response to growth in galactose. The HOG pathway was induced during growth in galactose, which required the nutrient regulatory AMP-dependent protein kinase (AMPK Snf1p, an intact respiratory chain, and a functional tricarboxylic acid (TCA cycle. The unfolded protein response (UPR kinase Ire1p was also required for HOG pathway activation in this context. Thus, the filamentous growth and HOG pathways are both active during growth in galactose. The two pathways redundantly promoted growth in galactose, but paradoxically, they also inhibited each other's activities. Such cross-modulation was critical to optimize the differentiation response. The human fungal pathogen Candida albicans showed a similar regulatory circuit. Thus, an evolutionarily conserved regulatory axis links metabolic respiration and AMPK to Ire1p, which regulates a differentiation response involving the modulated activity of ERK and p38 MAPK pathways.

  6. Sorbitol induces apoptosis of human colorectal cancer cells via p38 MAPK signal transduction.

    Science.gov (United States)

    Lu, Xue; Li, Chun; Wang, Yong-Kun; Jiang, Kun; Gai, Xiao-Dong

    2014-06-01

    Sorbitol has been reported to have anticancer effects in several tumor models, however its effects on colorectal cancer remain elusive. In the present study, the effects of sorbitol on growth inhibition and apoptosis in the colorectal cancer HCT116 cell line were evaluated and its mechanism of action was examined. An MTT assay was utilized to determine the effect of sorbitol on HCT116 cell proliferation at different time points and variable doses. Western blot analysis was used to examine the effect of sorbitol on apoptosis-related protein expression and the p38 MAPK signaling pathway. The results revealed that sorbitol may inhibit the growth of HCT116 cells in a time- and dose-dependent manner. Following treatment with sorbitol for 3 h, western blotting demonstrated cleavage of the caspase-3 zymogen protein and a cleavage product of poly (ADP-ribose) polymerase (PARP), a known substrate of caspase-3, was also evident. During sorbitol-induced apoptosis, the mitochondrial pathway was activated by a dose-dependent increase in Bax expression and cytochrome c release, while the expression of anti-apoptotic protein Bcl-2 was significantly decreased in a dose-dependent manner. The investigation for the downstream signal pathway revealed that sorbitol-induced apoptosis was mediated by an increase in phosphorylated p38 MAPK expression. Overall, the observations from the present study imply that sorbitol causes increased levels of Bax in response to p38 MAPK signaling, which results in the initiation of the mitochondrial death cascade. Therefore, sorbitol is a promising candidate as a potential chemotherapeutic agent for the treatment of colorectal cancer HCT116 cells.

  7. Protective Role of Fucoidan in Cerebral Ischemia-Reperfusion Injury through Inhibition of MAPK Signaling Pathway.

    Science.gov (United States)

    Che, Nan; Ma, Yijie; Xin, Yinhu

    2016-11-25

    Fucoidan has been reported to exhibit various beneficial activities ranging from to antivirus and anticancer properties. However, little information is available about the effects of fucoidan on cerebral ischemia-reperfusion injury (IRI). Our study aimed to explore the effects of fucoidan on cerebral IRI, as well as the underlying mechanisms. Sprague-Dawley (SD) rats were randomly subjected to four groups: Sham, IRI+saline (IRI+S), IRI+80 mg/kg fucoidan (IRI+F80), and IRI+160 mg/kg fucoidan (IRI+F160). Fucoidan (80 mg/kg or 160 mg/kg) was intraperitoneally injected from 7 days before the rats were induced to cerebral IRI model with middle cerebral artery occlusion (MCAO) method. At 24 h after reperfusion, neurological deficits and the total infarct volume were determined. The levels of inflammation-associated cytokines (interleukin (IL)-1β, IL-6, myeloperoxidase (MPO), and tumor necrosis factor (TNF)-α), oxidative stress-related proteins (malondialdehyde (MDA) and superoxide dismutase (SOD)) in the ischemic brain were measured by enzyme-linked immunosorbent assay (ELISA). Besides, the levels of apoptosis-related proteins (p-53, Bax, and B-cell lymphoma (Bcl)-2) and mitogen-activated protein kinase (MAPK) pathway (phosphorylation-extracellular signalregulated kinase (p-ERK), p-c-Jun N-terminal kinase (JNK), and p-p38) were measured. Results showed that administration of fucoidan significantly reduced the neurological deficits and infarct volume compared to the IRI+S group in a dose-dependent manner. Also, fucoidan statistically decreased the levels of inflammation-associated cytokines, and oxidative stress-related proteins, inhibited apoptosis, and suppressed the MAPK pathway. So, Fucoidan plays a protective role in cerebral IRI might be by inhibition of MAPK pathway.

  8. Intracellular signaling via ERK/MAPK completes the pathway for tubulogenic fibronectin in MDCK cells.

    Science.gov (United States)

    Liu, Zhao; Greco, Andres J; Hellman, Nathan E; Spector, June; Robinson, Jonathan; Tang, Oliver T; Lipschutz, Joshua H

    2007-02-16

    A classic in vitro model of branching morphogenesis utilizes the Madin-Darby canine kidney (MDCK) cell line. MDCK Strain II cells form hollow monoclonal cysts in a three-dimensional collagen matrix over the course of 10 days and tubulate in response to hepatocyte growth factor (HGF). We and our colleagues previously showed that activation of the extracellular-signal regulated kinase (ERK, aka MAPK) pathway is necessary and sufficient to induce tubulogenesis in MDCK cells. We also showed in a microarray study that one of the genes upregulated by HGF was the known tubulogene fibronectin. Given that HGF activates a multitude of signaling pathways, including ERK/MAPK, to test the intracellular regulatory pathway, we used two distinct inhibitors of ERK activation (U0126 and PD098059). Following induction of MDCK Type II cells with HGF, tubulogenic fibronectin mRNA was upregulated fourfold by real-time PCR, and minimal or no change in fibronectin expression was seen when HGF was added with either U0126 or PD098059. We confirmed these results using an MDCK cell line inducible for Raf, which is upstream of ERK. Following activation of Raf, fibronectin mRNA and protein expression were increased to a similar degree as was seen following HGF induction. Furthermore, MDCK Strain I cells, which originate from collecting ducts and have constitutively active ERK, spontaneously initiate tubulogenesis. We show here that MDCK Strain I cells have high levels of fibronectin mRNA and protein compared to MDCK Strain II cells. When U0126 and PD098059 were added to MDCK Strain I cells, fibronectin mRNA, and protein levels were decreased to levels seen in MDCK Strain II cells. These data allow us to complete what we believe is the first description of a tubulogenic pathway from receptor/ligand (HGF/CMET), through an intracellular signaling pathway (ERK/MAPK), to transcription and, finally, secretion of a critical tubuloprotein (fibronectin).

  9. Conservation of protein abundance patterns reveals the regulatory architecture of the EGFR-MAPK pathway.

    Science.gov (United States)

    Shi, Tujin; Niepel, Mario; McDermott, Jason E; Gao, Yuqian; Nicora, Carrie D; Chrisler, William B; Markillie, Lye M; Petyuk, Vladislav A; Smith, Richard D; Rodland, Karin D; Sorger, Peter K; Qian, Wei-Jun; Wiley, H Steven

    2016-07-12

    Various genetic mutations associated with cancer are known to alter cell signaling, but it is not clear whether they dysregulate signaling pathways by altering the abundance of pathway proteins. Using a combination of RNA sequencing and ultrasensitive targeted proteomics, we defined the primary components-16 core proteins and 10 feedback regulators-of the epidermal growth factor receptor (EGFR)-mitogen-activated protein kinase (MAPK) pathway in normal human mammary epithelial cells and then quantified their absolute abundance across a panel of normal and breast cancer cell lines as well as fibroblasts. We found that core pathway proteins were present at very similar concentrations across all cell types, with a variance similar to that of proteins previously shown to display conserved abundances across species. In contrast, EGFR and transcriptionally controlled feedback regulators were present at highly variable concentrations. The absolute abundance of most core proteins was between 50,000 and 70,000 copies per cell, but the adaptors SOS1, SOS2, and GAB1 were found at far lower amounts (2000 to 5000 copies per cell). MAPK signaling showed saturation in all cells between 3000 and 10,000 occupied EGFRs, consistent with the idea that adaptors limit signaling. Our results suggest that the relative stoichiometry of core MAPK pathway proteins is very similar across different cell types, with cell-specific differences mostly restricted to variable amounts of feedback regulators and receptors. The low abundance of adaptors relative to EGFR could be responsible for previous observations that only a fraction of total cell surface EGFR is capable of rapid endocytosis, high-affinity binding, and mitogenic signaling.

  10. Effects of MAPK and PI3K Pathways on PD-L1 Expression in Melanoma

    Science.gov (United States)

    Atefi, Mohammad; Avramis, Earl; Lassen, Amanda; Wong, Deborah; Robert, Lidia; Foulad, David; Cerniglia, Michael; Titz, Bjoern; Chodon, Thinle; Graeber, Thomas G.; Comin-Anduix, Begonya; Ribas, Antoni

    2014-01-01

    Purpose PD-L1 is the main ligand for the immune inhibitory receptor PD-1. This ligand is frequently expressed by melanoma cells. In this study we investigated whether PD-L1 expression is controlled by melanoma driver mutations and modified by oncogenic signaling inhibition. Experimental Design Expression of PD-L1 was investigated in a panel of 51 melanoma cell lines containing different oncogenic mutations, including cell lines with innate and acquired resistance to BRAF inhibitors. The effects of targeted therapy drugs on expression of PD-L1 by melanoma cells were investigated. Results No association was found between the level of PD-L1 expression and mutations in BRAF, NRAS, PTEN or amplification of AKT. Resistance to vemurafenib due to the activation of alternative signaling pathways was accompanied with the induction of PD-L1 expression, while the resistance due to the reactivation of the MAPK pathway had no effect on PD-L1 expression. In melanoma cell lines the effects of BRAF, MEK and PI3K inhibitors on expression of PD-L1 were variable from reduction to induction, particularly in the presence of INFγ. In PD-L1-exposed lymphocytes, vemurafenib paradoxically restored activity of the MAPK pathway and increased the secretion of cytokines. Conclusions In melanoma cell lines, including BRAF inhibitor-resistant cells, PD-L1 expression is variably regulated by oncogenic signaling pathways. PD-L1-exposed lymphocytes decrease MAPK signaling, which is corrected by exposure to vemurafenib, providing potential benefits of combining this drug with immunotherapies. PMID:24812408

  11. Acidic environment leads to ROS-induced MAPK signaling in cancer cells.

    Directory of Open Access Journals (Sweden)

    Anne Riemann

    Full Text Available Tumor micromilieu often shows pronounced acidosis forcing cells to adapt their phenotype towards enhanced tumorigenesis induced by altered cellular signalling and transcriptional regulation. In the presents study mechanisms and potential consequences of the crosstalk between extra- and intracellular pH (pH(e, pH(i and mitogen-activated-protein-kinases (ERK1/2, p38 was analyzed. Data were obtained mainly in AT1 R-3327 prostate carcinoma cells, but the principle importance was confirmed in 5 other cell types. Extracellular acidosis leads to a rapid and sustained decrease of pH(i in parallel to p38 phosphorylation in all cell types and to ERK1/2 phosphorylation in 3 of 6 cell types. Furthermore, p38 phosphorylation was elicited by sole intracellular lactacidosis at normal pH(e. Inhibition of ERK1/2 phosphorylation during acidosis led to necrotic cell death. No evidence for the involvement of the kinases c-SRC, PKC, PKA, PI3K or EGFR nor changes in cell volume in acidosis-induced MAPK activation was obtained. However, our data reveal that acidosis enhances the formation of reactive oxygen species (ROS, probably originating from mitochondria, which subsequently trigger MAPK phosphorylation. Scavenging of ROS prevented acidosis-induced MAPK phosphorylation whereas addition of H(2O(2 enhanced it. Finally, acidosis increased phosphorylation of the transcription factor CREB via p38, leading to increased transcriptional activity of a CRE-reporter even 24 h after switching the cells back to a normal environmental milieu. Thus, an acidic tumor microenvironment can induce a longer lasting p38-CREB-medited change in the transcriptional program, which may maintain the altered phenotype even when the cells leave the tumor environment.

  12. Isorhamnetin prevent endothelial cell injuries from oxidized LDL via activation of p38MAPK.

    Science.gov (United States)

    Bao, Meihua; Lou, Yijia

    2006-10-10

    The present investigation was undertaken to determine the protective effects of isorhamnetin on endothelial cell line EA.hy926 injuries induced by oxidized low-density lipoprotein (ox-LDL) and to uncover some of the underlying mechanisms of these effects. Indices such as cell viability, lactate dehydrogenase (LDH), and nitric oxide (NO) release were measured to evaluate the protective effects of isorhamnetin. 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity, superoxide dismutase (SOD), superoxide and reactive oxygen species (ROS) generation were also detected to evaluate the antioxidant effects of isorhamnetin. Reverse Transcription-Polymerase Chain Reaction (RT-PCR) was used to confirm the expression of endothelial nitric oxide synthase (eNOS) mRNA and lectin-like ox-LDL receptor-1 mRNA. Western blotting was used to evaluate the protein expression of this receptor and eNOS, as well as p38-mitogen-activated protein kinase (p38MAPK) phosphorylation and NF-kappaB p65 translocation. As a result, cell viability decreased significantly (Pisorhamnetin resulted in remarkable increase of cell viability (PIsorhamnetin pretreatment inhibited the ox-LDL-induced downregulation of eNOS, upregulation of lectin-like ox-LDL receptor-1, phosphorylation of the p38MAPK and translocation of NF-kappaB. Moreover, isorhamnetin exhibited strong antioxidant activity, which was shown by its inhibition effects on ox-LDL-induced superoxide, ROS overproduction and significant SOD reduction. The data indicated the protective effects of isorhamnetin on endothelial cell line EA.hy926 from ox-LDL-induced cell injuries. These effects were obtained via inhibition of lectin-like ox-LDL receptor-1 upregulation, interference of ox-LDL-mediated intracellular signaling pathway (p38MAPK activation, NF-kappaB nuclear translocation, eNOS expression) and the antioxidant activity of isorhamnetin.

  13. [Arnold-Chiari malformation in Noonan syndrome and other syndromes of the RAS/MAPK pathway].

    Science.gov (United States)

    Ejarque, Ismael; Millán-Salvador, José M; Oltra, Silvestre; Pesudo-Martínez, José V; Beneyto, Magdalena; Pérez-Aytés, Antonio

    2015-05-01

    Introduccion. El sindrome de Noonan (SN) y otros sindromes con fenotipo similar, como LEOPARD, cardiofaciocutaneo, Costello y Legius, estan asociados a mutaciones en genes incluidos en la via RAS/MAPK (rasopatias), una importante via de señalizacion relacionada con la proliferacion celular. El descenso de las amigdalas cerebelares dentro del canal medular cervical, conocido como malformacion de Arnold-Chiari (MAC), se ha descrito en pacientes afectos de SN, lo que ha llevado a sugerir que la MAC podria formar parte del espectro fenotipico del SN. Presentamos dos casos con SN y MAC. Casos clinicos. Caso 1: mujer de 29 años con fenotipo de Noonan. Fue intervenida a los 9 años de estenosis valvular pulmonar. A los 27 años, presento MAC sintomatica que preciso descompresion quirurgica. Presentaba mutacion c.922A>G (N308D) en el gen PTPN perteneciente a la via RAS/MAPK. Caso 2: niña de 10 años con fenotipo de Noonan y MAC asintomatica detectada en resonancia magnetica cerebral. Era portadora de la mutacion c.923A>G (N308S) en el gen PTPN11. Conclusiones. Hemos encontrado en la bibliografia seis pacientes con esta asociacion, cuatro con fenotipo Noonan y dos con LEOPARD. Nuestros dos pacientes aportan evidencia suplementaria a la hipotesis de que la MAC formaria parte del espectro fenotipico del SN. El escaso numero de pacientes publicados con esta asociacion no permite extraer recomendaciones sobre el momento y la frecuencia de estudio de neuroimagen; no obstante, una exploracion neurologica cuidadosa deberia incluirse en la guia anticipatoria de salud en los sindromes de la via RAS/MAPK.

  14. Extracellular ATP activates MAPK and ROS signaling during injury response in the fungus Trichoderma atroviride

    Directory of Open Access Journals (Sweden)

    Elizabeth eMedina-Castellanos

    2014-11-01

    Full Text Available The response to mechanical damage is crucial for the survival of multicellular organisms, enabling their adaptation to hostile environments. Trichoderma atroviride, a filamentous fungus of great importance in the biological control of plant diseases, responds to mechanical damage by activating regenerative processes and asexual reproduction (conidiation. During this response, reactive oxygen species (ROS are produced by the NADPH oxidase (Nox1/NoxR complex. To understand the underlying early signaling events, we evaluated molecules such as extracellular ATP (eATP and Ca2+ that are known to trigger wound-induced responses in plants and animals. Concretely, we investigated the activation of mitogen-activated protein kinase (MAPK pathways by eATP, Ca2+ and ROS. Indeed, application of exogenous ATP and Ca2+ triggered conidiation. Furthermore, eATP promoted the Nox1-dependent production of ROS and activated a MAPK pathway. Mutants in the MAPK-encoding genes tmk1 and tmk3 were affected in wound-induced conidiation, and phosphorylation of both Tmk1 and Tmk3 was triggered by eATP. We conclude that in this fungus, eATP acts as a damage-associated molecular pattern (DAMP. Our data indicate the existence of an eATP receptor and suggest that in fungi, eATP triggers pathways that converge to regulate asexual reproduction genes that are required for injury-induced conidiation. By contrast, Ca2+ is more likely to act as a downstream second messenger. The early steps of mechanical damage response in T. atroviride share conserved elements with those known from plants and animals.

