Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.12.2 (MEK)
18,161 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Granulocyte macrophage-colony-stimulating factor (GM-CSF), released from alveolar macrophages (AM), is an important regulator of eosinophil, T cell, and macrophage function and survival. We determined the mechanisms of GM-CSF regulation in AM from normal volunteers activated by lipopolysaccharide (LPS) by examining the role of nuclear factor-kappaB (NF-kappaB), and of p38 mitogen-activated protein (MAP) kinase and MAP kinase kinase (MKK-1). PD 098059 (10 microM), an inhibitor of upstream activator of MKK-1, inhibited GM-CSF expression, but the expression of GM-CSF was not inhibited by SB 203580 (10 microM), an inhibitor of p38-MAP kinase. Phosphorylation of extracellular signal-regulated kinase-1 (ERK-1), ERK-2, and p38 MAP kinase by LPS were demonstrated on Western blot analysis. LPS increased NF-kappaB:DNA binding as examined by electrophoretic mobility shift assay, but this was not suppressed by PD 098059 or by SB 203580. LPS induced an increase in NF-kappaB activation as examined by p50 translocation assay without suppression by PD 098059 or by SB 203580. SN50 (100 microM), an inhibitor of NF-kappaB translocation and the specific IKK-2-Inhibitor (AS602868; 10 microM), also prevented GM-CSF expression and release induced by LPS, indicating that GM-CSF release is NF-kappaB-dependent. PD 098059, but not SB 203580, inhibited LPS-induced histone acetyltransferase (HAT) activity, indicating chromatin modification. Furthermore, AS602868 and SN 50 suppressed LPS-induced HAT activity. TSA (10 ng/ml), an inhibitor of histone deacetylase (HDAC), reversed the inhibitory effect of PD 098059, SB 203580, SN 50 and AS602868 on GM-CSF release. GM-CSF expression and release in AM is controlled by NF-kappaB activation, and this is modulated by phosphorylation of MKK-1 and p38 MAP kinase acting on histone acetylation.
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PMID:Mitogen-activated protein kinase modulation of nuclear factor-kappaB-induced granulocyte macrophage-colony-stimulating factor release from human alveolar macrophages. 1287 51

Our previous studies have shown that the cell proliferation rate, mRNA levels of p450scc, p450c17, and 3betaHSD, and secretion of cortisol were significantly increased in human adrenocortical cells stably transfected with mutated K-ras expression plasmid "pK568MRSV" after being inducted with IPTG. In addition, the increased level was a time-dependent manner. However, the levels of p450, p450scc, p450c17, 3betaHSD, cortisol, and cell proliferation rate were inhibited by a MEK phospholation inhibitor, PD098059. The above results prove that mutated K-ras oncogene is able to regulate tumorigenesis and steroidogenesis through a Ras-RAF-MEK-MAPK signal transduction pathway. The aim of this study was to investigate regulated factors in this pathway and also examine whether the other signal transduction pathways or other moles involved in tumorigenesis or steroidogenesis. In the first year, we analyzed gene profiles of mutant K-ras-transfected adrenocortical cells by DNA microarray to determine the gene expression related to cell cycle, signal transduction, apoptosis, tumorigenesis, steroidogenesis, and other expressed sequence tag. After being affected by the K-ras mutant, gene expression was significantly increased in some upregulated genes. Human zinc-finger protein 22 increased by 28.5 times, Osteopontin increased by 5.8 times, LIM domain Kinase 2 (LIMK2) increased by 3.3 times, Homo sapiens dual-specificity tyrosine-(Y)-phosphorylation regulated Kinase 2 (DYRK2) increased by 2.2 times, and human syntaxin 3 increased by two times. On the other hand, significant decreases in gene expression were also observed in some downregulated genes. Retinoblastoma binding protein 1 (RBBP1) decreased by four times, Homo sapiens craniofacial development protein 1 (CFDP1) decreased by 2.4 times, DAP Kinase-related apoptosis-inducing protein Kinase 1 (DRAK1) decreased by 2.3 times, SKI-interacting protein (SKIP) decreased by 2.2 times, and human poly(A)-Binding protein (PABP) decreased by 2.1 times. In all significant differentially expressed genes, preliminary analysis by bioinformatics revealed that after induced K-ras mutant expression by isopropyl thiogalctoside (IPTG), the downregulation of RBBP1 gene was most correlated to cell proliferation. RBBP1 can bind with RB/E2F to form a mSIN3-HDAC complex, which induces cell cycle arrest in the G1/G0 stage by repressing transcription of E2F-regulated genes. The result of a Northern blot showed that RBBP1 were inhibited after an induction of IPTG for 36 h. Another Northern blot analysis proved that mRNA levels of cyclin D1 and c-myc increased in proportion to K-ras expression. Finally, Western blot was carried out, and the results showed that phosphorylated pRB also increased. Taken together, we infer that the mutant K-ras oncogene promoted the cells to proceed to the G1/S stage by the inhibiting the formation of RB/RBBP1-dependent repressor complex from binding with the SIN3-HDAC complex, which resulted in the acetylation of histone to active transcription of E2F-regulated genes. However, the roles of the other differentially expressed genes involved in cell proliferation, cell morphologic change, tumorigenesis, or steroidogenesis still need further investigation.
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PMID:Retinoblastoma protein (pRB) was significantly phosphorylated through a Ras-to-MAPK pathway in mutant K-ras stably transfected human adrenocortical cells. 1461 87