  15. Epidermal hyperplasia induced by Raf-MAPK signaling requires Stat3 activation.

    Science.gov (United States)

    Tarutani, Masahito; Nakajima, Kimiko; Takaishi, Mikiro; Ohko, Kentaro; Sano, Shigetoshi

    2013-11-01

    Raf is one of the downstream effectors of Ras GTPases, and plays a key role in regulating cell proliferation and differentiation through the activation of MAPK. We have previously demonstrated that temporal induction of Raf in the epidermis of K14-Raf:ER transgenic mice results in epidermal hyperplasia resembling squamous cell carcinoma and psoriasis. It has been demonstrated that epidermal Stat3 activation is required for psoriasis development, since keratinocyte-specific Stat3 activation in a mouse model elicits a psoriasis-like phenotype, which is reversed by inhibition of Stat3 signaling. The aim of this study was whether Stat3 signaling is involved in Raf-MAPK-dependent epidermal hyperplasia. K14-Raf:ER transgenic mice, in which the 4-hydroxytamoxifen (4OHT)-responsive mutant estrogen receptor ligand binding domain-Raf fusion gene is expressed under control of the keratin 14 promoter, were mated with epidermis-specific Stat3 null mice (K5-Cre.Stat3(flox/flox)). K5-Cre.Stat3(flox/flox) mice were used to define the impact of Stat3 deficiency on Raf-induced epidermal hyperplasia. Over-expression of Raf by 4OHT treatment in K14-Raf:ER;K5-Cre.Stat3(flox/flox) mice greatly attenuated the epidermal hyperplasia and dermal cell infiltrates compared with K14-Raf:ER;K5-Cre.Stat3(flox/WT) mice. Also, up-regulation of psoriasis-associated cytokine profiles, including VEGF, was inhibited in the skin from K14-Raf:ER;K5-Cre.Stat3(flox/flox) mice following 4OHT treatment. These results clearly indicate that Raf-MAPK-dependent psoriatic-like epidermal hyperplasia requires Stat3 signaling in keratinocytes. Copyright © 2013 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

  16. Morin Attenuates Ovalbumin-Induced Airway Inflammation by Modulating Oxidative Stress-Responsive MAPK Signaling

    Directory of Open Access Journals (Sweden)

    Yuan Ma

    2016-01-01

    Full Text Available Asthma is one of the most common inflammatory diseases characterized by airway hyperresponsiveness, inflammation, and remodeling. Morin, an active ingredient obtained from Moraceae plants, has been demonstrated to have promising anti-inflammatory activities in a range of disorders. However, its impacts on pulmonary diseases, particularly on asthma, have not been clarified. This study was designed to investigate whether morin alleviates airway inflammation in chronic asthma with an emphasis on oxidative stress modulation. In vivo, ovalbumin- (OVA- sensitized mice were administered with morin or dexamethasone before challenge. Bronchoalveolar lavage fluid (BALF and lung tissues were obtained to perform cell counts, histological analysis, and enzyme-linked immunosorbent assay. In vitro, human bronchial epithelial cells (BECs were challenged by tumor necrosis factor alpha (TNF-α. The supernatant was collected for the detection of the proinflammatory proteins, and the cells were collected for reactive oxygen species (ROS/mitogen-activated protein kinase (MAPK evaluations. Severe inflammatory responses and remodeling were observed in the airways of the OVA-sensitized mice. Treatment with morin dramatically attenuated the extensive trafficking of inflammatory cells into the BALF and inhibited their infiltration around the respiratory tracts and vessels. Morin administration also significantly suppressed goblet cell hyperplasia and collagen deposition/fibrosis and dose-dependently inhibited the OVA-induced increases in IgE, TNF-α, interleukin- (IL- 4, IL-13, matrix metalloproteinase-9, and malondialdehyde. In human BECs challenged by TNF-α, the levels of proteins such as eotaxin-1, monocyte chemoattractant protein-1, IL-8 and intercellular adhesion molecule-1, were consistently significantly decreased by morin. Western blotting and the 2′,7′-dichlorofluorescein assay revealed that the increases in intracellular ROS and MAPK phosphorylation were

  17. Activated p38 MAPK in Peripheral Blood Monocytes of Steroid Resistant Asthmatics.

    Directory of Open Access Journals (Sweden)

    Ling-Bo Li

    Full Text Available Steroid resistance is a significant problem in management of chronic inflammatory diseases, including asthma. Accessible biomarkers are needed to identify steroid resistant patients to optimize their treatment. This study examined corticosteroid resistance in severe asthma. 24 asthmatics with forced expiratory volume in one second of less then 80% predicted were classified as steroid resistant or steroid sensitive based on changes in their lung function following a week of treatment with oral prednisone. Heparinised blood was collected from patients prior to oral prednisone administration. Phosphorylated mitogen activated kinases (MAPK (extracellular regulated kinase (ERK, p38 and jun kinase (JNK were analyzed in whole blood samples using flow cytometry. Activation of phospho-p38 MAPK and phospho-mitogen- and stress-activated protein kinase 1 (MSK1 in asthmatics' peripheral blood mononuclear cells (PBMC were confirmed by Western blot. Dexamethasone suppression of the LPS-induced IL-8 mRNA production by steroid resistant asthmatics PBMC in the presence of p38 and ERK inhibitors was evaluated by real time PCR. Flow cytometry analysis identified significantly stronger p38 phosphorylation in CD14+ monocytes from steroid resistant than steroid sensitive asthmatics (p = 0.014, whereas no difference was found in phosphorylation of ERK or JNK in CD14+ cells from these two groups of asthmatics. No difference in phosphorylated p38, ERK, JNK was detected in CD4+, CD8+ T cells, B cells and NK cells from steroid resistant vs. steroid sensitive asthmatics. P38 MAPK pathway activation was confirmed by Western blot, as significantly higher phospho-p38 and phospho-MSK1 levels were detected in the PBMC lysates from steroid resistant asthmatics. P38 inhibitor significantly enhanced DEX suppression of LPS-induced IL-8 mRNA by PBMC of steroid resistant asthmatics. This is the first report demonstrating selective p38 MAPK pathway activation in blood monocytes of

  18. Synaptic plasticity, AMPA-R trafficking, and Ras-MAPK signaling

    Institute of Scientific and Technical Information of China (English)

    Yun GU; Ruth L STORNETTA

    2007-01-01

    Synaptic modification of transmission is a general phenomenon expressed at al-most every excitatory synapse in the mammalian brain. Over the last three decades,much has been discovered about the cellular, synaptic, molecular, and signalingmechanisms responsible for controlling synaptic transmission and plasticity. Here,we present a brief review of these mechanisms with emphasis on the currentunderstanding of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid recep-tor (AMPA-R) trafficking and Ras-mitogen-activated protein kinase (MAPK)signaling events involved in controlling synaptic transmission.

  19. Role of the fission yeast cell integrity MAPK pathway in response to glucose limitation

    Directory of Open Access Journals (Sweden)

    Madrid Marisa

    2013-02-01

    Full Text Available Abstract Background Glucose is a signaling molecule which regulates multiple events in eukaryotic organisms and the most preferred carbon source in the fission yeast Schizosaccharomyces pombe. The ability of this yeast to grow in the absence of glucose becomes strongly limited due to lack of enzymes of the glyoxylate cycle that support diauxic growth. The stress-activated protein kinase (SAPK pathway and its effectors, Sty1 MAPK and transcription factor Atf1, play a critical role in the adaptation of fission yeast to grow on alternative non-fermentable carbon sources by inducing the expression of fbp1+ gene, coding for the gluconeogenic enzyme fructose-1,6-bisphosphatase. The cell integrity Pmk1 pathway is another MAPK cascade that regulates various processes in fission yeast, including cell wall construction, cytokinesis, and ionic homeostasis. Pmk1 pathway also becomes strongly activated in response to glucose deprivation but its role during glucose exhaustion and ensuing adaptation to respiratory metabolism is currently unknown. Results We found that Pmk1 activation in the absence of glucose takes place only after complete depletion of this carbon source and that such activation is not related to an endogenous oxidative stress. Notably, Pmk1 MAPK activation relies on de novo protein synthesis, is independent on known upstream activators of the pathway like Rho2 GTPase, and involves PKC ortholog Pck2. Also, the Glucose/cAMP pathway is required operative for full activation of the Pmk1 signaling cascade. Mutants lacking Pmk1 displayed a partial growth defect in respiratory media which was not observed in the presence of glucose. This phenotype was accompanied by a decreased and delayed expression of transcription factor Atf1 and target genes fbp1+ and pyp2+. Intriguingly, the kinetics of Sty1 activation in Pmk1-less cells was clearly altered during growth adaptation to non-fermentable carbon sources. Conclusions Unknown upstream elements

  20. Socs36E Controls Niche Competition by Repressing MAPK Signaling in the Drosophila Testis.

    Directory of Open Access Journals (Sweden)

    Marc Amoyel

    2016-01-01

    Full Text Available The Drosophila testis is a well-established system for studying stem cell self-renewal and competition. In this tissue, the niche supports two stem cell populations, germ line stem cells (GSCs, which give rise to sperm, and somatic stem cells called cyst stem cells (CySCs, which support GSCs and their descendants. It has been established that CySCs compete with each other and with GSCs for niche access, and mutations have been identified that confer increased competitiveness to CySCs, resulting in the mutant stem cell and its descendants outcompeting wild type resident stem cells. Socs36E, which encodes a negative feedback inhibitor of the JAK/STAT pathway, was the first identified regulator of niche competition. The competitive behavior of Socs36E mutant CySCs was attributed to increased JAK/STAT signaling. Here we show that competitive behavior of Socs36E mutant CySCs is due in large part to unbridled Mitogen-Activated Protein Kinase (MAPK signaling. In Socs36E mutant clones, MAPK activity is elevated. Furthermore, we find that clonal upregulation of MAPK in CySCs leads to their outcompetition of wild type CySCs and of GSCs, recapitulating the Socs36E mutant phenotype. Indeed, when MAPK activity is removed from Socs36E mutant clones, they lose their competitiveness but maintain self-renewal, presumably due to increased JAK/STAT signaling in these cells. Consistently, loss of JAK/STAT activity in Socs36E mutant clones severely impairs their self-renewal. Thus, our results enable the genetic separation of two essential processes that occur in stem cells. While some niche signals specify the intrinsic property of self-renewal, which is absolutely required in all stem cells for niche residence, additional signals control the ability of stem cells to compete with their neighbors. Socs36E is node through which these processes are linked, demonstrating that negative feedback inhibition integrates multiple aspects of stem cell behavior.

  1. p38(MAPK)/p53-Mediated Bax induction contributes to neurons degeneration in rotenone-induced cellular and rat models of Parkinson's disease.

    Science.gov (United States)

    Wu, Feng; Wang, Zhong; Gu, Jin-Hua; Ge, Jian-Bin; Liang, Zhong-Qin; Qin, Zheng-Hong

    2013-09-01

    Rotenone is an environmental neurotoxin that induces degeneration of dopaminergic (DA) neurons in substantia nigra pars compacta (SNpc), which ultimately results in parkinsonism, but the molecular mechanisms of selective degeneration of nigral DA neurons are not fully understood. In the present study, we investigated the induction of p38(MAPK)/p53 and Bax in SNpc of Lewis rats after chronic treatment with rotenone and the contribution of Bax to rotenone-induced apoptotic commitment of differentiated PC12 cells. Lewis rats were subcutaneously treated with rotenone (1.5mg/kg) twice a day for 50days and the loss of tyrosine hydroxylase (THase), motor function impairment, and expression of p38(MAPK), P-p38(MAPK), p53, and Bax were assessed. After differentiated PC cells were treated with rotenone (500nM) for 6-36h, protein levels of p38(MAPK) and P-p38(MAPK), p53 nuclear translocation, Bax induction and cell death were measured. The results showed that rotenone administration significantly reduced motor activity and caused a loss of THase immunoreactivity in SNpc of Lewis rats. The degeneration of nigral DA neurons was accompanied by the increases in p38(MAPK), P-p38(MAPK), p53, and Bax protein levels. In cultured PC12 cells, rotenone also induced an upregulation of p38(MAPK), P-p38(MAPK), p53 and Bax. Pharmacological inhibition of p38(MAPK) with SB203580 (25μM) blunted rotenone-induced cell apoptosis. Treatment with SB203580 prevented the p53 nuclear translocation and upregulation of Bax. Inhibition of p53 with pifthrin-alpha or Bax with siRNAs significantly reduced rotenone-induced Bax induction and apoptotic cell death. These results suggest that the p38(MAPK)/p53-dependent induction of Bax contributes to rotenone's neurotoxicity in PD models.

  2. Cloning and Bioinformatics Analysis of MAPK13 Gene in Lanzhou Fat-tailed Sheep%兰州大尾羊MAPK13基因cDNA克隆及生物信息学分析

    Institute of Scientific and Technical Information of China (English)

    金方园; 徐红伟; 达小强; 臧荣鑫; 柏家林; 曹忻; 蔡勇; 冯玉兰; 杨具田

    2014-01-01

    The objective of this study is to reveal the biological function of mitogen-activated protein kinases(MAPK)gene in sheep and provide theoretical data for application by cloning MAPK and analyzing its sequence as well as genetic features. The specific primers on the CDS template of MAPK13 gene were designed and then the MAPK13 gene sequence of Lanzhou fat-tail sheep was cloned using RACE and RT-PCR technology. After that, its genetic characteristics were analyzed by bioinformatics methods. The 1 397 bp full-length cDNA of MAPK13 of Lanzhou fat-tailed sheep was obtained, in which the length of CDS is 1 102 bp. It encodes 367 amino acids in total. It was predicted that the molecular weight of MAPK13 protein of Lanzhou fat-tail sheep is 42.29 kD and its theoretical isoelectric point is 8.82. It is a non-transmembrane hydrophobin with no signal peptide, which locates mostly in cytoplasm by the result of subcellular level prediction and does not belong to the secretory protein. Its amino acid sequence contains 19 phosphorylation sites, 3 glycosylation sites, 3 phosphorylation domains, 4 other domains and 1 LCR domain. Also, its secondary structure is mainly randomly curly. Nucleotide sequence analysis revealed that the gene nucleotide sequence of Lanzhou fat-tail sheep has some different from the known sheep MAPK13 mRNA(Genbank No.:NM_001139455.1). Base transition occur in the 852th site is(C vs. A), respectively, the corresponding amino acids in the 265th is(S vs. T). Base transition occur in the 951th site is(T vs. G), but the corresponding amino acids do not change.The phylogenetic tree indicates that Lanzhou fat-tail sheep is close to bos. The fact that the structure of MAPK13 of Lanzhou fat-tail sheep highly similar with other species indicates that the gene is highly conservative. The S_TKc domain in the sequence can transfer the gamma phosphoryl of ATP to the serine/threonine residues of protein, leading to function imbalance and expression change in a series of

  3. RAS/MAPK Activation Drives Resistance to Smo Inhibition, Metastasis, and Tumor Evolution in Shh Pathway-Dependent Tumors.

    Science.gov (United States)

    Zhao, Xuesong; Ponomaryov, Tatyana; Ornell, Kimberly J; Zhou, Pengcheng; Dabral, Sukriti K; Pak, Ekaterina; Li, Wei; Atwood, Scott X; Whitson, Ramon J; Chang, Anne Lynn S; Li, Jiang; Oro, Anthony E; Chan, Jennifer A; Kelleher, Joseph F; Segal, Rosalind A

    2015-09-01

    Aberrant Shh signaling promotes tumor growth in diverse cancers. The importance of Shh signaling is particularly evident in medulloblastoma and basal cell carcinoma (BCC), where inhibitors targeting the Shh pathway component Smoothened (Smo) show great therapeutic promise. However, the emergence of drug resistance limits long-term efficacy, and the mechanisms of resistance remain poorly understood. Using new medulloblastoma models, we identify two distinct paradigms of resistance to Smo inhibition. Sufu mutations lead to maintenance of the Shh pathway in the presence of Smo inhibitors. Alternatively activation of the RAS-MAPK pathway circumvents Shh pathway dependency, drives tumor growth, and enhances metastatic behavior. Strikingly, in BCC patients treated with Smo inhibitor, squamous cell cancers with RAS/MAPK activation emerged from the antecedent BCC tumors. Together, these findings reveal a critical role of the RAS-MAPK pathway in drug resistance and tumor evolution of Shh pathway-dependent tumors.

  4. Euclidean distance harmonic method for establishing theoretical MAPK/Erk signaling pathway in treated breast cancer line MCF-7

    Institute of Scientific and Technical Information of China (English)

    ZHANG Hong-xin; LU Ying-hua; ZHANG Jin-ling

    2007-01-01

    Hierarchical clustering algorithms, such as Pearson's correlation, Euclidean distance, Euclidean distance harmonic,Spearman rank correlation, Kendall's tau, and City-block distance, were used to find the best way to establish theoretical MAPK/Erk signaling pathway on the basis of breast cancer line MCF-7 gene expressions. The algorithm consttucts a hierarchy from top to bottom on the basis of a self-organizing tree. It dynamically finds the number of clusters at each level. It was found that only Euclidean distance harmonic is fit for the analysis of the cascade composed from a RAF1 (c-Raf), a MKNK1, a MAPKK (MEK1/2) to MAPK (Erk) in breast cancer line MCF-7. The result is consistent with the biological experimental MAP/Erk signaling pathway, and the theoretical MAPK/Erk signaling pathway on breast cancer line MCF-7 is set up.

  5. Differential Roles of Grb2 and AP-2 in p38 MAPK- and EGF-Induced EGFR Internalization

    DEFF Research Database (Denmark)

    Grandal, Michael V; Grøvdal, Lene M; Henriksen, Lasse;

    2012-01-01

    also can induce EGFR endocytosis. This endocytosis lacks many of the characteristics of ligand-induced EGFR endocytosis. We compared the two types of endocytosis with regard to the requirements for proteins in the internalization machinery. Both types of endocytosis require clathrin, but while...... epidermal growth factor (EGF)-induced EGFR internalization also required Grb2, p38 MAPK-induced internalization did not. Interestingly, AP-2 knock down blocked p38 MAPK-induced EGFR internalization, but only mildly affected EGF-induced internalization. In line with this, simultaneously mutating two AP-2...... interaction sites in EGFR affected p38 MAPK-induced internalization much more than EGF-induced EGFR internalization. Thus, it seems that EGFR in the two situations uses different sets of internalization mechanisms....

  6. The high-osmolarity glycerol (HOG) and cell wall integrity (CWI) signalling pathways interplay: a yeast dialogue between MAPK routes.

    Science.gov (United States)

    Rodríguez-Peña, Jose Manuel; García, Raúl; Nombela, César; Arroyo, Javier

    2010-08-01

    Two mitogen-activated protein kinase (MAPK) pathways, viz. the high-osmolarity glycerol (HOG) and the cell wall integrity (CWI) pathways, regulate stress responses in the yeast Saccharomyces cerevisiae. Whereas the former is mainly involved in adaptation of yeast cells to hyperosmotic stress, the latter is activated under conditions leading to cell wall instability. Although MAPK signalling specificity can be conceived as requiring insulation of the different pathways, it is also becoming clear that the two pathways do not compete with each other but can be positively coordinated to regulate many stress responses. This review highlights our current knowledge about the collaboration between these two MAPK pathways to counteract different kinds of environmental stress.

  7. CCR7 regulates cell migration and invasion through MAPKs in metastatic squamous cell carcinoma of head and neck.

    Science.gov (United States)

    Liu, Fa-Yu; Safdar, Jawad; Li, Zhen-Ning; Fang, Qi-Gen; Zhang, Xu; Xu, Zhong-Fei; Sun, Chang-Fu

    2014-12-01

    Migration and invasion of tumor cells are essential prerequisites for the formation of metastasis in malignant diseases. Previously, we have reported that CC chemokine receptor 7 (CCR7) regulates the mobility of squamous cell carcinoma of head and neck (SCCHN) cells through several pathways, such as integrin and cdc42. In this study, we investigated the connection between CCR7 and mitogen-activated protein kinase (MAPK) family members, and their influence on cell invasion and migration in metastatic SCCHN cells. Western blotting, immunostaining and fluorescence microcopy were used to detect the protein expression and distribution of MAPKs, and the Migration assay, Matrigel invasion assay and wound-healing assay to detect the role of MAPKs in CCR7 regulating cell mobility. To analyze the correlation between CCR7 and MAPK activity and clinicopathological factors immunohistochemical staining was emplyed. The results showed stimulation of CCL19 and the activation of CCR7 could induce ERK1/2 and JNK phosphorylation, while it had no efect on p38. After activation, ERK1/2 and JNK promoted E-cadherin low expression and Vimentin high expression. The MAPK pathway not only mediated CCR7 induced cell migration, but also mediated invasion speed. The immunohistochemistry results showed that CCR7 was correlated with the phosphorylation of ERK1/2 and JNK in SCCHN, and these molecules were all associated with lymph node metastasis. Therefore, our study demonstrates that MAPK members (ERK1/2 and JNK) play a key role in CCR7 regulating SCCHN metastasis.

  8. The MAPK pathway signals telomerase modulation in response to isothiocyanate-induced DNA damage of human liver cancer cells.

    Directory of Open Access Journals (Sweden)

    Evelyn Lamy

    Full Text Available 4-methylthiobutyl isothiocyanate (MTBITC, an aliphatic, sulphuric compound from Brassica vegetables, possesses in vitro and in vivo antitumor activity. Recently we demonstrated the potent growth inhibitory potential of the DNA damaging agent MTBITC in human liver cancer cells. Here we now show that MTBITC down regulates telomerase which sensitizes cells to apoptosis induction. This is mediated by MAPK activation but independent from production of reactive oxygen species (ROS. Within one hour, MTBITC induced DNA damage in cancer cells correlating to a transient increase in hTERT mRNA expression which then turned into telomerase suppression, evident at mRNA as well as enzyme activity level. To clarify the role of MAPK for telomerase regulation, liver cancer cells were pre-treated with MAPK-specific inhibitors prior to MTBITC exposure. This clearly showed that transient elevation of hTERT mRNA expression was predominantly mediated by the MAPK family member JNK. In contrast, activated ERK1/2 and P38, but not JNK, signalled to telomerase abrogation and consequent apoptosis induction. DNA damage by MTBITC was also strongly abolished by MAPK inhibition. Oxidative stress, as analysed by DCF fluorescence assay, electron spin resonance spectroscopy and formation of 4-hydroxynonenal was found as not relevant for this process. Furthermore, N-acetylcysteine pre-treatment did not impact MTBITC-induced telomerase suppression or depolarization of the mitochondrial membrane potential as marker for apoptosis. Our data therefore imply that upon DNA damage by MTBITC, MAPK are essential for telomerase regulation and consequent growth impairment in liver tumor cells and this detail probably plays an important role in understanding the potential chemotherapeutic efficacy of ITC.

  9. Local overexpression of Su(H)-MAPK variants affects Notch target gene expression and adult phenotypes in Drosophila.

    Science.gov (United States)

    Auer, Jasmin S; Nagel, Anja C; Schulz, Adriana; Wahl, Vanessa; Preiss, Anette

    2015-12-01

    In Drosophila, Notch and EGFR signalling pathways are closely intertwined. Their relationship is mostly antagonistic, and may in part be based on the phosphorylation of the Notch signal transducer Suppressor of Hairless [Su(H)] by MAPK. Su(H) is a transcription factor that together with several cofactors regulates the expression of Notch target genes. Here we address the consequences of a local induction of three Su(H) variants on Notch target gene expression. To this end, wild-type Su(H), a phospho-deficient Su(H) (MAPK-) (ko) and a phospho-mimetic Su(H) (MAPK-ac) isoform were overexpressed in the central domain of the wing anlagen. The expression of the Notch target genes cut, wingless, E(spl)m8-HLH and vestigial, was monitored. For the latter two, reporter genes were used (E(spl)m8-lacZ, vg (BE) -lacZ). In general, Su(H) (MAPK-) (ko) induced a stronger response than wild-type Su(H), whereas the response to Su(H) (MAPK-ac) was very weak. Notch target genes cut, wingless and vg (BE) -lacZ were ectopically activated, whereas E(spl)m8-lacZ was repressed by overexpression of Su(H) proteins. In addition, in epistasis experiments an activated form of the EGF-receptor (DER (act) ) or the MAPK (rl (SEM) ) and individual Su(H) variants were co-overexpressed locally, to compare the resultant phenotypes in adult flies (thorax, wings and eyes) as well as to assay the response of the Notch target gene cut in cell clones.

  10. [OMT inhibited TGF-β1-induced cardiac fibroblast proliferation via down-regulating p38MAPK phosphorylation in vitro].

    Science.gov (United States)

    Xiao, Hai; Xu, Yi-ni; Luo, Hong; Chen, Yan; Zhang, Yan-yan; Tao, Ling; Jiang, Yan; Shen, Xiang-chun

    2015-06-01

    To investigate the inhibitory effects of OMT on TGF-β1-induced CFBs proliferation, and then explore the mechanism. The experiment was randomly divided into 6 groups as following: control group (serum free DMEM), model group (20 μg x L(-1) TGF-β1), OMT low dose group (1.89 x 10(-4) mol x L(-1) + 20 μg x L(-1) TGF-β1), OMT medium dose group (3.78 x 10(-4) mol x L(-1) + 20 μg x L(-1) TGF-β1), OMT high dose group (7.56 x 10(-4) mol x L(-1) + 20 μg x L(-1) TGF-β1), SB203580 group (p38MAPK blocking agent, 1 x 10(-5) mol x L(-1) + 20 μg x L(-1) TGF-β1). Vimentin of CFBs was identified by immunocytochemical methods, α-SMA of myFBs as well. Inhibitory effects of OMT on CFBs proliferation was detected by the MTT assay. Picric acid Sirius red staining was analyzed collagen type I and collagen type III deposition. Western blot was determined the expression of p38MAPK, p-p38MAPK, collagen type I and collagen type III. MTT results showed that OMT significantly inhibited CFBs proliferation induced by TGF-β1 (P OMT could protect against the CFBs proliferation. OMT could significantly decrease the deposition of collagen type I and collagen type III by Western bloting and picric acid Sirius red staining. Western blot results showed that TGF-β1 enhanced p38MAPK phosphorylation, however OMT attenuated the phosphorylation of p38MAPK induced by TGF-β1 (P OMT can inhibit the CFBs proliferation induced by TGF-β1, and its mechanism may be involved in inhibiting p38MAPK phosphorylation.

  11. Explore the variation of MMP3, JNK, p38 MAPKs, and autophagy at the early stage of osteoarthritis.

    Science.gov (United States)

    Shi, Jie; Zhang, Changjie; Yi, Zhongjie; Lan, Chunna

    2016-04-01

    Osteoarthritis is a chronic disease characterized by cartilage degeneration and chondrocyte apoptosis. Mitogen-activated protein kinase (MAPK) signaling pathway plays a key role in regulating OA process. Autophagy has an important effect on the OA process, and it is believed to be regulated by MAPKs. To reveal the mechanism and the effect of JNK and p38 MAPKs on matrix metalloproteinase 3 (MMP3) and autophagy in OA, the study established OA model in rabbits, used the measurement of the Osteoarthritis Research Society International scoring system to evaluate OA model, and conducted general observation, histological observation, and Western blotting of JNK, phosphorylate-JNK (P-JNK), p38, phosphorylate-p38 (P-p38), MMP3, and light-chain 3 (LC3)-II/LC3-I to explore the variation of JNK, p38 MAPKs, and autophagy at the early stage of OA. With OA progressing at the early stage, MMP3, P-p38, and P-JNK were gradually upregulated from the baseline to the peak in study groups when compared with the control group; JNK and p38 variated of turbulence without statistical difference; and LC3-II/LC3-I had a decreasing tendency from the 0- to 15-day group. This study identifies that compromised autophagy may be related to the OA progress and that JNK and p38 MAPKs have positive regulation on MMP3 and negative regulation on autophagy. It also implicates a new therapeutic strategy for OA and other degenerate diseases based on selective MAPK inhibitors, reduction of MMP3, and autophagy.

  12. TNF-α stimulates System A amino acid transport in primary human trophoblast cells mediated by p38 MAPK signaling.

    Science.gov (United States)

    Aye, Irving L M H; Jansson, Thomas; Powell, Theresa L

    2015-10-01

    Maternal obesity and gestational diabetes mellitus (GDM) increase the risk of delivering infants that are large for gestational age with greater adiposity, who are prone to the development of metabolic disease in childhood and beyond. These maternal conditions are also associated with increased levels of the proinflammatory cytokine TNF-α in maternal tissues and the placenta. Recent evidence suggests that changes in placental amino acid transport contribute to altered fetal growth. TNF-α was previously shown to stimulate System A amino acid transport in primary human trophoblasts (PHTs), however the molecular mechanisms remain unknown. In this study, we tested the hypothesis that TNF-α regulates amino acid uptake in cultured PHTs by a mitogen-activated protein kinase (MAPK)-dependent mechanism. Treatment of PHTs with TNF-α significantly increased System A amino acid transport, as well as Erk and p38 MAPK signaling. Pharmacological antagonism of p38, but not Erk MAPK activity, inhibited TNF-α stimulated System A activity. Silencing of p38 MAPK using siRNA transfections prevented TNF-α stimulated System A transport in PHTs. TNF-α significantly increased the protein expression of System A transporters SNAT1 and SNAT2, but did not affect their mRNA expression. The effects of TNF-α on SNAT1 and SNAT2 protein expression were reversed by p38 MAPK siRNA silencing. In conclusion, TNF-α regulates System A activity through increased SNAT1 and SNAT2 transporter protein expression in PHTs. These findings suggest that p38 MAPK may represent a critical mechanistic link between elevated proinflammatory cytokines and increased placental amino acid transport in obese and GDM pregnancies associated with fetal overgrowth.

  13. Mangiferin protect myocardial insults through modulation of MAPK/TGF-β pathways.

    Science.gov (United States)

    Suchal, Kapil; Malik, Salma; Gamad, Nanda; Malhotra, Rajiv Kumar; Goyal, Sameer N; Ojha, Shreesh; Kumari, Santosh; Bhatia, Jagriti; Arya, Dharamvir Singh

    2016-04-05

    Mangiferin, a xanthone glycoside isolated from leaves of Mangifera indica (Anacardiaceae) is known to modulate many biological targets in inflammation and oxidative stress. The present study was designed to investigate whether mangiferin exerts protection against myocardial ischemia-reperfusion (IR) injury and possible role of Mitogen Activated Protein Kinase (MAPKs) and Transforming Growth Factor-β (TGF-β) pathways in its cardioprotection. Male albino Wistar rats were treated with mangiferin (40 mg/kg, i.p.) for 15 days. At the end of the treatment protocol, rats were subjected to IR injury consisting of 45 min ischemia followed by 1h reperfusion. IR-control rats caused significant cardiac dysfunction, increased serum cardiac injury markers, lipid peroxidation and a significant decrease in tissue antioxidants as compared to sham group. Histopathological examination of IR rats revealed myocardial necrosis, edema and infiltration of inflammatory cells. However, pretreatment with mangiferin significantly restored myocardial oxidant-antioxidant status, maintained membrane integrity, and attenuated the levels of proinflammatory cytokines, pro-apoptotic proteins and TGF-β. Furthermore, mangiferin significantly reduced the phosphorylation of p38, and JNK and enhanced phosphorylation of ERK1/2. These results suggest that mangiferin protects against myocardial IR injury by modulating MAPK mediated inflammation and apoptosis.

  14. Cardiotoxin III Inhibits Proliferation and Migration of Oral Cancer Cells through MAPK and MMP Signaling

    Directory of Open Access Journals (Sweden)

    Ching-Yu Yen

    2013-01-01

    Full Text Available Cardiotoxin III (CTXIII, isolated from the snake venom of Formosan cobra Naja naja atra, has previously been found to induce apoptosis in many types of cancer. Early metastasis is typical for the progression of oral cancer. To modulate the cell migration behavior of oral cancer is one of the oral cancer therapies. In this study, the possible modulating effect of CTXIII on oral cancer migration is addressed. In the example of oral squamous carcinoma Ca9-22 cells, the cell viability was decreased by CTXIII treatment in a dose-responsive manner. In wound-healing assay, the cell migration of Ca9-22 cells was attenuated by CTXIII in a dose- and time-responsive manner. After CTXIII treatment, the MMP-2 and MMP-9 protein expressions were downregulated, and the phosphorylation of JNK and p38-MAPK was increased independent of ERK phosphorylation. In conclusion, CTXIII has antiproliferative and -migrating effects on oral cancer cells involving the p38-MAPK and MMP-2/-9 pathways.

  15. p38α MAPK Is Required for Tooth Morphogenesis and Enamel Secretion*

    Science.gov (United States)

    Greenblatt, Matthew B.; Kim, Jung-Min; Oh, Hwanhee; Park, Kwang Hwan; Choo, Min-Kyung; Sano, Yasuyo; Tye, Coralee E.; Skobe, Ziedonis; Davis, Roger J.; Park, Jin Mo; Bei, Marianna; Glimcher, Laurie H.; Shim, Jae-Hyuck

    2015-01-01

    An improved understanding of the molecular pathways that drive tooth morphogenesis and enamel secretion is needed to generate teeth from organ cultures for therapeutic implantation or to determine the pathogenesis of primary disorders of dentition (Abdollah, S., Macias-Silva, M., Tsukazaki, T., Hayashi, H., Attisano, L., and Wrana, J. L. (1997) J. Biol. Chem. 272, 27678–27685). Here we present a novel ectodermal dysplasia phenotype associated with conditional deletion of p38α MAPK in ectodermal appendages using K14-cre mice (p38αK14 mice). These mice display impaired patterning of dental cusps and a profound defect in the production and biomechanical strength of dental enamel because of defects in ameloblast differentiation and activity. In the absence of p38α, expression of amelogenin and β4-integrin in ameloblasts and p21 in the enamel knot was significantly reduced. Mice lacking the MAP2K MKK6, but not mice lacking MAP2K MKK3, also show the enamel defects, implying that MKK6 functions as an upstream kinase of p38α in ectodermal appendages. Lastly, stimulation with BMP2/7 in both explant culture and an ameloblast cell line confirm that p38α functions downstream of BMPs in this context. Thus, BMP-induced activation of the p38α MAPK pathway is critical for the morphogenesis of tooth cusps and the secretion of dental enamel. PMID:25406311

  16. Modulation of age-related NF-kappaB activation by dietary zingerone via MAPK pathway.

    Science.gov (United States)

    Kim, Mi Kyung; Chung, Sang Woon; Kim, Dae Hyun; Kim, Ji Min; Lee, Eun Kyeong; Kim, Ji Young; Ha, Young Mi; Kim, Yun Hee; No, Jae-Kyung; Chung, Hye Sun; Park, Kun-Young; Rhee, Sook Hee; Choi, Jae Sue; Yu, Byung Pal; Yokozawa, Takako; Kim, Young Jin; Chung, Hae Young

    2010-06-01

    Zingerone, a major component found in ginger root, has been known as anti-mutagenic and anti-carcinogenic activities that are often associated with its anti-oxidative and anti-inflammatory activities. In recent studies, we examined molecular mechanism of zingerone treatment on pro-inflammatory NF-kappaB activation via the redox-related NIK/IKK and MAPK pathways. Action mechanism of zingerone on NF-kappaB signaling was investigated in aged rat kidney and endothelial cells. The results showed that zingerone had not only the antioxidant effect by constitutive suppression of ROS, but also anti-inflammatory effects by suppression of nuclear factor (NF)-kappaB activation in aged rat. In addition, zingerone treatment suppressed gene activation of pro-inflammatory enzymes, COX-2 and iNOS, which were upregulated with aging through NF-kappaB activation and IKK/MAPK signaling pathway. These experiments strongly indicate that zingerone treatment exerts a beneficial efficacy by suppressing both oxidative stress and age-related inflammation through the modulation of several key pro-inflammatory genes and transcription factors. Thus, the significance of our findings is that the zingerone treatment may provide some preventive measure against chronic inflammatory conditions that underlie many age-related inflammatory diseases, such as metabolic syndrome, cardiovascular disease, dementia, arthritis, diabetes, osteoprosis, and cancers.

  17. Tangeretin Alleviates Cisplatin-Induced Acute Hepatic Injury in Rats: Targeting MAPKs and Apoptosis.

    Science.gov (United States)

    Omar, Hany A; Mohamed, Wafaa R; Arab, Hany H; Arafa, El-Shaimaa A

    2016-01-01

    Despite its broad applications, cisplatin affords considerable nephro- and hepatotoxicity through triggering inflammatory and oxidative stress cascades. The aim of the current investigation was to study the possible protective effects of tangeretin on cisplatin-induced hepatotoxicity. The impact of tangeretin on cisplatin-evoked hepatic dysfunction and histopathologic changes along with oxidative stress, inflammatory and apoptotic biomarkers were investigated compared to silymarin. Tangeretin pre-treatment significantly improved liver function tests (ALT and AST), inhibited cisplatin-induced lipid profile aberrations (total cholesterol and triglycerides) and diminished histopathologic structural damage in liver tissues. Tangeretin also attenuated cisplatin-induced hepatic inflammatory events as indicated by suppression of tumor necrosis factor-α (TNF-α) and enhancement of interleukin-10 (IL-10). Meanwhile, it lowered malondialdehyde (MDA), nitric oxide (NO) and nuclear factor erythroid 2-related factor 2 (NRF-2) levels with restoration of glutathione (GSH), and glutathione peroxidase (GPx). Regarding mitogen-activated protein kinase (MAPK) pathway, tangeretin attenuated cisplatin-induced increase in phospho-p38, phospho-c-Jun N-terminal kinase (p-JNK) and phospho-extracellular signal-regulated kinase (p-ERK1/2) in liver tissues. In addition, tangeretin downregulated Bax expression with augmentation of Bcl-2 promoting liver cell survival. Our results highlight the protective effects of tangeretin against cisplatin-induced acute hepatic injury via the concerted modulation of inflammation, oxidative stress, MAPKs and apoptotic pathways.

  18. Tangeretin Alleviates Cisplatin-Induced Acute Hepatic Injury in Rats: Targeting MAPKs and Apoptosis.

    Directory of Open Access Journals (Sweden)

    Hany A Omar

    Full Text Available Despite its broad applications, cisplatin affords considerable nephro- and hepatotoxicity through triggering inflammatory and oxidative stress cascades. The aim of the current investigation was to study the possible protective effects of tangeretin on cisplatin-induced hepatotoxicity. The impact of tangeretin on cisplatin-evoked hepatic dysfunction and histopathologic changes along with oxidative stress, inflammatory and apoptotic biomarkers were investigated compared to silymarin. Tangeretin pre-treatment significantly improved liver function tests (ALT and AST, inhibited cisplatin-induced lipid profile aberrations (total cholesterol and triglycerides and diminished histopathologic structural damage in liver tissues. Tangeretin also attenuated cisplatin-induced hepatic inflammatory events as indicated by suppression of tumor necrosis factor-α (TNF-α and enhancement of interleukin-10 (IL-10. Meanwhile, it lowered malondialdehyde (MDA, nitric oxide (NO and nuclear factor erythroid 2-related factor 2 (NRF-2 levels with restoration of glutathione (GSH, and glutathione peroxidase (GPx. Regarding mitogen-activated protein kinase (MAPK pathway, tangeretin attenuated cisplatin-induced increase in phospho-p38, phospho-c-Jun N-terminal kinase (p-JNK and phospho-extracellular signal-regulated kinase (p-ERK1/2 in liver tissues. In addition, tangeretin downregulated Bax expression with augmentation of Bcl-2 promoting liver cell survival. Our results highlight the protective effects of tangeretin against cisplatin-induced acute hepatic injury via the concerted modulation of inflammation, oxidative stress, MAPKs and apoptotic pathways.

  19. FGFR3 promotes synchondrosis closure and fusion of ossification centers through the MAPK pathway

    Science.gov (United States)

    Matsushita, Takehiko; Wilcox, William R.; Chan, Yuk Yu; Kawanami, Aya; Bükülmez, Hülya; Balmes, Gener; Krejci, Pavel; Mekikian, Pertchoui B.; Otani, Kazuyuki; Yamaura, Isakichi; Warman, Matthew L.; Givol, David; Murakami, Shunichi

    2009-01-01

    Activating mutations in FGFR3 cause achondroplasia and thanatophoric dysplasia, the most common human skeletal dysplasias. In these disorders, spinal canal and foramen magnum stenosis can cause serious neurologic complications. Here, we provide evidence that FGFR3 and MAPK signaling in chondrocytes promote synchondrosis closure and fusion of ossification centers. We observed premature synchondrosis closure in the spine and cranial base in human cases of homozygous achondroplasia and thanatophoric dysplasia as well as in mouse models of achondroplasia. In both species, premature synchondrosis closure was associated with increased bone formation. Chondrocyte-specific activation of Fgfr3 in mice induced premature synchondrosis closure and enhanced osteoblast differentiation around synchondroses. FGF signaling in chondrocytes increases Bmp ligand mRNA expression and decreases Bmp antagonist mRNA expression in a MAPK-dependent manner, suggesting a role for Bmp signaling in the increased bone formation. The enhanced bone formation would accelerate the fusion of ossification centers and limit the endochondral bone growth. Spinal canal and foramen magnum stenosis in heterozygous achondroplasia patients, therefore, may occur through premature synchondrosis closure. If this is the case, then any growth-promoting treatment for these complications of achondroplasia must precede the timing of the synchondrosis closure. PMID:18923003

  20. Kaempferol Attenuates Cardiac Hypertrophy via Regulation of ASK1/MAPK Signaling Pathway and Oxidative Stress.

    Science.gov (United States)

    Feng, Hong; Cao, Jianlei; Zhang, Guangyu; Wang, Yanggan

    2017-02-20

    Kaempferol has been demonstrated to provide benefits for the treatment of atherosclerosis, coronary heart disease, hyperlipidemia, and diabetes through its antioxidant and anti-inflammatory properties. However, its role in cardiac hypertrophy remains to be elucidated. The aim of our study was to investigate the effects of kaempferol on cardiac hypertrophy and the underlying mechanism. Mice subjected to aorta banding were treated with or without kaempferol (100 mg/kg/d, p. o.) for 6 weeks. Echocardiography was performed to evaluate cardiac function. Mice hearts were collected for pathological observation and molecular mechanism investigation. H9c2 cardiomyocytes were stimulated with or without phenylephrine for in vitro study. Kaempferol significantly attenuated cardiac hypertrophy induced by aorta banding as evidenced by decreased cardiomyocyte areas and interstitial fibrosis, accompanied with improved cardiac functions and decreased apoptosis. The ASK1/MAPK signaling pathways (JNK1/2 and p38) were markedly activated in the aorta banding mouse heart but inhibited by kaempferol treatment. In in vitro experiments, kaempferol also inhibited the activity of ASK1/JNK1/2/p38 signaling pathway and the enlargement of H9c2 cardiomyocytes. Furthermore, our study revealed that kaempferol could protect the mouse heart and H9c2 cells from pathological oxidative stress. Our investigation indicated that treatment with kaempferol protects against cardiac hypertrophy, and its cardioprotection may be partially explained by the inhibition of the ASK1/MAPK signaling pathway and the regulation of oxidative stress.

  1. Apigenin promotes osteogenic differentiation of human mesenchymal stem cells through JNK and p38 MAPK pathways.

    Science.gov (United States)

    Zhang, Xue; Zhou, Chenhui; Zha, Xuan; Xu, Zhoumei; Li, Li; Liu, Yuyu; Xu, Liangliang; Cui, Liao; Xu, Daohua; Zhu, Baohua

    2015-09-01

    Apigenin is a plant-derived flavonoid and has been reported to prevent bone loss in ovariectomized mice, but the role of apigenin on osteogenic differentiation of human mesenchymal stem cells (hMSCs) has not been reported. In the present study, the effect of apigenin on osteogenic differentiation of hMSCs was explored. Our results showed that apigenin treatment significantly increased alkaline phosphatase (ALP) activity and mineralization in hMSCs. RT-PCR revealed that apigenin markedly up-regulated the mRNA expression of osteopontin (OPN) and the transcription factors runt-related transcription factor 2 (Runx2). The expression of Runx2 and osterix (OSX) proteins were also increased in hMSCs differentiating into osteoblasts after treatment with apigenin. Furthermore, we investigated the signaling pathways responsible for osteogenic differentiation of apigenin in hMSCs. We found that apigenin treatment significantly increased the levels of p-JNK, p-p38 in hMSCs and addition of the inhibitors of JNK (SP600125) or p38 MAPK (SB203580) eliminated the stimulating effects of apigenin. In addition, addition of SP600125 or SB203580 also blocked apigenin-induced ALP activity, OPN, Runx2, and OSX expression and meanwhile inhibited bone nodule formation. Taken together, these findings suggest apigenin promotes the osteogenesis of hMSCs through activation of JNK and p38 MAPK signal pathways which leads to Runx2 and OSX expressions to induce the formation of bone nodule.