Transcriptional activation of eukaryotic genes depends on the precise and ordered recruitment of activators, chromatin modifiers/remodelers, coactivators, and general transcription factors to the promoters of target genes. Using the human matrix metalloproteinase 9 (MMP-9) gene as a model system, we investigated the sequential assembly and dynamic formation of transcription complexes on a human promoter under the influence of mitogen signaling. We find that, coincident with activation of the MMP-9 gene, activators, chromatin remodeling complexes, and coactivators are recruited to the preassembled MMP-9 promoter in a stepwise and coordinated order, which is dependent on activation of MEK-1/extracellular signal-regulated kinase and NF-kappa B signaling pathways. Conversely, corepressor complexes are released from the MMP-9 promoter after transcriptional activation. Histone modifications shift from repressive to permissive modifications concurrent with activation of the MMP-9 gene. Chromatin remodeling induced by Brg-1 is required for MMP-9 gene transcription, which is concomitant with initiation of transcription. Therefore, coordination of cell signaling, chromatin remodeling, histone modifications, and stepwise recruitment of transcription regulators is critical to precisely regulate MMP-9 gene transcription in a temporally and spatially dependent manner. Given the important role of MMP-9 in both normal development and pathological conditions, understanding MMP-9 gene regulation is of great relevance.
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PMID:Coordination of cell signaling, chromatin remodeling, histone modifications, and regulator recruitment in human matrix metalloproteinase 9 gene transcription. 1516 10

The relationship between breast cancer-associated fatty acid synthase (FAS; oncogenic antigen-519) and chemotherapy-induced cell damage has not been studied. We examined the ability of C75, a synthetic slow-binding inhibitor of FAS activity, to modulate the cytotoxic activity of the microtubule-interfering agent Taxol (paclitaxel) in SK-Br3, MDA-MB-231, MCF-7 and multidrug-resistant MDR-1 (P-Glycoprotein)-overexpressing MCF-7/AdrR breast cancer cells. When the combination of C75 with Taxol in either concurrent (C75 + Taxol 24 hr) or sequential (C75 24 hr --> Taxol 24 hr) schedules were tested for synergism, addition or antagonism using the isobologram and the median-effect plot analyses, co-exposure of C75 and Taxol mostly demonstrated synergistic effects, whereas sequential exposure to C75 followed by Taxol mainly showed additive or antagonistic interactions. Because the nature of the cytotoxic interactions was definitely schedule-dependent in MCF-7 cells, we next evaluated the effects of C75 on Taxol-induced apoptosis as well as Taxol-activated cell death and cell survival-signaling pathways in this breast cancer cell model. An ELISA for histone-associated DNA fragments demonstrated that C75 and Taxol co-exposure caused a synergistic enhancement of apoptotic cell death, whereas C75 pre-treatment did not enhance the apoptosis-inducing activity of Taxol. Co-exposure to C75 and Taxol induced a remarkable nuclear accumulation of activated p38 mitogen-activated protein kinase (p38 MAPK), which was accompanied by a synergistic nuclear accumulation of the p53 tumor-suppressor protein that was phosphorylated at Ser46, a p38 MAPK-regulated pro-apoptotic modification of p53. As single agents, FAS blocker C75 and Taxol induced a significant stimulation of the proliferation and cell survival mitogen-activated protein kinase extracellular signal-regulated kinase (ERK1/ERK2 MAPK) activity, whereas, in combination, they interfered with ERK1/ERK2 activation. Moreover, the combined treatment of C75 and Taxol inactivated the anti-apoptotic AKT (protein kinase B) kinase more than either agent alone, as evidenced by a synergistic down-regulation of AKT phosphorylation at its activating site Ser(473) without affecting AKT protein levels. To rule out a role for non-FAS C75-mediated effects, we finally used the potent and highly sequence-specific mechanism of RNA interference (RNAi) to block FAS-dependent signaling. Importantly, SK-Br3 and multi-drug resistant MCF-7/AdrR cells transiently transfected with sequence-specific double-stranded RNA oligonucleotides targeting FAS gene demonstrated hypersensitivity to Taxol-induced apoptotic cell death. Our findings establish for the first time that FAS blockade augments the cytotoxicity of anti-mitotic drug Taxol against breast cancer cells and that this chemosensitizing effect is schedule-dependent. We suggest that the alternate activation of both the pro-apoptotic p38 MAPK-p53 signaling and the cytoprotective MEK1/2 --> ERK1/2 cascade, as well as the inactivation of the anti-apoptotic AKT activity may explain, at least in part, the sequence-dependent enhancement of Taxol-induced cytotoxicity and apoptosis that follows inhibition of FAS activity in breast cancer cells. If chemically stable FAS inhibitors demonstrate systemic anticancer effects of FAS inhibition in vivo, these findings may render FAS as a valuable molecular target to enhance the efficacy of taxanes-based chemotherapy in human breast cancer.
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PMID:Pharmacological and small interference RNA-mediated inhibition of breast cancer-associated fatty acid synthase (oncogenic antigen-519) synergistically enhances Taxol (paclitaxel)-induced cytotoxicity. 1565

Transcription of the prolactin gene is dynamically controlled by positive and negative hormone signals that target the regulatory promoter region. Based on the inducibility of prolactin gene expression by inhibitors of histone deacetylases (HDACs), we examined the role of histone acetylation at the genomic prolactin promoter as a late step in transcriptional regulation. Chromatin immunoprecipitation analysis of GH4 cells revealed elevated levels of acetylated histones in the promoter and enhancer regions of the gene, compared with downstream intron sequences. 17beta-Estradiol stimulated histone H4 acetylation in the promoter region by 2- to 3-fold within 30 min. Dopamine inhibited histone H4 acetylation by 2-fold in 30 min, an effect mimicked by the MAPK kinase (MEK1) inhibitor U0126. In contrast, the synthetic glucocorticoid dexamethasone, which inhibits prolactin transcription, failed to alter histone acetylation over the same time frame. Association of transcription activator Pit-1 with the prolactin promoter was unchanged by hormone treatment. However, in response to dopamine, histone deacetylase HDAC2 and corepressor mSin3A were rapidly recruited to the prolactin promoter, and association was sustained above basal levels over a 1-h period. Consistent with this corepressor function, depletion of endogenous mSin3A by small interfering RNA was sufficient to enhance prolactin gene expression by 70%, comparable to the induction by the HDAC inhibitor, trichostatin A. These studies demonstrate that dopamine D2 receptor activation and inhibition of MAPK (ERK1/2) signaling lead to rapid deacetylation of histones at the genomic prolactin promoter. Recruitment of specific HDAC/ corepressor complexes may be an important mechanism for repression of target gene transcription by Gi/o-coupled receptors.
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PMID:Epigenetic mechanisms in the dopamine D2 receptor-dependent inhibition of the prolactin gene. 1573 Nov 70

We established that such stress agent as antibiotic anisomycin was capable at very low concentrations to activate transcription of immediate-early c-fos gene, which generally was under negative control in fibroblasts transformed by E1A and cHa-ras oncogenes. Activation of c-fos gene is short-term reaching a maximum in 1 hour after anisomycin action then its transcription level declines that is typical for immediate-early genes. Transcription of two other examined immediate-early genes, c-jun and Egr-1, is at high enough level in this cell line, however its additional activation does take place under anisomycin action too. It is shown that in E1A + ras transformants, anisomycin induces MEK/ERK and JNK MAP kinase pathways ofsignal transduction whereas p38 kinase cascade is not activated. Using specific inhibitors of various MAP kinase cascades it has been shown for E1A + ras transformed cells that the anisomycin-induced transcription of c-fos gene is mainly adjusted through MEK/ERK kinase pathway while high level of c-jun gene transcription is supported by activity of JNK kinases. Even at higher concentration, anisomycin does not lead finally to accumulation of acetylated forms of core histones, in particular H3 histone therefore the transcriptional activation of immediate-early genes induced by anisomycin is not bound apparently with decompactization of chromatin structure in the region of promoters of these genes.
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PMID:[Effect of anisomycin on activation of early response genes c-fos, c-jun, Egr-1 in cells transformed by E1A and cHa-ras oncogenes]. 1577 51