  2. Clinical candidates of small molecule p38 MAPK inhibitors for inflammatory diseases

    Directory of Open Access Journals (Sweden)

    Li Xing

    2016-01-01

    Full Text Available The trigger and etiology of chronic inflammatory diseases are not well understood, hindering the development of efficient therapeutic approaches. The observation that abnormal activity of the p38 MAPK is common to all inflammatory diseases raised the expectation that p38 inhibitors would serve as general anti-inflammatory therapeutics. A large number of inhibitors were consequently discovered. Several compounds of different scaffolds, blocking the p38 MAPK signaling pathway, have entered phase II clinical trials for rheumatoid arthritis, chronic obstructive pulmonary disease, pain, cardiovascular diseases, and cancer. As I review here, in almost all cases the clinical trials have failed, leading to re-design of compounds and re-evaluation of p38 as a suitable target. I describe how structural features, unique to p38α, have been employed in the inhibitor design and achieved high degree of kinome selectivity. I then focus on some of the drugs that reached human trials and summarize their in vitro/in vivo pharmacological profiles and the related outcomes from clinical investigations. These compounds include VX-745, VX-702, RO-4402257, SCIO- 469, BIRB-796, SD-0006, PH-797804, AMG-548, LY2228820, SB-681323 and GW-856553. Finally, I discuss novel suggested approaches for the use of p38 inhibitors such as combining p38 inhibition with inhibiting other targets that function in parallel inflammatory pathways for achieving efficacy in treating inflammatory diseases.

  3. MALT1 Inhibition of Oral Carcinoma Cell Invasion and ERK/MAPK Activation.

    Science.gov (United States)

    Chiba, T; Soeno, Y; Shirako, Y; Sudo, H; Yagishita, H; Taya, Y; Kawashiri, S; Okada, Y; Imai, K

    2016-04-01

    The expression of mucosa-associated lymphoid tissue 1 (MALT1) that activates nuclear factor (NF)-κB in lymphocyte lineages is rapidly inactivated in oral carcinoma cells at the invasive front and the patients with worst prognosis. However, its mechanism to accelerate carcinoma progression remains unknown, and this study was carried out to examine the role in invasion. HSC2 oral carcinoma cells stably expressing wild-type MALT1 (wtMALT1) reduced the invasion of basement membrane matrices and collagen gels, and the dominant-negative form (∆MALT1)-expressing cells aggressively invaded into collagen gels. MALT1 decelerated proliferation and migration of cells and downregulated expression of matrix metalloproteinase 2 and 9, which were confirmed by short interfering RNA transfections. Reporter assays and immunoblot analysis showed that MALT1 does not affect the NF-κB pathway but inhibits ERK/MAPK activation. This was confirmed by endogenous MALT1 expression in oral carcinoma cell lines. Orthotopic implantation of ∆MALT1-expressing HSC2 cells in mice grew rapid expansive and invasive tongue tumors in contrast to an absence of tumor formation by wtMALT1-expressing cells. These results demonstrate that MALT1 suppresses oral carcinoma invasion by inhibiting proliferation, migration, and extracellular matrix degradation and that the ERK/MAPK pathway is a target of MALT1 and further suggests a role as a suppressor of carcinoma progression.

  4. Interaction between PGE2 and EGF receptor through MAPKs in mouse embryonic stem cell proliferation.

    Science.gov (United States)

    Yun, S P; Lee, M Y; Ryu, J M; Han, H J

    2009-05-01

    Identifying the small molecules that permit precise regulation of embryonic stem (ES) cell proliferation should further support our understanding of the underlying molecular mechanisms of self renewal. In the present study, we showed that PGE(2) increased [(3)H]-thymidine incorporation in a time and dose dependent manner. In addition, PGE(2) increased the expression of cell cycle regulatory proteins, the percentage of cells in S phase and the total number of cells. PGE(2) obviously increased E-type prostaglandin (EP) receptor 1 mRNA expression level compare to 2, 3, 4 subtypes. EP1 antagonist also blocked PGE(2)-induced cell cycle regulatory protein expression and thymidine incorporation. PGE(2) caused phosphorylation of protein kinase C, Src, epidermal growth factor (EGF) receptor, phosphatidylinositol 3-kinase (PI3K)/Akt phosphorylation, and p44/42 mitogen-activated protein kinase (MAPK), which were blocked by each inhibitors. In conclusion, PGE(2)-stimulated proliferation is mediated by MAPK via EP1 receptor-dependent PKC and EGF receptor-dependent PI3K/Akt signaling pathways in mouse ES cells.

  5. Prostaglandin E2 induces expression of MAPK phosphatase 1 (MKP-1) in airway smooth muscle cells.

    Science.gov (United States)

    Rumzhum, Nowshin N; Ammit, Alaina J

    2016-07-05

    Prostaglandin E2 (PGE2) is a prostanoid with diverse actions in health and disease. In chronic respiratory diseases driven by inflammation, PGE2 has both positive and negative effects. An enhanced understanding of the receptor-mediated cellular signalling pathways induced by PGE2 may help us separate the beneficial properties from unwanted actions of this important prostaglandin. PGE2 is known to exert anti-inflammatory and bronchoprotective actions in human airways. To date however, whether PGE2 increases production of the anti-inflammatory protein MAPK phosphatase 1 (MKP-1) was unknown. We address this herein and use primary cultures of human airway smooth muscle (ASM) cells to show that PGE2 increases MKP-1 mRNA and protein upregulation in a concentration-dependent manner. We explore the signalling pathways responsible and show that PGE2-induces CREB phosphorylation, not p38 MAPK activation, in ASM cells. Moreover, we utilize selective antagonists of EP2 (PF-04418948) and EP4 receptors (GW 627368X) to begin to identify EP-mediated functional outcomes in ASM cells in vitro. Taken together with earlier studies, our data suggest that PGE2 increases production of the anti-inflammatory protein MKP-1 via cAMP/CREB-mediated cellular signalling in ASM cells and demonstrates that EP2 may, in part, be involved. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Weaning Induced Hepatic Oxidative Stress, Apoptosis, and Aminotransferases through MAPK Signaling Pathways in Piglets

    Science.gov (United States)

    Luo, Zhen; Zhu, Wei; Guo, Qi; Luo, Wenli; Zhang, Jing; Xu, Weina

    2016-01-01

    This study investigated the effects of weaning on the hepatic redox status, apoptosis, function, and the mitogen-activated protein kinase (MAPK) signaling pathways during the first week after weaning in piglets. A total of 12 litters of piglets were weaned at d 21 and divided into the weaning group (WG) and the control group (CG). Six piglets from each group were slaughtered at d 0 (d 20, referred to weaning), d 1, d 4, and d 7 after weaning. Results showed that weaning significantly increased the concentrations of hepatic free radicals H2O2 and NO, malondialdehyde (MDA), and 8-hydroxy-2′-deoxyguanosine (8-OHdG), while significantly decreasing the inhibitory hydroxyl ability (IHA) and glutathione peroxidase (GSH-Px), and altered the level of superoxide dismutase (SOD). The apoptosis results showed that weaning increased the concentrations of caspase-3, caspase-8, caspase-9 and the ratio of Bax/Bcl-2. In addition, aspartate aminotransferase transaminase (AST) and alanine aminotransferase (ALT) in liver homogenates increased after weaning. The phosphorylated JNK and ERK1/2 increased, while the activated p38 initially decreased and then increased. Our results suggested that weaning increased the hepatic oxidative stress and aminotransferases and initiated apoptosis, which may be related to the activated MAPK pathways in postweaning piglets.

  7. Signaling intermediates (MAPK and PI3K) as therapeutic targets in NSCLC.

    Science.gov (United States)

    Ciuffreda, Ludovica; Incani, Ursula Cesta; Steelman, Linda S; Abrams, Stephen L; Falcone, Italia; Curatolo, Anais Del; Chappell, William H; Franklin, Richard A; Vari, Sabrina; Cognetti, Francesco; McCubrey, James A; Milella, Michele

    2014-01-01

    The RAS/RAF/MEK/ ERK and the PI3K/AKT/mTOR pathways govern fundamental physiological processes, such as cell proliferation, differentiation, metabolism, cytoskeleton reorganization and cell death and survival. Constitutive activation of these signal transduction pathways is a required hallmark of cancer and dysregulation, on either genetic or epigenetic grounds, of these pathways has been implicated in the initiation, progression and metastastic spread of lung cances. Targeting components of the MAPK and PI3K cascades is thus an attractive strategy in the development of novel therapeutic approaches to treat lung cancer, although the use of single pathway inhibitors has met with limited clinical success so far. Indeed, the presence of intra- and inter-pathway compensatory loops that re-activate the very same cascade, either upstream or downstream the point of pharmacological blockade, or activate the alternate pathway following the blockade of one signaling cascade has been demonstrated, potentially driving preclinical (and possibly clinical) resistance. Therefore, the blockade of both pathways with combinations of signaling inhibitors might result in a more efficient anti-tumor effect, and thus potentially overcome and/or delay clinical resistance, as compared with single agent. The current review aims at summarizing the current status of preclinical and clinical research with regard to pathway crosstalks between the MAPK and PI3K cascades in NSCLC and the rationale for combined therapeutic pathway targeting.

  8. Adapting glycolysis to cancer cell proliferation: the MAPK pathway focuses on PFKFB3.

    Science.gov (United States)

    Bolaños, Juan P

    2013-06-15

    Besides the necessary changes in the expression of cell cycle-related proteins, cancer cells undergo a profound series of metabolic adaptations focused to satisfy their excessive demand for biomass. An essential metabolic transformation of these cells is increased glycolysis, which is currently the focus of anticancer therapies. Several key players have been identified, so far, that adapt glycolysis to allow an increased proliferation in cancer. In this issue of the Biochemical Journal, Novellasdemunt and colleagues elegantly identify a novel mechanism by which MK2 [MAPK (mitogen-activated protein kinase)-activated protein kinase 2], a key component of the MAPK pathway, up-regulates glycolysis in response to stress in cancer cells. The authors found that, by phosphorylating specific substrate residues, MK2 promotes both increased the gene transcription and allosteric activation of PFKFB3 (6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3), a key glycolysis-promoting enzyme. These results reveal a novel pathway through which MK2 co-ordinates metabolic adaptation to cell proliferation in cancer and highlight PFKFB3 as a potential therapeutic target in this devastating disease.

  9. SIRT1 regulates MAPK pathways in vitiligo skin: insight into the molecular pathways of cell survival.

    Science.gov (United States)

    Becatti, Matteo; Fiorillo, Claudia; Barygina, Victoria; Cecchi, Cristina; Lotti, Torello; Prignano, Francesca; Silvestro, Agrippino; Nassi, Paolo; Taddei, Niccolò

    2014-03-01

    Vitiligo is an acquired and progressive hypomelanotic disease that manifests as circumscribed depigmented patches on the skin. The aetiology of vitiligo remains unclear, but recent experimental data underline the interactions between melanocytes and other typical skin cells, particularly keratinocytes. Our previous results indicate that keratinocytes from perilesional skin show the features of damaged cells. Sirtuins (silent mating type information regulation 2 homolog) 1, well-known modulators of lifespan in many species, have a role in gene repression, metabolic control, apoptosis and cell survival, DNA repair, development, inflammation, neuroprotection and healthy ageing. In the literature there is no evidence for SIRT1 signalling in vitiligo and its possible involvement in disease progression. Here, biopsies were taken from the perilesional skin of 16 patients suffering from non-segmental vitiligo and SIRT1 signalling was investigated in these cells. For the first time, a new SIRT1/Akt, also known as Protein Kinase B (PKB)/mitogen-activated protein kinase (MAPK) signalling has been revealed in vitiligo. SIRT1 regulates MAPK pathway via Akt-apoptosis signal-regulating kinase-1 and down-regulates pro-apoptotic molecules, leading to decreased oxidative stress and apoptotic cell death in perilesional vitiligo keratinocytes. We therefore propose SIRT1 activation as a novel way of protecting perilesional vitiligo keratinocytes from damage.

  10. CHIP promotes thyroid cancer proliferation via activation of the MAPK and AKT pathways

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Li [Department of Pharmacy, Urumchi General Hospital of Lanzhou Military Region, Urumchi, Xinjiang 830000 (China); Liu, Lianyong [Medical College of Soochow University, Suzhou, Jiangsu 215123 (China); Department of Endocrinology, Shanghai Punan Hospital, Shanghai 200125 (China); He, Xiaohua; Shen, Yunling; Liu, Xuerong; Wei, Jing; Yu, Fang [Department of Endocrinology, Urumchi General Hospital of Lanzhou Military Region, Urumchi, Xinjiang 830000 (China); Tian, Jianqing, E-mail: jianqing0991@163.com [Department of Endocrinology, Urumchi General Hospital of Lanzhou Military Region, Urumchi, Xinjiang 830000 (China)

    2016-08-26

    The carboxyl terminus of Hsp70-interacting protein (CHIP) is a U box-type ubiquitin ligase that plays crucial roles in various biological processes, including tumor progression. To date, the functional mechanism of CHIP in thyroid cancer remains unknown. Here, we obtained evidence of upregulation of CHIP in thyroid cancer tissues and cell lines. CHIP overexpression markedly enhanced thyroid cancer cell viability and colony formation in vitro and accelerated tumor growth in vivo. Conversely, CHIP knockdown impaired cell proliferation and tumor growth. Notably, CHIP promoted cell growth through activation of MAPK and AKT pathways, subsequently decreasing p27 and increasing cyclin D1 and p-FOXO3a expression. Our findings collectively indicate that CHIP functions as an oncogene in thyroid cancer, and is therefore a potential therapeutic target for this disease. - Highlights: • CHIP is significantly upregulated in thyroid cancer cells. • Overexpression of CHIP facilitates proliferation and tumorigenesis of thyroid cancer cells. • Silencing of CHIP inhibits the proliferation and tumorigenesis of thyroid cancer cells. • CHIP promotes thyroid cancer cell proliferation via activating the MAPK and AKT pathways.

  11. Klf4 Is a Transcriptional Regulator of Genes Critical for EMT, Including Jnk1 (Mapk8)

    Science.gov (United States)

    Tiwari, Neha; Meyer-Schaller, Nathalie; Arnold, Phil; Antoniadis, Helena; Pachkov, Mikhail; van Nimwegen, Erik; Christofori, Gerhard

    2013-01-01

    We have identified the zinc-finger transcription factor Kruppel-like factor 4 (Klf4) among the transcription factors that are significantly downregulated in their expression during epithelial-mesenchymal transition (EMT) in mammary epithelial cells and in breast cancer cells. Loss and gain of function experiments demonstrate that the down-regulation of Klf4 expression is required for the induction of EMT in vitro and for metastasis in vivo. In addition, reduced Klf4 expression correlates with shorter disease-free survival of subsets of breast cancer patients. Yet, reduced expression of Klf4 also induces apoptosis in cells undergoing TGFβ-induced EMT. Chromatin immunoprecipitation/deep-sequencing in combination with gene expression profiling reveals direct Klf4 target genes, including E-cadherin (Cdh1), N-cadherin (Cdh2), vimentin (Vim), β-catenin (Ctnnb1), VEGF-A (Vegfa), endothelin-1 (Edn1) and Jnk1 (Mapk8). Thereby, Klf4 acts as a transcriptional activator of epithelial genes and as a repressor of mesenchymal genes. Specifically, increased expression of Jnk1 (Mapk8) upon down-regulation of its transcriptional repressor Klf4 is required for EMT cell migration and for the induction of apoptosis. The data demonstrate a central role of Klf4 in the maintenance of epithelial cell differentiation and the prevention of EMT and metastasis. PMID:23451207

  12. Phosphoproteomic analysis of protein kinase C signaling in Saccharomyces cerevisiae reveals Slt2 mitogen-activated protein kinase (MAPK)-dependent phosphorylation of eisosome core components.

    Science.gov (United States)

    Mascaraque, Victoria; Hernáez, María Luisa; Jiménez-Sánchez, María; Hansen, Rasmus; Gil, Concha; Martín, Humberto; Cid, Víctor J; Molina, María

    2013-03-01

    The cell wall integrity (CWI) pathway of the model organism Saccharomyces cerevisiae has been thoroughly studied as a paradigm of the mitogen-activated protein kinase (MAPK) pathway. It consists of a classic MAPK module comprising the Bck1 MAPK kinase kinase, two redundant MAPK kinases (Mkk1 and Mkk2), and the Slt2 MAPK. This module is activated under a variety of stimuli related to cell wall homeostasis by Pkc1, the only member of the protein kinase C family in budding yeast. Quantitative phosphoproteomics based on stable isotope labeling of amino acids in cell culture is a powerful tool for globally studying protein phosphorylation. Here we report an analysis of the yeast phosphoproteome upon overexpression of a PKC1 hyperactive allele that specifically activates CWI MAPK signaling in the absence of external stimuli. We found 82 phosphopeptides originating from 43 proteins that showed enhanced phosphorylation in these conditions. The MAPK S/T-P target motif was significantly overrepresented in these phosphopeptides. Hyperphosphorylated proteins provide putative novel targets of the Pkc1-cell wall integrity pathway involved in diverse functions such as the control of gene expression, protein synthesis, cytoskeleton maintenance, DNA repair, and metabolism. Remarkably, five components of the plasma-membrane-associated protein complex known as eisosomes were found among the up-regulated proteins. We show here that Pkc1-induced phosphorylation of the eisosome core components Pil1 and Lsp1 was not exerted directly by Pkc1, but involved signaling through the Slt2 MAPK module.

  13. Resilience to audiogenic seizures is associated with p-ERK1/2 dephosphorylation in the subiculum of Fmr1 knockout mice

    Directory of Open Access Journals (Sweden)

    Giulia eCuria

    2013-04-01

    Full Text Available Young, but not adult, Fmr1 knockout (KO mice display audiogenic seizures (AGS that can be prevented by inhibiting extracellular signal-regulated kinases 1/2 (ERK1/2 phosphorylation. In order to identify the cerebral regions involved in these phenomena, we characterized the response to AGS in Fmr1 KO mice and wild type (WT controls at postnatal day (P 45 and P90. To characterize the diverse response to AGS in various cerebral regions, we evaluated the activity markers FosB/ΔFosB and phosphorylated ERK1/2 (p-ERK1/2. Wild running (100% of tested mice followed by clonic/tonic seizures (30% were observed in P45 Fmr1 KO mice, but not in WT mice. In P90 Fmr1 KO mice, wild running was only present in 25% of tested animals. Basal FosB/ΔFosB immunoreactivity was higher (P<0.01 vs WT in the CA1 and subiculum of P45 Fmr1 KO mice. Following the AGS test, FosB/ΔFosB expression consistently increased in most of the analyzed regions in both groups at P45, but not at P90. Interestingly, FosB/ΔFosB immunoreactivity was significantly higher in P45 Fmr1 KO mice in the medial geniculate body (P<0.05 vs WT and CA3 (P<0.01. Neurons presenting with immunopositivity to p-ERK1/2 were more abundant in the subiculum of Fmr1 KO mice in control condition (P<0.05 vs WT, in both age groups. In this region, p-ERK1/2-immunopositive cells significantly decreased (-75%, P<0.01 in P90 Fmr1 KO mice exposed to the AGS test, but no changes were found in P45 mice or in other brain regions. In both age groups of WT mice, p-ERK1/2-immunopositive cells increased in the subiculum after exposure to the acoustic test. Our findings illustrate that FosB/ΔFosB markers are overexpressed in the medial geniculate body and CA3 in Fmr1 KO mice experiencing AGS, and that p-ERK1/2 is markedly decreased in the subiculum of Fmr1 KO mice resistant to AGS induction. These findings suggest that resilience to AGS is associated with dephosphorylation of p-ERK1/2 in the subiculum of mature Fmr1 KO mice.

  14. Activation of the MAPK11/12/13/14 (p38 MAPK) pathway regulates the transcription of autophagy genes in response to oxidative stress induced by a novel copper complex in HeLa cells.