Ethanol treatment increases gene expression in the liver through mechanisms that are not clearly understood. Histone acetylation has been shown to induce transcriptional activation. We have investigated the characteristics and mechanisms of ethanol-induced histone H3 acetylation in rat hepatocytes. Immunocytochemical and immunoblot analysis revealed that ethanol treatment significantly increased H3 acetylation at Lys9 with negligible effects at Lys14, -18, and -23. Acute in vivo administration of alcohol in rats produced the same results as in vitro observations. Nuclear extracts from ethanol-treated hepatocytes increased acetylation in H3 peptide to a greater extent than extracts from untreated cells, suggesting that ethanol either increased the expression level or the specific activity of histone acetyltransferases (HAT). Use of different H3 peptides indicated that ethanol selectively modulated HAT(s) targeting H3-Lys9. Treatment with acetate, an ethanol metabolite, also increased acetylation of H3-Lys9 and modulated HAT(s) in the same manner as ethanol, suggesting that acetate mediates the ethanol-induced effect on HAT. Inhibitors of MEK (U0126) and JNK (SP600125), but not p38 MAPK inhibitor (SB203580), suppressed ethanol-induced H3 acetylation. However, U0126 and SP600125 did not significantly affect ethanol-induced effect on HAT, suggesting that ERK and JNK regulate histone acetylation through a separate pathway(s) that does not involve modulation of HAT. Chromatin immunoprecipitation assay demonstrated that ethanol treatment increased the association of the class I alcohol dehydrogenase (ADH I) gene with acetylated H3-Lys9. These data provide first evidence that ethanol increases acetylation of H3-Lys9 through modulation of HAT(s) and that histone acetylation may underlie the mechanism for ethanol-induced ADH I gene expression.
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PMID:Involvement of histone acetyltransferase (HAT) in ethanol-induced acetylation of histone H3 in hepatocytes: potential mechanism for gene expression. 1608 63

Epigenetic histone modifications contribute to the regulation of eukaryotic gene transcription. The role of epigenetic regulation in immunity to intracellular pathogens is poorly understood. We tested the hypothesis that epigenetic histone modifications influence cytokine expression by intracellular bacteria. Intracellular Listeria monocytogenes, but not noninvasive Listeria innocua, induced release of distinct CC and CXC chemokines, as well as Th1 and Th2 cytokines and growth factors by endothelial cells. Cytokine expression was in part dependent on p38 MAPK and MEK1. We analyzed global histone modification and modifications in detail at the gene promoter of IL-8, which depended on both kinase pathways, and of IFN-gamma, which was not blocked by kinase inhibition. Intracellular Listeria induced time-dependent acetylation (lysine 8) of histone H4 and phosphorylation/acetylation (serine 10/lysine 14) of histone H3 globally and at the il8 promoter in HUVEC, as well as recruitment of the histone acetylase CREB-binding protein. Inhibitors of p38 MAPK and MEK1 reduced lysine 8 acetylation of histone H4 and serine 10/lysine 14 phosphorylation/acetylation of histone H3 in Listeria-infected endothelial cells and disappearance of histone deacetylase 1 at the il8 promoter in HUVEC. In contrast, IFN-gamma gene transcription was activated by Listeria monocytogenes independent of p38 MAPK and MEK1, and histone phosphorylation/acetylation remained unchanged in infected cells at the IFN-gamma promoter. Specific inhibition of histone deacetylases by trichostatin A increased Listeria-induced expression of IL-8, but not of IFN-gamma, underlining the specific physiological impact of histone acetylation. In conclusion, MAPK-dependent epigenetic modifications differentially contributed to L. monocytogenes-induced cytokine expression by human endothelial cells.
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PMID:Intracellular bacteria differentially regulated endothelial cytokine release by MAPK-dependent histone modification. 1611 70