    Science.gov (United States)

    Zhong, Wu; Zhu, Haichuan; Sheng, Fugeng; Tian, Yonglu; Zhou, Jun; Chen, Yingyu; Li, Song; Lin, Jian

    2014-07-01

    Transition metal copper (Cu) can exist in oxidized or reduced states in cells, leading to cytotoxicity in cancer cells through oxidative stress. Recently, copper complexes are emerging as a new class of anticancer compounds. Here, we report that a novel anticancer copper complex (HYF127c/Cu) induces oxidative stress-dependent cell death in cancer cells. Further, transcriptional analysis revealed that oxidative stress elicits broad transcriptional changes of genes, in which autophagy-related genes are significantly changed in HYF127c/Cu-treated cells. Consistently, autophagy was induced in HYF127c/Cu-treated cells and inhibitors of autophagy promoted cell death induced by HYF127c/Cu. Further analysis identified that the MAPK11/12/13/14 (formerly known as p38 MAPK) pathway was also activated in HYF127c/Cu-treated cells. Meanwhile, the MAPK11/12/13/14 inhibitor SB203580 downregulated autophagy by inhibiting the transcription of the autophagy genes MAP1LC3B, BAG3, and HSPA1A, and promoted HYF127c/Cu-induced cell death. These data suggest that copper-induced oxidative stress will induce protective autophagy through transcriptional regulation of autophagy genes by activation of the MAPK11/12/13/14 pathway in HeLa cells.

  15. Sequential allergen desensitization of basophils is non-specific and may involve p38 MAPK.

    Science.gov (United States)

    Witting Christensen, S K; Kortekaas Krohn, I; Thuraiaiyah, J; Skjold, T; Schmid, J M; Hoffmann, H J H

    2014-10-01

    Sequential allergen desensitization provides temporary tolerance for allergic patients. We adapted a clinical protocol to desensitize human blood basophils ex vivo and investigated the mechanism and allergen specificity. We included 28 adult, grass allergic subjects. The optimal, activating allergen concentration was determined by measuring activated CD63(+) CD193(+) SS(Low) basophils in a basophil activation test with 8 log-dilutions of grass allergen. Basophils in whole blood were desensitized by incubation with twofold to 2.5-fold increasing allergen doses in 10 steps starting at 1 : 1000 of the optimal dose. Involvement of p38 mitogen-activated protein kinase (MAPK) was assessed after 3 min of allergen stimulation (n = 7). Allergen specificity was investigated by desensitizing cells from multi-allergic subjects with grass allergen and challenging with optimal doses of grass, birch, recombinant house dust mite (rDer p2) allergen or anti-IgE (n = 10). Desensitization reduced the fraction of blood basophils responding to challenge with an optimal allergen dose from a median (IQR) 81.0% (66.3-88.8) to 35.4% (19.8-47.1, P desensitized with grass allergen. Challenge with grass allergen resulted in 39.6% activation (15.8-58.3). An unrelated challenge (birch, rDer p2 or anti-IgE) resulted in 53.4% activation (30.8-66.8, P = 0.16 compared with grass). Desensitization reduced p38 MAPK phosphorylation from a median 48.1% (15.6-92.8) to 26.1% (7.4-71.2, P = 0.047) and correlated with decrease in CD63 upregulation (n = 7, r > 0.79, P Desensitization attenuated basophil response rapidly and non-specifically at a stage before p38 MAPK phosphorylation. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  16. Deoxynivalenol induced mouse skin cell proliferation and inflammation via MAPK pathway

    Energy Technology Data Exchange (ETDEWEB)

    Mishra, Sakshi [Food Drug and Chemical Toxicology, CSIR-Indian Institute of Toxicology Research (CSIR-IITR), P.O. Box No. 80, Mahatma Gandhi Marg, Lucknow 226 001 (India); Department of Biochemistry, Banaras Hindu University (BHU), Varanasi (India); Tripathi, Anurag [Food Drug and Chemical Toxicology, CSIR-Indian Institute of Toxicology Research (CSIR-IITR), P.O. Box No. 80, Mahatma Gandhi Marg, Lucknow 226 001 (India); Chaudhari, Bhushan P. [Pathology Laboratory, CSIR-Indian Institute of Toxicology Research (CSIR-IITR), Mahatma Gandhi Marg, PO Box 80, Lucknow 226001, Uttar Pradesh (India); Dwivedi, Premendra D. [Food Drug and Chemical Toxicology, CSIR-Indian Institute of Toxicology Research (CSIR-IITR), P.O. Box No. 80, Mahatma Gandhi Marg, Lucknow 226 001 (India); Pandey, Haushila P. [Department of Biochemistry, Banaras Hindu University (BHU), Varanasi (India); Das, Mukul, E-mail: mditrc@rediffmail.com [Food Drug and Chemical Toxicology, CSIR-Indian Institute of Toxicology Research (CSIR-IITR), P.O. Box No. 80, Mahatma Gandhi Marg, Lucknow 226 001 (India)

    2014-09-01

    Several toxicological manifestations of deoxynivalenol (DON), a mycotoxin, are well documented; however, dermal toxicity is not yet explored. The effect of topical application of DON to mice was studied using markers of skin proliferation, inflammation and tumor promotion. Single topical application of DON (84–672 nmol/mouse) significantly enhanced dermal hyperplasia and skin edema. DON (336 and 672 nmol) caused significant enhancement in [{sup 3}H]-thymidine uptake in DNA along with increased myeloperoxidase and ornithine decarboxylase activities, suggesting tissue inflammation and cell proliferation. Furthermore, DON (168 nmol) caused enhanced expression of RAS, and phosphorylation of PI3K/Akt, ERK, JNK and p38 MAPKs. DON exposure also showed activation of transcription factors, c-fos, c-jun and NF-κB along with phosphorylation of IkBα. Enhanced phosphorylation of NF-κB by DON caused over expression of target proteins, COX-2, cyclin D1 and iNOS in skin. Though a single topical application of DMBA followed by twice weekly application of DON (84 and 168 nmol) showed no tumorigenesis after 24 weeks, however, histopathological studies suggested hyperplasia of the epidermis and hypertrophy of hair follicles. Interestingly, intestine was also found to be affected as enlarged Peyer's patches were observed, suggesting inflammatory effects which were supported by elevation of inflammatory cytokines after 24 weeks of topical application of DON. These results suggest that DON induced cell proliferation in mouse skin is through the activation of MAPK signaling pathway involving transcription factors NFκB and AP-1, further leading to transcriptional activation of downstream target proteins c-fos, c-jun, cyclin D1, iNOS and COX-2 which might be responsible for its inflammatory potential. - Highlights: • Topical application of DON enhanced epidermal inflammation and cell proliferation. • DON follows PI3K/Akt/MAPK signaling cascade, with activation of AP-1 and NF

  17. Polycyclic aromatic hydrocarbon (PAH)-mediated upregulation of hepatic microRNA-181 family promotes cancer cell migration by targeting MAPK phosphatase-5, regulating the activation of p38 MAPK

    Energy Technology Data Exchange (ETDEWEB)

    Song, Mi-Kyung [Center for Integrated Risk Research, Cellular and Molecular Toxicology Laboratory, Korea Institute of Science and Technology (KIST) (Korea, Republic of); School of Life Sciences and Biotechnology, Korea University, Anam-Dong, Seoungbuk-Gu, Seoul 136-701 (Korea, Republic of); Park, Yong-Keun [School of Life Sciences and Biotechnology, Korea University, Anam-Dong, Seoungbuk-Gu, Seoul 136-701 (Korea, Republic of); Ryu, Jae-Chun, E-mail: ryujc@kist.re.kr [Center for Integrated Risk Research, Cellular and Molecular Toxicology Laboratory, Korea Institute of Science and Technology (KIST) (Korea, Republic of)

    2013-11-15

    Growing evidence indicates that changes in microRNA (miRNA) expression in cancer induced by chemical carcinogens play an important role in cancer development and progression by regulating related genes. However, the mechanisms underlying miRNA involvement in hepatocarcinogenesis induced by polycyclic aromatic hydrocarbons (PAHs) remain unclear. Thus, the identification of aberrant miRNA expression during PAH-induced cancer cell migration will lead to a better understanding of the substantial role of miRNAs in cancer progression. In the present study, miRNA expression profiling showed significant upregulation of miR-181a, -181b, and -181d in human hepatocellular carcinoma cells (HepG2 line) exposed to benzo[a]anthracene (BA) and benzo[k]fluoranthene (BF). MAPK phosphatase-5 (MKP-5), a validated miR-181 target that deactivates MAPKs, was markedly suppressed while phosphorylation of p38 MAPK was increased after BA and BF exposure. The migration of HepG2 cells, observed using the scratch wound-healing assay, also increased in a dose-dependent manner. Depletion of miR-181 family members by miRNA inhibitors enhanced the expression of MKP-5 and suppressed the phosphorylation of p38 MAPK. Furthermore, the depletion of the miR-181 family inhibited cancer cell migration. Based on these results, we conclude that the miR-181 family plays a critical role in PAH-induced hepatocarcinogenesis by targeting MKP-5, resulting in the regulation of p38 MAPK activation. - Highlights: • We found significant upregulation of miR-181 family in HCC exposed to BA and BF. • We identified the MKP-5 as a putative target of miR-181 family. • MKP-5 was suppressed while p-P38 was increased after BA and BF exposure. • The migration of HepG2 cells increased in a dose-dependent manner.

  18. Reactive oxygen species mediate nitric oxide production through ERK/JNK MAPK signaling in HAPI microglia after PFOS exposure.

    Science.gov (United States)

    Wang, Cheng; Nie, Xiaoke; Zhang, Yan; Li, Ting; Mao, Jiamin; Liu, Xinhang; Gu, Yiyang; Shi, Jiyun; Xiao, Jing; Wan, Chunhua; Wu, Qiyun

    2015-10-15

    Perfluorooctane sulfonate (PFOS), an emerging persistent contaminant that is commonly encountered during daily life, has been shown to exert toxic effects on the central nervous system (CNS). However, the molecular mechanisms underlying the neurotoxicity of PFOS remain largely unknown. It has been widely acknowledged that the inflammatory mediators released by hyper-activated microglia play vital roles in the pathogenesis of various neurological diseases. In the present study, we examined the impact of PFOS exposure on microglial activation and the release of proinflammatory mediators, including nitric oxide (NO) and reactive oxidative species (ROS). We found that PFOS exposure led to concentration-dependent NO and ROS production by rat HAPI microglia. We also discovered that there was rapid activation of the ERK/JNK MAPK signaling pathway in the HAPI microglia following PFOS treatment. Moreover, the PFOS-induced iNOS expression and NO production were attenuated after the inhibition of ERK or JNK MAPK by their corresponding inhibitors, PD98059 and SP600125. Interestingly, NAC, a ROS inhibitor, blocked iNOS expression, NO production, and activation of ERK and JNK MAPKs, which suggested that PFOS-mediated microglial NO production occurs via a ROS/ERK/JNK MAPK signaling pathway. Finally, by exposing SH-SY5Y cells to PFOS-treated microglia-conditioned medium, we demonstrated that NO was responsible for PFOS-mediated neuronal apoptosis.

  19. Mitogen-activated protein kinases (MAPKs) regulate IL-6 over-production during concomitant influenza virus and Staphylococcus aureus infection

    Science.gov (United States)

    Klemm, Carolin; Bruchhagen, Christin; van Krüchten, Andre; Niemann, Silke; Löffler, Bettina; Peters, Georg; Ludwig, Stephan; Ehrhardt, Christina

    2017-01-01

    Bacterial super-infections are a major complication of influenza virus (IV) infections and often lead to severe pneumonia. One hallmark of IV-associated Staphylococcus aureus (S. aureus) infection is rapid progression to a serious disease outcome. Changes in immune and inflammatory host responses increase morbidity and complicate efficient therapy. A key player during inflammation is the multifunctional cytokine IL-6. Although increased IL-6 levels have been observed after severe disease upon IV and/or bacterial super-infection, the underlying molecular mechanisms still remain to be elucidated. In the present study, we focused on cellular signalling pathways regulating IL-6 production upon IV/S. aureus super-infection. Additionally, infection with viable bacteria was mimicked by lipoteichoic acid stimulation in this model. Analyses of cellular signalling mechanisms revealed synergistically increased activation of the MAPK p38 as well as enhanced phosphorylation of the MAPKs ERK1/2 and JNK in the presence of super-infecting bacteria. Interestingly, inhibition of MAPK activity indicated a strong dependence of IL-6 expression on p38 and ERK1/2, while the MAPK JNK seems not to be involved. Thus, our results provide new molecular insights into the regulation of IL-6, a marker of severe disease, which might contribute to the lethal synergism of IV and S. aureus. PMID:28195157

  20. Internalization of EGF receptor following lipid rafts disruption in keratinocytes is delayed and dependent on p38 MAPK activation

    DEFF Research Database (Denmark)

    Lambert, S.; Ameels, H.; Gniadecki, R.;

    2008-01-01

    internalization without participation of the ligand under the control of p38 MAPK during stress conditions. Since cholesterol depletion using methyl-beta-cyclodextrin is known to induce ligand-independent activation of EGFR in keratinocytes, we investigated by confocal microscopy and ligand-binding tests...

  1. Small Molecule APY606 Displays Extensive Antitumor Activity in Pancreatic Cancer via Impairing Ras-MAPK Signaling

    Science.gov (United States)

    Guo, Na; Liu, Zuojia; Zhao, Wenjing; Wang, Erkang; Wang, Jin

    2016-01-01

    Pancreatic cancer has been found with abnormal expression or mutation in Ras proteins. Oncogenic Ras activation exploits their extensive signaling reach to affect multiple cellular processes, in which the mitogen-activated protein kinase (MAPK) signaling exerts important roles in tumorigenesis. Therapies targeted Ras are thus of major benefit for pancreatic cancer. Although small molecule APY606 has been successfully picked out by virtual drug screening based on Ras target receptor, its in-depth mechanism remains to be elucidated. We herein assessed the antitumor activity of APY606 against human pancreatic cancer Capan-1 and SW1990 cell lines and explored the effect of Ras-MAPK and apoptosis-related signaling pathway on the activity of APY606. APY606 treatment resulted in a dose- and time-dependent inhibition of cancer cell viability. Additionally, APY606 exhibited strong antitumor activity, as evidenced not only by reduction in tumor cell invasion, migration and mitochondrial membrane potential but also by alteration in several apoptotic indexes. Furthermore, APY606 treatment directly inhibited Ras-GTP and the downstream activation of MAPK, which resulted in the down-regulation of anti-apoptotic protein Bcl-2, leading to the up-regulation of mitochondrial apoptosis pathway-related proteins (Bax, cytosolic Cytochrome c and Caspase 3) and of cyclin-dependent kinase 2 and Cyclin A, E. These data suggest that impairing Ras-MAPK signaling is a novel mechanism of action for APY606 during therapeutic intervention in pancreatic cancer. PMID:27223122

  2. Mos in the Oocyte: How to Use MAPK Independently of Growth Factors and Transcription to Control Meiotic Divisions

    Directory of Open Access Journals (Sweden)

    Aude Dupré

    2011-01-01

    Full Text Available In many cell types, the mitogen-activated protein kinase (MAPK also named extracellular signal-regulated kinase (ERK is activated in response to a variety of extracellular growth factor-receptor interactions and leads to the transcriptional activation of immediate early genes, hereby influencing a number of tissue-specific biological activities, as cell proliferation, survival and differentiation. In one specific cell type however, the female germ cell, MAPK does not follow this canonical scheme. In oocytes, MAPK is activated independently of growth factors and tyrosine kinase receptors, acts independently of transcriptional regulation, plays a crucial role in controlling meiotic divisions, and is under the control of a peculiar upstream regulator, the kinase Mos. Mos was originally identified as the transforming gene of Moloney murine sarcoma virus and its cellular homologue was the first proto-oncogene to be molecularly cloned. What could be the specific roles of Mos that render it necessary for meiosis? Which unique functions could explain the evolutionary cost to have selected one gene to only serve for few hours in one very specific cell type? This review discusses the original features of MAPK activation by Mos and the roles of this module in oocytes.

  3. p38 MAPK mediates cardiovascular and behavioral responses induced by central IL-1β and footshock in conscious rats

    Institute of Scientific and Technical Information of China (English)

    Rui-mao ZHENG; Chang-jiang ZOU; Shi-gong ZHU

    2004-01-01

    AIM: To investigate the roles of p38 mitogen-activated protein kinase (p38 MAPK) in the cardiovascular and behavioral responses induced by intracerebral ventricular injection (icv) of interleukin- 1 β (IL- 1 β) or footshock.METHODS: We examined the effects of p38 MAPK on mean artery blood pressure (mABP), heart rate (HR), and motor activity (MA) during central administration of IL- 1 β, or footshock after icv SB203580 (a specific inhibitor of the p38 MAPK) with Cardiovascular and Behavior Telemetry System in conscious SD rats. RESULTS: (1) IL-1 β (icy) or footshock remarkably rise the mABP, and the maximal changes are (7.8± 1.8) and (12.3±3.5) mmHg,respectively, which was abrogated by the pretreatment with p38 inhibitor SB203580 intracerebroventricularly. (2)Compared with icv saline group, the motor activity was significantly decreased in SB203580 group with maximal changes (-7.6± 1.1) counts/min after footshock. CONCLUSION: p38 MAPK plays an important role in the pressor response induced by central administration of IL- 1 β or footshock and change of motor activity after footshock in conscious rats.

  4. p38α MAPK regulates proliferation and differentiation of osteoclast progenitors and bone remodeling in an aging-dependent manner

    Science.gov (United States)

    Cong, Qian; Jia, Hao; Li, Ping; Qiu, Shoutao; Yeh, James; Wang, Yibin; Zhang, Zhen-Lin; Ao, Junping; Li, Baojie; Liu, Huijuan

    2017-01-01

    Bone mass is determined by the balance between bone formation, carried out by mesenchymal stem cell-derived osteoblasts, and bone resorption, carried out by monocyte-derived osteoclasts. Here we investigated the potential roles of p38 MAPKs, which are activated by growth factors and cytokines including RANKL and BMPs, in osteoclastogenesis and bone resorption by ablating p38α MAPK in LysM+monocytes. p38α deficiency promoted monocyte proliferation but regulated monocyte osteoclastic differentiation in a cell-density dependent manner, with proliferating p38α−/− cultures showing increased differentiation. While young mutant mice showed minor increase in bone mass, 6-month-old mutant mice developed osteoporosis, associated with an increase in osteoclastogenesis and bone resorption and an increase in the pool of monocytes. Moreover, monocyte-specific p38α ablation resulted in a decrease in bone formation and the number of bone marrow mesenchymal stem/stromal cells, likely due to decreased expression of PDGF-AA and BMP2. The expression of PDGF-AA and BMP2 was positively regulated by the p38 MAPK-Creb axis in osteoclasts, with the promoters of PDGF-AA and BMP2 having Creb binding sites. These findings uncovered the molecular mechanisms by which p38α MAPK regulates osteoclastogenesis and coordinates osteoclastogenesis and osteoblastogenesis. PMID:28382965

  5. Increases in cAMP, MAPK activity, and CREB phosphorylation during REM sleep: implications for REM sleep and memory consolidation.

    Science.gov (United States)

    Luo, Jie; Phan, Trongha X; Yang, Yimei; Garelick, Michael G; Storm, Daniel R

    2013-04-10

    The cyclic adenosine monophosphate (cAMP), mitogen-activated protein kinase (MAPK), and cAMP response element-binding protein (CREB) transcriptional pathway is required for consolidation of hippocampus-dependent memory. In mice, this pathway undergoes a circadian oscillation required for memory persistence that reaches a peak during the daytime. Because mice exhibit polyphasic sleep patterns during the day, this suggested the interesting possibility that cAMP, MAPK activity, and CREB phosphorylation may be elevated during sleep. Here, we report that cAMP, phospho-p44/42 MAPK, and phospho-CREB are higher in rapid eye movement (REM) sleep compared with awake mice but are not elevated in non-REM sleep. This peak of activity during REM sleep does not occur in mice lacking calmodulin-stimulated adenylyl cyclases, a mouse strain that learns but cannot consolidate hippocampus-dependent memory. We conclude that a preferential increase in cAMP, MAPK activity, and CREB phosphorylation during REM sleep may contribute to hippocampus-dependent memory consolidation.

  6. Failure to Target RANKL Signaling Through p38-MAPK Results in Defective Osteoclastogenesis in the Microphthalmia Cloudy-Eyed Mutant.

    Science.gov (United States)

    Carey, Heather A; Bronisz, Agnieszka; Cabrera, Jennifer; Hildreth, Blake E; Cuitiño, Maria; Fu, Qi; Ahmad, Asrar; Toribio, Ramiro E; Ostrowski, Michael C; Sharma, Sudarshana M

    2016-03-01

    The Microphthalmia-associated transcription factor (MITF) is a basic helix-loop-helix leucine zipper family factor that is essential for terminal osteoclast differentiation. Previous work demonstrates that phosphorylation of MITF by p38 MAPK downstream of Receptor Activator of NFkB Ligand (RANKL) signaling is necessary for MITF activation in osteoclasts. The spontaneous Mitf cloudy eyed (ce) allele results in production of a truncated MITF protein that lacks the leucine zipper and C-terminal end. Here we show that the Mitf(ce) allele leads to a dense bone phenotype in neonatal mice due to defective osteoclast differentiation. In response to RANKL stimulation, in vitro osteoclast differentiation was impaired in myeloid precursors derived from neonatal or adult Mitf(ce/ce) mice. The loss of the leucine zipper domain in Mitf(ce/ce) mice does not interfere with the recruitment of MITF/PU.1 complexes to target promoters. Further, we have mapped the p38 MAPK docking site within the region deleted in Mitf(ce). This interaction is necessary for the phosphorylation of MITF by p38 MAPK. Site-directed mutations in the docking site interfered with the interaction between MITF and its co-factors FUS and BRG1. MITF-ce fails to recruit FUS and BRG1 to target genes, resulting in decreased expression of target genes and impaired osteoclast function. These results highlight the crucial role of signaling dependent MITF/p38 MAPK interactions in osteoclast differentiation.

  7. Reduced Drought Tolerance by CRISPR/Cas9-Mediated SlMAPK3 Mutagenesis in Tomato Plants.

    Science.gov (United States)

    Wang, Liu; Chen, Lin; Li, Rui; Zhao, Ruirui; Yang, Meijing; Sheng, Jiping; Shen, Lin

    2017-10-04

    Drought stress is one of the most destructive environmental factors that affect tomato plants adversely. Mitogen-activated protein kinases (MAPKs) are important signaling molecules that respond to drought stress. In this study, SlMAPK3 was induced by drought stress, and the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (CRISPR/Cas9) system was utilized to generate slmapk3 mutants. Two independent T1 transgenic lines and wild-type (WT) tomato plants were used for analysis of drought tolerance. Compared with WT plants, slmapk3 mutants exhibited more severe wilting symptom, higher hydrogen peroxide content, lower antioxidant enzymes activities, and suffered more membrane damage under drought stress. Furthermore, knockout of SlMAPK3 led to up- or down-regulated expressions of drought stress-responsive genes including SlLOX, SlGST, and SlDREB. The results suggest that SlMAPK3 is involved in drought response in tomato plants by protecting cell membranes from oxidative damage and modulating transcription of stress-related genes.

  8. Inhibition of MAPK-mediated ACE expression by compound C66 prevents STZ-induced diabetic nephropathy.

    Science.gov (United States)

    Pan, Yong; Huang, Yi; Wang, Zhe; Fang, Qilu; Sun, Yusheng; Tong, Chao; Peng, Kesong; Wang, Yangwei; Miao, Lining; Cai, Lu; Zhao, Yunjie; Liang, Guang

    2014-02-01

    A range of in vitro, experimental and clinical intervention studies have implicated an important role for hyperglycaemia-induced activation of the renin-angiotensin system (RAS) in the development and progression of diabetic nephropathy (DN). Blockade of RAS by angiotensin converting enzyme (ACE) inhibitors is an effective strategy in treating diabetic kidney diseases. However, few studies demonstrate the mechanism by which hyperglycaemia up-regulates the expression of ACE gene. Our previous studies have identified a novel curcumin analogue, (2E,6E)-2,6-bis(2-(trifluoromethyl)benzylidene)cyclohexanone (C66), which could inhibit the high glucose (HG)-induced phosphorylation of mitogen-activated protein kinases in mouse macrophages. In this study, we found that the renal protection of C66 in diabetic mice was associated with mitogen-activated protein kinase (MAPK) inactivation and ACE/angiotensin II (Ang II) down-regulation. Generally, MAPKs have been considered as a downstream signalling of Ang II and a mediator for Ang II-induced pathophysiological actions. However, using C66 and specific inhibitors as small molecule probes, in vitro experiments demonstrate that the MAPK signalling pathway regulates ACE expression under HG stimulation, which contributes to renal Ang II activation and the development of DN. This study indicates that C66 is a potential candidate of DN therapeutic agents, and more importantly, that reduction in ACE expression by MAPKs inhibition seems to be an alternative strategy for the treatment of DN.

  9. CONTRIBUTION OF INSPIRATORY FLOW TO ACTIVATION OF EGFR, RAS, MAPK, ATF-2 AND C-JUN DURING LUNG STRETCH

    Science.gov (United States)

    Contribution of Inspiratory Flow to Activation of EGFR, Ras, MAPK, ATF-2 and c-Jun during Lung StretchR. Silbajoris 1, Z. Li 2, J. M. Samet 1 and Y. C. Huang 1. 1 NHEERL, ORD, US EPA, RTP, NC and 2 CEMALB, UNC-CH, Chapel Hill, NC . Mechanical ventilation with larg...

  10. The effect of cytoplasmic regions of LIF receptor on activating MAPK p42/44 in HL-60 cell

    Institute of Scientific and Technical Information of China (English)

    LIU Hou-qi; SHAN Ji-kuan; TANG Shu-ping; LIU Shan-rong; JI Kai-hong

    2001-01-01

    Objective: To study the relation between the cytoplasmic region of each subunit of leukemia inhibitory factor (LIF) receptor and mitogen-activated protein kinase (MAPK) in human leukemia line HL-60 for studying the mechanism of leukemia cell proliferation disorder. Methods: We prepared the chimerical receptor with exchanged cytoplasmic domain (190/130,130/190), expressed it on the membrane of HL-60, and then bound LIF with the wild type receptor competitively. The cytoplasmic region homodimerization (190cyt- 190cyt, 130cyt- 130cyt) initiated intracellular signal transduction, cell proliferation and differentiation. MAPK phosphorylation level was observed by means of immunoblotting and immunobiochemistry. Results: Higher level ofMAPK p42/p44 phosphorylation (Thr202Tyr204) and faster cell proliferation were determined in group 190/130 compared with the wild type receptor. Those in group 130/190 were lower than the others. Conclusion: The cytoplasmic domain ofgpl30 involves in the induction of MAPK activation and the cell proliferation of HL-60.

  11. Regulation of EGF-induced ERK/MAPK Activation and EGFR Internalization by G Protein-coupled Receptor Kinase 2

    Institute of Scientific and Technical Information of China (English)

    Jingxia GAO; Jiali LI; Lan MA

    2005-01-01

    G protein-coupled receptor kinases (GRKs) mediate agonist-induced phosphorylation and desensitization of various G protein-coupled receptors (GPCRs). We investigate the role of GRK2 on epidermal growth factor (EGF) receptor signaling, including EGF-induced extracellular signal-regulated kinase and mitogen-activated protein kinase (ERK/MAPK) activation and EGFR internalization. Immunoprecipitation and immunofluorescence experiments show that EGF stimulates GRK2 binding to EGFR complex and GRK2 translocating from cytoplasm to the plasma membrane in human embryonic kidney 293 cells. Western blotting assay shows that EGF-induced ERK/MAPK phosphorylation increases 1.9-fold, 1.1-fold and 1.5-fold (P<0.05) at time point 30, 60 and 120 min, respectively when the cells were transfected with GRK2,suggesting the regulatory role of GRK2 on EGF-induced ERK/MAPK activation. Flow cytometry experiments show that GRK2 overexpression has no effect on EGF-induced EGFR internalization, however, it increases agonist-induced G protein-coupled δ opioid receptor internalization by approximately 40% (P<0.01). Overall,these data suggest that GRK2 has a regulatory role in EGF-induced ERK/MAPK activation, and that the mechanisms underlying the modulatory role of GRK2 in EGFR and GPCR signaling pathways are somewhat different at least in receptor internalization.

  12. Egr-1 decreases adipocyte insulin sensitivity by tilting PI3K/Akt and MAPK signal balance in mice

    Science.gov (United States)

    Yu, Xiao; Shen, Ning; Zhang, Ming-Liang; Pan, Fei-Yan; Wang, Chen; Jia, Wei-Ping; Liu, Chang; Gao, Qian; Gao, Xiang; Xue, Bin; Li, Chao-Jun

    2011-01-01

    It is well known that insulin can activate both PI3K/Akt pathway, which is responsible for glucose uptake, and MAPK pathway, which is crucial for insulin resistance formation. But, it is unclear exactly how the two pathways coordinate to regulate insulin sensitivity upon hyperinsulinism stress of type 2 diabetes mellitus (T2DM). Here, we show that an early response transcription factor Egr-1 could tilt the signalling balance by blocking PI3K/Akt signalling through PTEN and augmenting Erk/MAPK signalling through GGPPS, resulting in insulin resistance in adipocytes. Egr-1, PTEN and GGPPS are upregulated in the fat tissue of T2DM patients and db/db mice. Egr-1 overexpression in epididymal fat induced systematic insulin resistance in wild-type mice, and loss of Egr-1 function improved whole-body insulin sensitivity in diabetic mice, which is mediated by Egr-1 controlled PI3K/Akt and Erk/MAPK signalling balance. Therefore, we have revealed, for the first time, the mechanism by which Egr-1 induces insulin resistance under hyperinsulinism stress, which provides an ideal pharmacological target since inhibiting Egr-1 can simultaneously block MAPK and augment PI3K/Akt activation during insulin stimulation. PMID:21829168

  13. The role of the MAPK pathway alterations in GM-CSF modulated human neutrophil apoptosis with aging

    Directory of Open Access Journals (Sweden)

    Fortin Carl

    2005-03-01

    Full Text Available Abstract Background Neutrophils represent the first line of defence against aggressions. The programmed death of neutrophils is delayed by pro-inflammatory stimuli to ensure a proper resolution of the inflammation in time and place. The pro-inflammatory stimuli include granulocyte-macrophage colony-stimulating factor (GM-CSF. Recently, we have demonstrated that although neutrophils have an identical spontaneous apoptosis in elderly subjects compared to that in young subjects, the GM-CSF-induced delayed apoptosis is markedly diminished. The present study investigates whether an alteration of the GM-CSF stimulation of MAPKs play a role in the diminished rescue from apoptosis of PMN of elderly subjects. Methods Neutrophils were separated from healthy young and elderly donors satisfying the SENIEUR protocol. Neutrophils were stimulated with GM-CSF and inhibitors of the MAPKinase pathway. Apoptosis commitment, phosphorylation of signaling molecules, caspase-3 activities as well as expression of pro- and anti-apoptotic molecules were performed in this study. Data were analyzed using Student's two-tailed t-test for independent means. Significance was set for p ≤ 0.05 unless stated otherwise. Results In this paper we present evidence that an alteration in the p42/p44 MAPK activation occurs in PMN of elderly subjects under GM-CSF stimulation and this plays a role in the decreased delay of apoptosis of PMN in elderly. We also show that p38 MAPK does not play a role in GM-CSF delayed apoptosis in PMN of any age-groups, while it participates to the spontaneous apoptosis. Our results also show that the alteration of the p42/p44 MAPK activation contributes to the inability of GM-CSF to decrease the caspase-3 activation in PMN of elderly subjects. Moreover, GM-CSF converts the pro-apoptotic phenotype to an anti-apoptotic phenotype by modulating the bcl-2 family members Bax and Bcl-xL in PMN of young subjects, while this does not occur in PMN of elderly

  14. MMPs 2 and 9 are essential for coronary collateral growth and are prominently regulated by p38 MAPK

    Science.gov (United States)

    Dodd, Tracy; Jadhav, Rashmi; Wiggins, Luke; Stewart, James; Smith, Erika; Russell, James C.; Rocic, Petra

    2011-01-01

    Transient, repetitive ischemia (RI) stimulates coronary collateral growth (CCG) in normal, healthy (SD) rats, which requires p38 MAPK activation. In contrast, RI does not induce CCG in the metabolic syndrome (JCR) rats, which is associated with lack of p38 MAPK activation. The functional consequences of p38 MAPK activation in CCG remain unknown. Theoretically, effective collateral growth would require extracellular matrix remodeling; however, direct assessment as well as identification of proteases responsible for this degradation are lacking. In this study, we investigated the role of p38 MAPK in the regulation of matrix metalloproteinases 2 and 9 (MMPs 2 and 9) and their requirement for CCG in SD vs. JCR rats. The rats underwent the RI protocol (8 LAD occlusions, 40 sec each, every 20 min, in 8 hr cycles for 0, 3, 6, or 9 days). MMP expression was measured in the ischemic, collateral-dependent zone (CZ) and the normal zone (NZ) by Western blot, and MMP activity by zymography. Expression and activation of MMP 2 and 9 were significantly increased (~3.5 fold) on day 3 of RI in the CZ of SD rats. In vivo p38 MAPK inhibition completely blocked RI-induced MMP 2 and 9 expression and activation. MMP activation correlated with increased degradation of components of the basement membrane and the vascular elastic laminae: elastin (~3 fold), laminin (~3 fold) and type IV collagen (~2 fold). This was blocked by MMP 2 and 9 inhibition, which also abolished RI-induced CCG. In contrast, in JCR rats, RI did not induce expression or activation of MMP 2 or 9 and there was no associated degradation of elastin, laminin or type IV collagen. In conclusion, MMP 2 and 9 activation is essential for CCG and is mediated, in part, by p38 MAPK. Furthermore, compromised CCG in the metabolic syndrome may be partially due to the lack of p38 MAPK-dependent activation of MMP 2 and 9 and resultant decreased extracellular matrix degradation. PMID:21884701

  15. A new NFIA:RAF1 fusion activating the MAPK pathway in pilocytic astrocytoma

    DEFF Research Database (Denmark)

    Yde, Christina Westmose; Sehested, Astrid; Mateu-Regué, Àngels

    2016-01-01

    Pilocytic astrocytoma (PA) is one of the most common brain cancers among children and activation of the Mitogen-Activated Protein Kinase (MAPK) pathway is considered the hallmark. In the majority of cases, oncogenic BRAF fusions or BRAF V600E mutations are observed, while RAF1 or NF1 alterations...... are more rarely found. However, in some cases, no apparent cancer driver events can be identified. Here, we describe a novel fusion between the transcription factor nuclear factor 1A (NFIA) and Raf-1 proto-oncogene (RAF1) in a 5-year old boy with PA. The novel fusion was identified as part...... of a comprehensive genomic tumor profiling. We show that the NFIA:RAF1 fusion results in constitutive Raf1 kinase activity, leading to activation of downstream MEK1/2 cascade and increased proliferation of cancer cells. The NFIA:RAF1 fusion displayed distinct subcellular localization towards the plasma membrane...

  16. Cordyceps bassiana inhibits smooth muscle cell proliferation via the ERK1/2 MAPK signaling pathway.

    Science.gov (United States)

    Jin, Enze; Han, Seongho; Son, Mina; Kim, Sung-Whan

    2016-01-01

    Cordyceps belongs to a genus of acormycete fungi and is known to exhibit various pharmacological effects. The aim of this study was to investigate the effect of Cordyceps species on the proliferation of vascular smooth muscle cells (VSMC) and their underlying molecular mechanism. A cell proliferation assay showed that Cordyceps bassiana ethanol extract (CBEE) significantly inhibited VSMC proliferation. In addition, neointimal formation was significantly reduced by treatment with CBEE in the carotid artery of balloon-injured rats. We also investigated the effects of CBEE on the extracellular signal-regulated kinase (ERK) signal pathway. Western blot analysis revealed increased ERK 1/2 phosphorylation in VSMCs treated with CBEE. Pretreatment with U0126 completely abrogated CBEE-induced ERK 1/2 phosphorylation. In conclusion, CBEE exhibited anti-proliferative properties that affected VSMCs through the ERK1/2 MAPK signaling pathway. Our data may elucidate the inhibitory mechanism of this natural product.

  17. CHIP promotes thyroid cancer proliferation via activation of the MAPK and AKT pathways.

    Science.gov (United States)

    Zhang, Li; Liu, Lianyong; He, Xiaohua; Shen, Yunling; Liu, Xuerong; Wei, Jing; Yu, Fang; Tian, Jianqing

    2016-08-26

    The carboxyl terminus of Hsp70-interacting protein (CHIP) is a U box-type ubiquitin ligase that plays crucial roles in various biological processes, including tumor progression. To date, the functional mechanism of CHIP in thyroid cancer remains unknown. Here, we obtained evidence of upregulation of CHIP in thyroid cancer tissues and cell lines. CHIP overexpression markedly enhanced thyroid cancer cell viability and colony formation in vitro and accelerated tumor growth in vivo. Conversely, CHIP knockdown impaired cell proliferation and tumor growth. Notably, CHIP promoted cell growth through activation of MAPK and AKT pathways, subsequently decreasing p27 and increasing cyclin D1 and p-FOXO3a expression. Our findings collectively indicate that CHIP functions as an oncogene in thyroid cancer, and is therefore a potential therapeutic target for this disease.

  18. ATXN1L, CIC, and ETS Transcription Factors Modulate Sensitivity to MAPK Pathway Inhibition

    Science.gov (United States)

    Wang, Belinda; Krall, Elsa Beyer; Aguirre, Andrew James; Kim, Miju; Widlund, Hans Ragnar; Doshi, Mihir Bhavik; Sicinska, Ewa; Sulahian, Rita; Goodale, Amy; Cowley, Glenn Spencer; Piccioni, Federica; Doench, John Gerard; Root, David Edward; Hahn, William Chun

    2017-01-01

    SUMMARY Intrinsic resistance and RTK-RAS-MAPK pathway reactivation has limited the effectiveness of MEK and RAF inhibitors (MAPKi) in RAS- and RAF-mutant cancers. To identify genes that modulate sensitivity to MAPKi, we performed genome scale CRISPR-Cas9 loss-of-function screens in two KRAS-mutant pancreatic cancer cell lines treated with the MEK1/2 inhibitor trametinib. Loss of CIC, a transcriptional repressor of ETV1, 4, and 5, promoted survival in the setting of MAPKi in cancer cells derived from several lineages. ATXN1L deletion, which reduces CIC protein, or ectopic expression of ETV1, 4, or 5 also modulated sensitivity to trametinib. ATXN1L expression inversely correlates with response to MAPKi inhibition in clinical studies. These observations identify the ATXN1L-CIC-ETS transcription factor axis as a mediator of resistance to MAPKi. PMID:28178529

  19. A new NFIA:RAF1 fusion activating the MAPK pathway in pilocytic astrocytoma

    DEFF Research Database (Denmark)

    Yde, Christina Westmose; Sehested, Astrid; Regué, Àngels Mateu

    2016-01-01

    are more rarely found. However, in some cases, no apparent cancer driver events can be identified. Here, we describe a novel fusion between the transcription factor nuclear factor 1A (NFIA) and Raf-1 proto-oncogene (RAF1) in a 5-year old boy with PA. The novel fusion was identified as part......Pilocytic astrocytoma (PA) is one of the most common brain cancers among children and activation of the Mitogen-Activated Protein Kinase (MAPK) pathway is considered the hallmark. In the majority of cases, oncogenic BRAF fusions or BRAF V600E mutations are observed, while RAF1 or NF1 alterations...... of a comprehensive genomic tumor profiling. We show that the NFIA:RAF1 fusion results in constitutive Raf1 kinase activity, leading to activation of downstream MEK1/2 cascade and increased proliferation of cancer cells. The NFIA:RAF1 fusion displayed distinct subcellular localization towards the plasma membrane...

  20. Low cell cholesterol levels increase NFkappaB activity through a p38 MAPK-dependent mechanism.

    Science.gov (United States)

    Calleros, Laura; Lasa, Marina; Toro, María J; Chiloeches, Antonio

    2006-12-01

    Cholesterol, p38 MAPK and NFkappaB have been shown to participate in inflammation and cellular differentiation. Here, we examined the effect of cholesterol on NFkappaB-dependent transcription and the mechanisms underlying this effect in NIH3T3 cells. We show that chronic cholesterol depletion achieved with lipoprotein-deficient serum (LPDS) and 25-hydroxycholesterol (25-HC) treatment resulted in a significant increase in NFkappaB-dependent transcription, NFkappaB-DNA binding, IkappaBalpha degradation and p65/NFkappaB translocation to the nucleus, and the addition of exogenous cholesterol reversed these effects. Previously, we have shown that low cell cholesterol levels activate p38 MAPK. Here, we found that inhibition of p38 MAPK with the specific inhibitor SB203580 blocked the increase in NFkappaB activity, IkappaBalpha degradation and p65/NFkappaB translocation to the nucleus induced by cholesterol depletion. Moreover, the inhibition of the p38 MAPK downstream effector MSK1 with the specific inhibitor H89, or the overexpression of a kinase defective MSK1 abrogated the NFkappaB-dependent transcription induced by cholesterol depletion. On the other hand, the transactivation potential of p65/NFkappaB depends on phosphorylation of S276 by MSK1. We observed that cholesterol depletion increased the p65/NFkappaB transactivation capacity. This effect was reversed by cell cholesterol repletion or incubation with the SB203580 inhibitor. Moreover, the expression of a p65/NFkappaB S276A mutant was insensitive to cholesterol depletion. Together, our results demonstrate that cholesterol depletion induces NFkappaB transcriptional activity, not only by affecting the IkappaBalpha degradation and the translocation of p65/NFkappaB to the nucleus, but also regulating the p65/NFkappaB transactivating potential through a p38 MAPK/MSK1 mediated pathway.