Sulforaphane (SFN) is a major isothiocyanate compound in cruciferous vegetables such as broccoli, cauliflower, and Brussels sprouts. Preclinical animal models have recently shown that SFN and other isothiocyanates may be useful for prostrate cancer (PCa) chemoprevention. In this study we used a DU145 human PCa cell culture model to investigate the role of protein kinase signaling pathway(s) in SFN-induced cell cycle arrest and apoptosis and whether another chemopreventive agent selenium enhances the apoptosis potency of SFN. The results showed that SFN exposure for 24 h or longer significantly decreased the number of viable DU145 cells in a dose-dependent manner with an IC50 of asymptotically equal to 10 microM. The decreased cell number was associated with G2/M phase arrest and apoptotic cell death, with the latter being evidenced by caspase-mediated cleavage of poly(ADP-ribose) polymerase and increased release of histone-associated DNA fragments. A peptide inhibitor of caspase-8 completely blocked SFN-induced apoptosis and that for caspase-9 exerted a major protection; however, neither inhibitor attenuated SFN-induced G2/M arrest. Regarding potential mediators, SFN treatment induced a transient rise of reactive oxygen species (ROS) peaking within (1/2) h and the activation of JNK within 1 h but did not have any detectable effect on the phosphorylation of p38MAPK or ERK1/2 from 6 h to 24 h. Pretreatment of cells with N-acetylcysteine to enrich intracellular glutathione blocked SFN-induced ROS and apoptotic cell death. Inhibiting the JNK activity with a pharmacologic inhibitor SP600125 abolished the induction of G2/M arrest and apoptosis by SFN, whereas chemical inhibitors for p38MAPK and MEK1/2 did not have any modulating effect on SFN-induced apoptosis. Taken together, the data indicate that SFN decreased viable DU145 cell number in large part through the generation of ROS and JNK-mediated signaling to G2/M arrest and caspase-dependent apoptosis. Selenium in the form of inorganic sodium selenite salt or methylseleninic acid did not enhance SFN-induced apoptosis in this cell culture model.
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PMID:Involvement of c-Jun N-terminal kinase in G2/M arrest and caspase-mediated apoptosis induced by sulforaphane in DU145 prostate cancer cells. 1620 52

Isothiocyanates (ITCs) are potentially important cancer chemopreventive compounds found in cruciferous vegetables. In this study, three ITCs: allyl ITC, benzyl ITC and phenylethyl ITC, induced DNA cell-cycle changes and cell death in undifferentiated Caco-2 cells and their roles in PI3K/Akt and MEK/ERK signaling pathways have been investigated. Flow cytometric analysis was used to measure cell-cycle distribution, expression of mitotic marker (phosphorylated H3 histone), mitochondrial transmembrane potential for the determination of ITC-induced apoptosis measured by Annexin V-FITC staining and metabolic conversion of fluorescein diacetate, and quantification of sub-G1 population. Cellular MAPK and phosphorylated MAPK were measured using western blot analysis. All ITCs tested induced G2/M cell-cycle arrest after 24-h treatment, a time- and concentration-dependent activation of ERK1/2, dissipation of mitochondrial transmembrane potential and apoptosis. Both PI3K/Akt and MEK/ERK inhibitors, LY294002 and PD98059, attenuated the extent of BITC-induced cell death. Pretreatment of cells with either the PD98059 or LY294002 inhibitor, caused a dose-dependent inhibition of histone H3 (p-H3) phosphorylation. Despite the LY294002 inhibitor having no effect on the proportion of ITC-induced G2/M arrested cells, a significant decrease of p-H3/(G2/M) ratio in both PD98059- and LY294002-treated cells was observed. We suggest that the decrease of mitotic cells was compensated for by an increase of cells in G2 phase. LY294002 and PD98059 affect cell transition from G2 to M phase and from S to G2 phase respectively. These results indicate that isothiocyanates can induce cell cycle-change through multiple signaling pathways and more detailed study is merited to further unravel the chemopreventive and chemotherapeutic mechanisms of ITCs.
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PMID:Effects of MEK1 and PI3K inhibitors on allyl-, benzyl- and phenylethyl-isothiocyanate-induced G2/M arrest and cell death in Caco-2 cells. 1621 Dec 42


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