  1. Missense Variant in MAPK Inactivator PTPN5 Is Associated with Decreased Severity of Post-Burn Hypertrophic Scarring.

    Directory of Open Access Journals (Sweden)

    Ravi F Sood

    Full Text Available Hypertrophic scarring (HTS is hypothesized to have a genetic mechanism, yet its genetic determinants are largely unknown. The mitogen-activated protein kinase (MAPK pathways are important mediators of inflammatory signaling, and experimental evidence implicates MAPKs in HTS formation. We hypothesized that single-nucleotide polymorphisms (SNPs in MAPK-pathway genes would be associated with severity of post-burn HTS.We analyzed data from a prospective-cohort genome-wide association study of post-burn HTS. We included subjects with deep-partial-thickness burns admitted to our center who provided blood for genotyping and had at least one Vancouver Scar Scale (VSS assessment. After adjusting for HTS risk factors and population stratification, we tested MAPK-pathway gene SNPs for association with the four VSS variables in a joint regression model. In addition to individual-SNP analysis, we performed gene-based association testing.Our study population consisted of 538 adults (median age 40 years who were predominantly White (76% males (71% admitted to our center from 2007-2014 with small-to-moderate-sized burns (median burn size 6% total body surface area. Of 2,146 SNPs tested, a rare missense variant in the PTPN5 gene (rs56234898; minor allele frequency 1.5% was significantly associated with decreased severity of post-burn HTS (P = 1.3×10-6. In gene-based analysis, PTPN5 (P = 1.2×10-5 showed a significant association and BDNF (P = 9.5×10-4 a borderline-significant association with HTS severity.We report PTPN5 as a novel genetic locus associated with HTS severity. PTPN5 is a MAPK inhibitor expressed in neurons, suggesting a potential role for neurotrophic factors and neuroinflammatory signaling in HTS pathophysiology.

  2. Missense Variant in MAPK Inactivator PTPN5 Is Associated with Decreased Severity of Post-Burn Hypertrophic Scarring.

    Science.gov (United States)

    Sood, Ravi F; Arbabi, Saman; Honari, Shari; Gibran, Nicole S

    2016-01-01

    Hypertrophic scarring (HTS) is hypothesized to have a genetic mechanism, yet its genetic determinants are largely unknown. The mitogen-activated protein kinase (MAPK) pathways are important mediators of inflammatory signaling, and experimental evidence implicates MAPKs in HTS formation. We hypothesized that single-nucleotide polymorphisms (SNPs) in MAPK-pathway genes would be associated with severity of post-burn HTS. We analyzed data from a prospective-cohort genome-wide association study of post-burn HTS. We included subjects with deep-partial-thickness burns admitted to our center who provided blood for genotyping and had at least one Vancouver Scar Scale (VSS) assessment. After adjusting for HTS risk factors and population stratification, we tested MAPK-pathway gene SNPs for association with the four VSS variables in a joint regression model. In addition to individual-SNP analysis, we performed gene-based association testing. Our study population consisted of 538 adults (median age 40 years) who were predominantly White (76%) males (71%) admitted to our center from 2007-2014 with small-to-moderate-sized burns (median burn size 6% total body surface area). Of 2,146 SNPs tested, a rare missense variant in the PTPN5 gene (rs56234898; minor allele frequency 1.5%) was significantly associated with decreased severity of post-burn HTS (P = 1.3×10-6). In gene-based analysis, PTPN5 (P = 1.2×10-5) showed a significant association and BDNF (P = 9.5×10-4) a borderline-significant association with HTS severity. We report PTPN5 as a novel genetic locus associated with HTS severity. PTPN5 is a MAPK inhibitor expressed in neurons, suggesting a potential role for neurotrophic factors and neuroinflammatory signaling in HTS pathophysiology.

  3. l-carnitine preserves cardiac function by activating p38 MAPK/Nrf2 signalling in hearts exposed to irradiation.

    Science.gov (United States)

    Fan, Zhigang; Han, Yang; Ye, Yuanpeng; Liu, Chao; Cai, Hui

    2017-06-05

    Radiation-induced heart damage (RIHD) is now considered to be one of the causes of mortality in cancer patients undergoing radiotherapy. Cardiac function impairments are clinical manifestations of RIHD. L-carnitine shows protective effects against irradiation and heart disease. This study was aimed to investigate the cardioprotective effects and potential molecular mechanisms of L-carnitine against RIHD. Mouse hearts were exposed to γ-radiation to induce RIHD. L-carnitine at doses of 100mg/Kg and 200mg/Kg was used to treat animals intraperitoneally. Additionally, a specific inhibitor of p38 MAPK was used to treat animals by intraperitoneal injections. Cardiac systolic/diastolic functions were determined using invasive hemodynamic methods; myocyte apoptosis was assessed using the TUNEL assay; intracellular reactive oxygen species production was measured using DHE staining; and western blotting was used to evaluate the phosphorylation of p38MAPK, phosphorylation of Nrf2, and expression levels of HO1, NQO1, caspase3 and bax. L-carnitine treatments inhibited irradiation induced cardiac function impairments. Radiation exposure induced myocyte apoptosis and reactive oxygen species production, which were attenuated by L-carnitine treatments. However, administration of a p38 MAPK inhibitor (SB203580) dramatically impaired L-carnitine's effect on attenuating apoptosis, reactive oxygen species accumulation and cardiac functions in irradiated hearts. Our study showed that L-carnitine administration activated p38MAPK/Nrf2 signalling, initiating the expression of HO1 and NQO1, which have anti-apoptotic and anti-oxidative effects, respectively. In conclusion, L-carnitine attenuates cardiac function loss by inhibiting reactive oxygen species production and apoptosis in hearts exposed to radiation. The cardioprotective effects of L-carnitine were mediated by p38MAPK/Nrf2 signalling. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. The MAPK pathway is involved in the regulation of rapid pacing-induced ionic channel remodeling in rat atrial myocytes.

    Science.gov (United States)

    Cheng, Wei; Zhu, Yun; Wang, Haidong

    2016-03-01

    Alterations to the expression L‑type calcium channels (LTCCs) and Kv4.3 potassium channels form the possible basis of atrial electrical remodeling during rapid pacing. The mitogen‑activated protein kinase (MAPK) pathway is affected by increases in cytoplasmic Ca2+, and therefore represents an attractive candidate for the regulation and mediation of Ca2+‑induced ion channel remodeling. The present study aimed to investigate alterations to the ion channel‑MAPK axis, and to determine its influence on ion channel remodeling during atrial fibrillation. Rat atrial myocytes were isolated, cultured, and in vitro rapid pacing was established. Intracellular Ca2+ signals were monitored using the Fluo‑3/AM Ca2+ indicator. Verapamil, PD98058 and SB203580 were added to the culture medium of various groups at specific time‑points. The mRNA expression levels of LTCC‑α1c and Kv4.3 potassium channels were detected by reverse transcription‑polymerase chain reaction. Western blotting was performed to determine the expression levels of channel and signaling proteins. The results demonstrated that fast pacing significantly increased the intracellular Ca2+ concentration in atrial myocytes, whereas treatment with verapamil markedly inhibited this increase. In addition, verapamil significantly antagonized the rapid pacing‑induced activation of extracellular signal‑regulated kinase (ERK) and p38MAPK. These results indicated that the MAPK pathway may have an important role in the opening of LTCCs, and alterations to MAPK molecule expression could affect the expression and remodeling of ion channels.

  5. LPS Induces Occludin Dysregulation in Cerebral Microvascular Endothelial Cells via MAPK Signaling and Augmenting MMP-2 Levels

    Directory of Open Access Journals (Sweden)

    Lan-hui Qin

    2015-01-01

    Full Text Available Disrupted blood-brain barrier (BBB integrity contributes to cerebral edema during central nervous system infection. The current study explored the mechanism of lipopolysaccharide- (LPS- induced dysregulation of tight junction (TJ proteins. Human cerebral microvascular endothelial cells (hCMEC/D3 were exposed to LPS, SB203580 (p38MAPK inhibitor, or SP600125 (JNK inhibitor, and cell vitality was determined by MTT assay. The proteins expressions of p38MAPK, JNK, and TJs (occludin and zonula occludens- (ZO- 1 were determined by western blot. The mRNA levels of TJ components and MMP-2 were measured with quantitative real-time polymerase chain reaction (qRT-PCR, and MMP-2 protein levels were determined by enzyme-linked immunosorbent assay (ELISA. LPS, SB203580, and SP600125 under respective concentrations of 10, 7.69, or 0.22 µg/mL had no effects on cell vitality. Treatment with LPS decreased mRNA and protein levels of occludin and ZO-1 and enhanced p38MAPK and JNK phosphorylation and MMP-2 expression. These effects were attenuated by pretreatment with SB203580 or SP600125, but not in ZO-1 expression. Both doxycycline hyclate (a total MMP inhibitor and SB-3CT (a specific MMP-2 inhibitor partially attenuated the LPS-induced downregulation of occludin. These data suggest that MMP-2 overexpression and p38MAPK/JNK pathways are involved in the LPS-mediated alterations of occludin in hCMEC/D3; however, ZO-1 levels are not influenced by p38MAPK/JNK.

  6. TRPV1 modulates morphine-induced conditioned place preference via p38 MAPK in the nucleus accumbens.

    Science.gov (United States)

    Hong, Sa-Ik; Nguyen, Thi-Lien; Ma, Shi-Xun; Kim, Hyoung-Chun; Lee, Seok-Yong; Jang, Choon-Gon

    2017-09-15

    Emerging evidence suggests that the transient receptor potential vanilloid type 1 channel (TRPV1) is a novel target for the treatment of drug addiction, such as cocaine and morphine. Previously we reported that TRPV1 inhibition reduced morphine reward in the dorsal striatum (DSt) of mice and morphine self-administration through a decrease in accumbal activity in rats. However, the role of TRPV1 on morphine-conditioned reward in addiction-related brain regions, such as the nucleus accumbens (NAc), has not been previously established. Here, we investigated the effects of TRPV1 on morphine conditioned place preference (CPP) and intracellular mechanisms of TRPV1 using Western blot analysis and immunohistochemistry (IHC) in morphine-administered mice. TRPV1 knockout mice did not exhibit morphine reward responses, and both i.p. and intra-NAc injections of SB366791, a selective TRPV1 antagonist, reduced morphine-induced CPP in wild-type mice. Furthermore, i.p. injection of SB203580, a selective p38 MAPK inhibitor, also dampened morphine-induced CPP. To determine the molecular mechanisms of the TRPV1/p38 MAPK pathway in morphine CPP, we investigated the expression of adenylyl cyclase type 1 (AC1) and phospho-p38 mitogen-activated protein kinase (MAPK) and nuclear factor-kappa B (NF-κB) in the NAc. Either SB366791 or SB203580 decreased the protein expression levels of phospho-p38 MAPK, phosphor-NF-κB, and AC1 in the NAc of morphine CPP mice. Taken together, our findings suggest that TRPV1 may modulate morphine-induced conditioned reward effects via the p38 MAPK signaling pathway in the NAc. Therefore, blockade of TRPV1 may provide a novel therapeutic approach for the prevention and treatment of opioid addiction. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Immunosuppressant MPA Modulates Tight Junction through Epigenetic Activation of MLCK/MLC-2 Pathway via p38MAPK

    Directory of Open Access Journals (Sweden)

    Niamat Khan

    2015-12-01

    Full Text Available Background: Mycophenolic acid (MPA is an important immunosuppressive drug (ISD prescribed to prevent graft rejection in the organ transplanted patients, however, its use is also associated with adverse side effects like sporadic gastrointestinal (GI disturbances. Recently, we reported the MPA induced tight junctions (TJs deregulation which involves MLCK/MLC-2 pathway. Here, we investigated the global histone acetylation as well as gene-specific chromatin signature of several genes associated with TJs regulation in Caco-2 cells after MPA treatment.Results: The epigenetic analysis shows that MPA treatment increases the global histone acetylation levels as well as the enrichment for transcriptional active histone modification mark (H3K4me3 at promoter regions of p38MAPK, ATF-2, MLCK, and MLC-2. In contrast, the promoter region of occludin was enriched for transcriptional repressive histone modification mark (H3K27me3 after MPA treatment. In line with the chromatin status, MPA treatment increased the expression of p38MAPK, ATF-2, MLCK, and MLC-2 both at transcriptional and translational level, while occludin expression was negatively influenced. Interestingly, the MPA induced gene expression changes and functional properties of Caco-2 cells could be blocked by the inhibition of p38MAPK using a chemical inhibitor (SB203580.Conclusions: Collectively, our results highlight that MPA disrupts the structure of TJs via p38MAPK-dependent activation of MLCK/MLC-2 pathway that results in decreased integrity of Caco-2 monolayer. These results led us to suggest that p38MAPK-mediated lose integrity of epithelial monolayer could be the possible cause of GI disturbance (barrier dysfunction in the intestine, leading to leaky style diarrhea observed in the organ-transplanted patients treated with MPA.

  8. Daily sesame oil supplementation attenuates local renin-angiotensin system via inhibiting MAPK activation and oxidative stress in cardiac hypertrophy.

    Science.gov (United States)

    Liu, Chuan-Teng; Liu, Ming-Yie

    2017-04-01

    The renin-angiotensin system (RAS) is involved in the development of left ventricular hypertrophy (LVH) by which increases cardiac morbidity and mortality. Activation of mitogen-activated protein kinases (MAPKs) and oxidative stress are important in RAS-mediated cardiac hypertrophy. Sesame oil, a potent antioxidant, attenuates hypertension-dependent LVH. We examined the protective role of sesame oil on RAS-mediated MAPK activation and oxidative stress in rats. We induced LVH using a hypertensive model by subcutaneously injecting deoxycorticosterone acetate (DOCA; 15 mg/ml/kg in mineral oil; twice weekly for 5 weeks) and supplementing with 1% sodium chloride drinking water (DOCA/salt) to uninephrectomized rats. Sesame oil was gavaged (0.5 or 1 ml/kg/day for 7 days) after 4 weeks of DOCA/salt treatment. Cardiac histopathology, RAS parameters, expression of MAPKs, reactive oxygen species and lipid peroxidation were assessed 24 h after the last dose of sesame oil. Sesame oil significantly decreased the size of cardiomyocytes and the levels of cardiac renin, angiotensin-converting enzyme and angiotensin II. In addition, sesame oil down-regulated the expression of angiotensin type 1 receptor, JNK and p38 MAPK and apoptosis signal regulating kinase 1, c-Fos and c-Jun in rats receiving DOCA/salt. Furthermore, the induction of nicotinamide adenine dinucleotide phosphate oxidase, superoxide anion and hydroxyl radical and lipid peroxidation by DOCA/salt were inhibited by sesame oil. Sesame oil modulates cardiac RAS to ameliorate LVH by inhibiting MAPK activation and lowering oxidative stress. Copyright © 2016 Elsevier Inc. All rights reserved.

  9. Activation of MAPK/ERK signaling by Burkholderia pseudomallei cycle inhibiting factor (Cif)

    Science.gov (United States)

    Ng, Mei Ying; Wang, Mei; Casey, Patrick J.; Gan, Yunn-Hwen; Hagen, Thilo

    2017-01-01

    Cycle inhibiting factors (Cifs) are virulence proteins secreted by the type III secretion system of some Gram-negative pathogenic bacteria including Burkholderia pseudomallei. Cif is known to function to deamidate Nedd8, leading to inhibition of Cullin E3 ubiquitin ligases (CRL) and consequently induction of cell cycle arrest. Here we show that Cif can function as a potent activator of MAPK/ERK signaling without significant activation of other signaling pathways downstream of receptor tyrosine kinases. Importantly, we found that the ability of Cif to activate ERK is dependent on its deamidase activity, but independent of Cullin E3 ligase inhibition. This suggests that apart from Nedd8, other cellular targets of Cif-dependent deamidation exist. We provide evidence that the mechanism involved in Cif-mediated ERK activation is dependent on recruitment of the Grb2-SOS1 complex to the plasma membrane. Further investigation revealed that Cif appears to modify the phosphorylation status of SOS1 in a region containing the CDC25-H and proline-rich domains. It is known that prolonged Cullin E3 ligase inhibition leads to cellular apoptosis. Therefore, we hypothesize that ERK activation is an important mechanism to counter the pro-apoptotic effects of Cif. Indeed, we show that Cif dependent ERK activation promotes phosphorylation of the proapoptotic protein Bim, thereby potentially conferring a pro-survival signal. In summary, we identified a novel deamidation-dependent mechanism of action of the B. pseudomallei virulence factor Cif/CHBP to activate MAPK/ERK signaling. Our study demonstrates that bacterial proteins such as Cif can serve as useful molecular tools to uncover novel aspects of mammalian signaling pathways. PMID:28166272

  10. Acidic environment activates inflammatory programs in fibroblasts via a cAMP-MAPK pathway.

    Science.gov (United States)

    Riemann, A; Ihling, A; Thomas, J; Schneider, B; Thews, O; Gekle, M

    2015-02-01

    The tissue micromilieu in disorders (inflammation, ischemia, tumor) often shows pronounced metabolic acidosis that may alter signaling and transcriptional activity in resident cells which can be of special importance for omnipresent fibroblasts. In the present study we investigated the impact of metabolic acidosis on rat fibroblasts with special emphasis on their role in inflammation by regulation of TNF-α, MCP-1, COX-2 and iNOS expression and the signaling pathways involved. Extracellular acidosis led to an enhanced expression of TNF-α, COX-2 and iNOS in parallel to an activation of p38 and ERK1/2 kinases that was not observed by sole intracellular acidosis. Accordingly, the protein amounts of TNF-α and COX-2 as well as the production of nitrate and nitrite were elevated. Acidosis-induced expression of COX-2 and iNOS depended on p38 kinase, but not on ERK1/2. In contrast acidosis-induced TNF-α expression was independent of both kinases. Although GPR4, GPR68 and GPR132 are expressed in fibroblasts, the involvement of these potential candidate pH sensors could be ruled out since no acidosis-induced elevation in intracellular cAMP or free calcium content was observed. Furthermore our data show that MAPK activation by an acidic micromilieu depends on Ser/Thr phosphatase activity, but not on the production of reactive oxygen species and is sensitive to cAMP antagonism by Rp-cAMPS. In conclusion, our results show that an acidic microenvironment induces a differential transcriptional program of pathological relevant genes in fibroblasts via the cAMP-phosphatase-MAPK pathway and thereby generates a parainflammatory situation that can result in tissue remodeling.

  11. Intraluminal pressure stimulates MAPK phosphorylation in arterioles: temporal dissociation from myogenic contractile response.

    Science.gov (United States)

    Spurrell, Brian E; Murphy, Timothy V; Hill, Michael A

    2003-10-01

    Members of the MAPK family of enzymes, p42/44 and p38, have been implicated in both the regulation of contractile function and growth responses in vascular smooth muscle. We determined whether such kinases are activated during the arteriolar myogenic response after increases in intraluminal pressure. Particular emphasis was placed on temporal aspects of activation to determine whether such phosphorylation events parallel the known time course for myogenic contraction. Experiments used single cannulated arterioles isolated from the cremaster muscle of rats with some vessels loaded with the fluorescent Ca2+-sensitive dye fura 2 (2 microM). The p42/44 inhibitor PD-98059 (50 microM) caused vasodilation but did not prevent pressure-induced myogenic constriction. The vasodilator response was accompanied by decreased smooth muscle intracellular Ca2+. Western blotting revealed a significant increase in the level of phosphorylation of p42/44 15 min after the application of a 30- to 100-mmHg pressure step. Phosphorylation of p42/44 was a late event that appeared to be temporally dissociated from contraction, which was complete within 1-5 min. EGF (80 nM) caused marked phosphorylation of p42/44 but only acted as a weak vasoconstrictor. The p38 inhibitor SB-203580 (10 microM) did not alter baseline diameter, nor did it prevent myogenic vasoconstriction. Consistent with these observations, SB-203580 did not cause a measurable change in intracellular Ca2+. The results demonstrate activation of the p42/44 class of MAPK resulting from increased transmural pressure. Such activation is, however, dissociated from the acute pressure-induced vasoconstrictor response in terms of time course and may represent the activation of compensatory, but parallel, pathways, including those related to growth and remodeling.

  12. Trefoil factors: Tumor progression markers and mitogens via EGFR/MAPK activation in cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Kanuengnuch Kosriwong; Trevelyan R Menheniott; Andrew S Giraud; Patcharee Jearanaikoon; Banchob Sripa; Temduang Limpaiboon

    2011-01-01

    AIM: To investigate trefoil factor (TFF ) gene copy number, mRNA and protein expression as potential biomarkers in cholangiocarcinoma (CCA). METHODS: TFF mRNA levels, gene copy number and protein expression were determined respectively by quantitative reverse transcription polymerase chain reaction (PCR), quantitative PCR and immunohistochemistry in bile duct epithelium biopsies collected from individuals with CCA, precancerous bile duct dysplasia and from disease-free controls. The functional impact of recombinant human (rh)TFF2 peptide treatment on proliferation and epidermal growth factor receptor (EGFR)/mitogenactivated protein kinase (MAPK) signaling was assessed in the CCA cell line, KMBC, by viable cell counting and immunoblotting, respectively. RESULTS: TFF1 , TFF2 and TFF3 mRNA expression was significantly increased in CCA tissue compared to disease-free controls, and was unrelated to gene copy number. TFF1 immunoreactivity was strongly increased in both dysplasia and CCA, whereas TFF2 immunoreactivity was increased only in CCA compared to diseasefree controls. By contrast, TFF3 immunoreactivity was moderately decreased in dysplasia and further decreased in CCA. Kaplan-Meier analysis found no association of TFF mRNA, protein and copy number with age, gender, histological subtype, and patient survival time. Treatment of KMBC cells with rhTFF2 stimulated proliferation, triggered phosphorylation of EGFR and downstream extracellular signal related kinase (ERK), whereas co-incubation with the EGFR tyrosine kinase inhibitor, PD153035, blocked rhTFF2-dependent proliferation and EGFR/ERK responses. CONCLUSION: TFF mRNA/protein expression is indicative of CCA tumor progression, but not predictive for histological sub-type or survival time. TFF2 is mitogenic in CCA via EGFR/MAPK activation.

  13. p38 MAPK inhibition reduces diabetes-induced impairment of wound healing

    Directory of Open Access Journals (Sweden)

    Satyanarayana Medicherla

    2009-06-01

    Full Text Available Satyanarayana Medicherla1, Scott Wadsworth2, Breda Cullen3, Derek Silcock3, Jing Y Ma1, Ruban Mangadu1, Irene Kerr1, Sarvajit Chakravarty1, Gregory L Luedtke1, Sundeep Dugar1, Andrew A Protter1, Linda S Higgins11Scios Inc., Fremont, CA, USA; 2Center for Biomaterials and Advanced Technologies, Somerville, NJ, USA; 3Johnson & Johnson Wound Management, Gargrave, UKAbstract: In healthy tissue, a wound initiates an inflammatory response characterized by the presence of a hematoma, infiltration of inflammatory cells into the wound and, eventually, wound healing. In pathological conditions like diabetes mellitus, wound healing is impaired by the presence of chronic nonresolving inflammation. p38 mitogen-activated protein kinase (MAPK inhibitors have demonstrated anti-inflammatory effects, primarily by inhibiting the expression of inflammatory cytokines and regulating cellular traffic into wounds. The db/db mouse model of type 2 diabetes was used to characterize the time course of expression of activated p38 during impaired wound healing. The p38α-selective inhibitor, SCIO-469, was applied topically and effects on p38 activation and on wound healing were evaluated. A topical dressing used clinically, PromogranTM, was used as a comparator. In this study, we established that p38 is phosphorylated on Days 1 to 7 post-wounding in db/db mice. Further, we demonstrated that SCIO-469, at a dose of 10 µg/wound, had a positive effect on wound contraction, granulation tissue formation, and re-epithelialization, and also increased wound maturity during healing. These effects were similar to or greater than those observed with PromogranTM. These results suggest a novel approach to prophylactic and therapeutic management of chronic wounds associated with diabetes or other conditions in which healing is impaired.Keywords: p38 MAPK ihibition, diabetic wound healing, db/db mouse, nonresolving healing, PromogranTM

  14. Phosphorylation of the Polarity Protein BASL Differentiates Asymmetric Cell Fate through MAPKs and SPCH.

    Science.gov (United States)

    Zhang, Ying; Guo, Xiaoyu; Dong, Juan

    2016-11-07

    Cell polarization is commonly used for the regulation of stem cell asymmetric division in both animals and plants. Stomatal development in Arabidopsis, a process that produces breathing pores in the epidermis, requires asymmetric cell division to differentiate highly specialized guard cells while maintaining a stem cell population [1, 2]. The BREAKING OF ASYMMETRY IN THE STOMATAL LINEAGE (BASL) protein exhibits a polarized localization pattern in the cell and is required for differential cell fates resulting from asymmetric cell division [3]. The polarization of BASL is made possible by a positive feedback loop with a canonical mitogen-activated protein kinase (MAPK) pathway that recruits the MAPKK kinase YODA (YDA) and MAPK 6 (MPK6) to the cortical polarity site [4]. Here, we study BASL intracellular dynamics and show that the membrane-associated BASL is slowly replenished at the cortical polarity site and that the mobility is tightly linked to its phosphorylation status. Because BASL polarity is only exhibited by one daughter cell after an asymmetric cell division, we study how BASL differentially functions in the two daughter cells. The YDA MAPK cascade transduces upstream ligand-receptor signaling [5-13] to the transcription factor SPEECHLESS (SPCH), which controls stomatal initiation and is directly suppressed by MPK3/6-mediated phosphorylation [14, 15]. We show that BASL polarization leads to elevated nuclear MPK6 signaling and lowered SPCH abundance in one of the two daughter cells. Therefore, two daughter cells are differentiated by BASL polarity-mediated differential suppression of SPCH, which may provide developmental plasticity in plant stem cell asymmetric cell division (ACD).

  15. MAPK Activation Is Essential for Waddlia chondrophila Induced CXCL8 Expression in Human Epithelial Cells.

    Directory of Open Access Journals (Sweden)

    Skye Storrie

    Full Text Available Waddlia chondrophila (W. chondrophila is an emerging agent of respiratory and reproductive disease in humans and cattle. The organism is a member of the order Chlamydiales, and shares many similarities at the genome level and in growth studies with other well-characterised zoonotic chlamydial agents, such as Chlamydia abortus (C. abortus. The current study investigated the growth characteristics and innate immune responses of human and ruminant epithelial cells in response to infection with W. chondrophila.Human epithelial cells (HEp2 were infected with W. chondrophila for 24h. CXCL8 release was significantly elevated in each of the cell lines by active-infection with live W. chondrophila, but not by exposure to UV-killed organisms. Inhibition of either p38 or p42/44 MAPK significantly inhibited the stimulation of CXCL8 release in each of the cell lines. To determine the pattern recognition receptor through which CXCL8 release was stimulated, wild-type HEK293 cells which express no TLR2, TLR4, NOD2 and only negligible NOD1 were infected with live organisms. A significant increase in CXCL8 was observed.W. chondrophila actively infects and replicates within both human and ruminant epithelial cells stimulating CXCL8 release. Release of CXCL8 is significantly inhibited by inhibition of either p38 or p42/44 MAPK indicating a role for this pathway in the innate immune response to W. chondrophila infection. W. chondrophila stimulation of CXCL8 secretion in HEK293 cells indicates that TLR2, TLR4, NOD2 and NOD1 receptors are not essential to the innate immune response to infection.

  16. Interleukin-35 Inhibits Endothelial Cell Activation by Suppressing MAPK-AP-1 Pathway.

    Science.gov (United States)

    Sha, Xiaojin; Meng, Shu; Li, Xinyuan; Xi, Hang; Maddaloni, Massimo; Pascual, David W; Shan, Huimin; Jiang, Xiaohua; Wang, Hong; Yang, Xiao-feng

    2015-07-31

    Vascular response is an essential pathological mechanism underlying various inflammatory diseases. This study determines whether IL-35, a novel responsive anti-inflammatory cytokine, inhibits vascular response in acute inflammation. Using a mouse model of LPS-induced acute inflammation and plasma samples from sepsis patients, we found that IL-35 was induced in the plasma of mice after LPS injection as well as in the plasma of sepsis patients. In addition, IL-35 decreased LPS-induced proinflammatory cytokines and chemokines in the plasma of mice. Furthermore, IL-35 inhibited leukocyte adhesion to the endothelium in the vessels of lung and cremaster muscle and decreased the numbers of inflammatory cells in bronchoalveolar lavage fluid. Mechanistically, IL-35 inhibited the LPS-induced up-regulation of endothelial cell (EC) adhesion molecule VCAM-1 through IL-35 receptors gp130 and IL-12Rβ2 via inhibition of the MAPK-activator protein-1 (AP-1) signaling pathway. We also found that IL-27, which shares the EBI3 subunit with IL-35, promoted LPS-induced VCAM-1 in human aortic ECs and that EBI3-deficient mice had similar vascular response to LPS when compared with that of WT mice. These results demonstrated for the first time that inflammation-induced IL-35 inhibits LPS-induced EC activation by suppressing MAPK-AP1-mediated VCAM-1 expression and attenuates LPS-induced secretion of proinflammatory cytokines/chemokines. Our results provide insight into the control of vascular inflammation by IL-35 and suggest that IL-35 is an attractive novel therapeutic reagent for sepsis and cardiovascular diseases.

  17. Activation of MAPK/ERK signaling by Burkholderia pseudomallei cycle inhibiting factor (Cif).

    Science.gov (United States)

    Ng, Mei Ying; Wang, Mei; Casey, Patrick J; Gan, Yunn-Hwen; Hagen, Thilo

    2017-01-01

    Cycle inhibiting factors (Cifs) are virulence proteins secreted by the type III secretion system of some Gram-negative pathogenic bacteria including Burkholderia pseudomallei. Cif is known to function to deamidate Nedd8, leading to inhibition of Cullin E3 ubiquitin ligases (CRL) and consequently induction of cell cycle arrest. Here we show that Cif can function as a potent activator of MAPK/ERK signaling without significant activation of other signaling pathways downstream of receptor tyrosine kinases. Importantly, we found that the ability of Cif to activate ERK is dependent on its deamidase activity, but independent of Cullin E3 ligase inhibition. This suggests that apart from Nedd8, other cellular targets of Cif-dependent deamidation exist. We provide evidence that the mechanism involved in Cif-mediated ERK activation is dependent on recruitment of the Grb2-SOS1 complex to the plasma membrane. Further investigation revealed that Cif appears to modify the phosphorylation status of SOS1 in a region containing the CDC25-H and proline-rich domains. It is known that prolonged Cullin E3 ligase inhibition leads to cellular apoptosis. Therefore, we hypothesize that ERK activation is an important mechanism to counter the pro-apoptotic effects of Cif. Indeed, we show that Cif dependent ERK activation promotes phosphorylation of the proapoptotic protein Bim, thereby potentially conferring a pro-survival signal. In summary, we identified a novel deamidation-dependent mechanism of action of the B. pseudomallei virulence factor Cif/CHBP to activate MAPK/ERK signaling. Our study demonstrates that bacterial proteins such as Cif can serve as useful molecular tools to uncover novel aspects of mammalian signaling pathways.

  18. PLA2G16 promotes osteosarcoma metastasis and drug resistance via the MAPK pathway

    Science.gov (United States)

    Li, Lin; Liang, Shoulei; Wasylishen, Amanda R.; Zhang, Yanqin; Yang, Xueli; Zhou, Bingzheng; Shan, Luling; Han, Xiuxin; Mu, Tianyang; Wang, Guowen; Xiong, Shunbin

    2016-01-01

    The prognosis of metastatic osteosarcoma is dismal and a better understanding of the mechanisms underlying disease progression is essential to improve treatment options and patient outcomes. We previously demonstrated Pla2g16 overexpression in mouse osteosarcoma contributes to metastasis phenotypes and increased expression of PLA2G16 is associated with metastasis and poor prognosis in human tumors. To further examine the mechanisms through which PLA2G16 contributes to human osteosarcoma metastasis and explore the potential of PLA2G16 as a therapeutic target in osteosarcoma, we generated a panel of human osteosarcoma cell lines expressing different levels of PLA2G16. The functional analyses of these cell lines demonstrated high levels of PLA2G16 expression increased osteosarcoma cell migration, invasion, clonogenic survival, and anchorage-independent colony formation. Importantly, this activity was dependent on the phospholipase activity of PLA2G16. Additionally, PLA2G16 overexpression decreased the sensitivity of cells to a panel of chemotherapeutic agents. Analysis of downstream pathways revealed the pro-metastasis functions of PLA2G16 were mediated through the MAPK pathway, as knockdown of PLA2G16 decreased ERK1/2 phosphorylation and pharmacological inhibition of MEK significantly repressed PLA2G16 mediated cell migration and clonogenic survival. Furthermore, PLA2G16 overexpression promoted xenograft tumor growth in vivo, and these tumors exhibit increased ERK1/2 phosphorylation. Lastly, the expression of PLA2G16 is strongly correlated with the increased ERK1/2 phosphorylation in human osteosarcoma samples, and the combined lesions are associated with reduced overall and metastasis-free survival. Collectively, these results demonstrate increased PLA2G16 expression activates the MAPK pathway to enhance osteosarcoma metastasis and may be a novel therapeutic target for these cancers. PMID:26933804

  19. Quantitative cell signalling analysis reveals down-regulation of MAPK pathway activation in colorectal cancer.

    LENUS (Irish Health Repository)

    Gulmann, Christian

    2009-08-01

    Mitogen-activated protein kinases (MAPK) are considered to play significant roles in colonic carcinogenesis and kinase inhibitor therapy has been proposed as a potential tool in the treatment of this disease. Reverse-phase microarray assays using phospho-specific antibodies can directly measure levels of phosphorylated protein isoforms. In the current study, samples from 35 cases of untreated colorectal cancer colectomies were laser capture-microdissected to isolate epithelium and stroma from cancer as well as normal (i.e. uninvolved) mucosa. Lysates generated from these four tissue types were spotted onto reverse-phase protein microarrays and probed with a panel of antibodies to ERK, p-ERK, p38, p-p38, p-JNK, MEK and p-MEK. Whereas total protein levels were unchanged, or slightly elevated (p38, p = 0.0025) in cancers, activated isoforms, including p-ERK, p-p38 and p-JNK, were decreased two- to four-fold in cancers compared with uninvolved mucosa (p < 0.0023 in all cases except for p-JNK in epithelium, where decrement was non-significant). This was backed up by western blotting. Dukes\\' stage B and C cancers displayed lower p-ERK and p-p38 expression than Dukes\\' stage A cancers, although this was not statistically significant. It is concluded that MAPK activity may be down-regulated in colorectal cancer and that further exploration of inhibitory therapy in this system should be carefully evaluated if this finding is confirmed in larger series.

  20. Novel mechanisms and therapeutic approaches in melanoma: targeting the MAPK pathway.

    Science.gov (United States)

    Grimaldi, Antonio Maria; Simeone, Ester; Festino, Lucia; Vanella, Vito; Palla, Marco; Ascierto, Paolo Antonio

    2015-06-01

    The development of novel treatments that selectively inhibit the RAS-RAF-MAPK pathway represents a milestone in the history of melanoma treatment. BRAF mutations occur in approximately 45% of cutaneous melanomas, while mutations in NRAS occur in 15-25%. Vemurafenib was the first BRAF inhibitor to be approved in 2011, based on the results of a phase III trial (BRIM-3) that showed higher progression-free survival and overall survival compared with dacarbazine chemotherapy in metastatic BRAF-mutated melanoma. Dabrafenib, another BRAF inhibitor, has shown similar results and was approved in 2013. Preclinical studies suggested that another novel group of agents, the MEK inhibitors, showed stronger inhibition of both mutated BRAF and NRAS cell cultures than vemurafenib. Trametinib was the first MEK inhibitor approved in 2014, both as a single agent and in combination with dabrafenib for the treatment of advanced BRAF-mutated melanoma. Other MEK inhibitors are also in development. Concomitant inhibition of both MEK and BRAF has shown more durable and greater tumor response than BRAF monotherapy, by overcoming the multiple genetic mechanisms of escape. Combined therapy prevents the development of acquired resistance as well as decreasing cutaneous toxicity secondary to paradoxical activation of the MAPK pathway induced by BRAF inhibitors. Various combinations of BRAF and MEK inhibitors have shown promising results. Moreover, triple combination therapies involved other agents with novel mechanisms of action are also being evaluated. These and other combination strategies involving immunotherapies and targeted therapies offer the hope of improving outcomes beyond those already achieved with anti-BRAF treatments.

  1. MAGI1 inhibits migration and invasion via blocking MAPK/ERK signaling pathway in gastric cancer

    Science.gov (United States)

    Jia, Shuqin; Lu, Jiajia; Qu, Tingting; Feng, Yi; Wang, Xiaohong; Liu, Caixia; Ji, Jiafu

    2017-01-01

    Objective To explore the association of membrane-associated guanylate kinase inverted 1 (MAGI1) with gastric cancer (GC) and the related molecular mechanisms. Methods The reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry (IHC) were utilized to measure the MAGI1 expression level in GC tissues. Quantitative real-time PCR and Western blotting were used to ensure the MAGI1 expression in GC cell lines. Small hairpin RNA (shRNA) was applied for knockdown of endogenous MAGI1 in GC cells. MTT assay and colony formation assay, scratch wounding migration assay and transwell chamber migration assay, as well as transwell chamber invasion assay were employed respectively to investigate the GC cell proliferation, migration and invasion in MAGI1-knockdown and control GC cells. The potential molecular mechanism mediated by MAGI1 was studied using Western blotting and RT- PCR. Results RT-PCR and IHC verified MAGI1 was frequently expressed in matched adjacent noncancerous mucosa compared with GC tissues and the expression of MAGI1 was related to clinical pathological parameters. Functional assays indicated that MAGI1 knockdown significantly promoted GC cell migration and invasion. Further mechanism investigation demonstrated that one pathway of MAGI1 inhibiting migration and invasion was mainly by altering the expression of matrix metalloproteinases (MMPs) and epithelial-mesenchymal transition (EMT)-related molecules via inhibiting MAPK/ERK signaling pathway. Conclusions MAGI1 was associated with GC clinical pathological parameters and acted as a tumor suppressor via inhibiting of MAPK/ERK signaling pathway in GC. PMID:28373751

  2. Neospora caninum Activates p38 MAPK as an Evasion Mechanism against Innate Immunity

    Science.gov (United States)

    Mota, Caroline M.; Oliveira, Ana C. M.; Davoli-Ferreira, Marcela; Silva, Murilo V.; Santiago, Fernanda M.; Nadipuram, Santhosh M.; Vashisht, Ajay A.; Wohlschlegel, James A.; Bradley, Peter J.; Silva, João S.; Mineo, José R.; Mineo, Tiago W. P.

    2016-01-01

    Due to the high prevalence and economic impact of neosporosis, the development of safe and effective vaccines and therapies against this parasite has been a priority in the field and is crucial to limit horizontal and vertical transmission in natural hosts. Limited data is available regarding factors that regulate the immune response against this parasite and such knowledge is essential in order to understand Neospora caninum induced pathogenesis. Mitogen-activated protein kinases (MAPKs) govern diverse cellular processes, including growth, differentiation, apoptosis, and immune-mediated responses. In that sense, our goal was to understand the role of MAPKs during the infection by N. caninum. We found that p38 phosphorylation was quickly triggered in macrophages stimulated by live tachyzoites and antigen extracts, while its chemical inhibition resulted in upregulation of IL-12p40 production and augmented B7/MHC expression. In vivo blockade of p38 resulted in an amplified production of cytokines, which preceded a reduction in latent parasite burden and enhanced survival against the infection. Additionally, the experiments indicate that the p38 activation is induced by a mechanism that depends on GPCR, PI3K and AKT signaling pathways, and that the phenomena here observed is distinct that those induced by Toxoplasma gondii’s GRA24 protein. Altogether, these results showed that N. caninum manipulates p38 phosphorylation in its favor, in order to downregulate the host’s innate immune responses. Additionally, those results infer that active interference in this signaling pathway may be useful for the development of a new therapeutic strategy against neosporosis. PMID:27679624

  3. Cholesterol Enhances Colorectal Cancer Progression via ROS Elevation and MAPK Signaling Pathway Activation

    Directory of Open Access Journals (Sweden)

    Caihua Wang

    2017-06-01

    Full Text Available Background/Aims: Elevated serum cholesterol levels were linked to a higher risk of colorectal adenoma and colorectal cancer (CRC, while the effect of cholesterol on CRC metastasis has not been widely studied. Methods: CRC patients were enrolled to evaluate the association between low-density lipoprotein cholesterol (LDL and CRC metastases, and LDL receptor (LDLR level of the CRC tissue was assessed by immunohistochemistry. The effects of LDL on cell proliferation, migration and stemness were assessed in CRC cells in vitro, and the effects of high fat diet (HFD on tumor growth and intestinal tumorigenicity were investigated in vivo. ROS assays, gene expression array analysis and western blot were used to explore the mechanisms o