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:2.7.10.1 (ERK)
95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cellular stress activates multiple mitogen-activated protein kinase (MAPK) cascades and immediate-early gene (IEG) transcription. To address how these events are linked, we investigated the endogenous signaling/transcription factor network driving IEG activation by arsenite and anisomycin in the human osteosarcoma cell line HOS/TE-85. Induction of IEG transcription by both stresses corresponded temporally with the phosphorylation of the regulatory factors Elk-1 and cAMP response element-binding protein (CREB), along with activation of the extracellular signal-regulated kinase (ERK), stress-activated protein kinase (SAPK) and p38 MAPK cascades. To assess the role of the different cascades, they were selectively inhibited with PD98059, SP600125 and SB203580, respectively. This implicated all three cascades in Elk-1 phosphorylation after arsenite treatment, whereas ERK and SAPK inhibition diminished this, and IEG mRNA levels, downstream of anisomycin. SB blocked phosphorylation of both serum response factor (SRF) and CREB, and strongly reduced IEG activation by both stresses. Combining PD with SB further reduced arsenite induction of IEG transcription. Thus, all three MAPK cascades mediate anisomycin- and arsenite-induced signaling to IEG promoters in HOS cells through the differential targeting of Elk-1, SRF and CREB.
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PMID:Immediate-early gene induction by the stresses anisomycin and arsenite in human osteosarcoma cells involves MAPK cascade signaling to Elk-1, CREB and SRF. 1266 Aug 19

The MADS-box transcription factor Mcm1p and forkhead (FKH) transcription factor Fkh2p act in a DNA-bound complex to regulate cell-cycle dependent expression of the CLB2 cluster in Saccharomyces cerevisiae. Binding of Fkh2p requires prior binding by Mcm1p. Here we have investigated the molecular determinants governing the formation of the Mcm1p- Fkh2p complex. Fkh2p exhibits cooperativity in complex formation with Mcm1p and we have mapped a small region of Fkh2p located immediately upstream of the FKH DNA binding domain that is required for this cooperativity. This region is lacking in the related protein Fkh1p that cannot form ternary complexes with Mcm1p. A second region is identified that inhibits Mcm1p-independent DNA binding by Fkh2p. The spacing between the Mcm1p and Fkh2p binding sites is also a critical determinant for complex formation. We also show that Fkh2p can form ternary complexes with the human counterpart of Mcm1p, serum response factor (SRF). Mutations at analogous positions in Mcm1p, which are known to affect SRF interaction with its partner protein Elk-1, abrogate complex formation with Fkh2p, demonstrating evolutionary conservation of coregulatory protein binding surfaces. Our data therefore provide molecular insights into the mechanisms of Mcm1p- Fkh2p complex formation and more generally aid our understanding of MADS-box protein function.
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PMID:Molecular determinants of the cell-cycle regulated Mcm1p-Fkh2p transcription factor complex. 1271 72

The chaperonin-containing t-complex polypeptide 1 (CCT) is a molecular chaperone that facilitates protein folding in eukaryotic cytosol, and the expression of CCT is highly dependent on cell growth. We show here that transcription of the gene encoding the theta subunit of mouse CCT, Cctq, is regulated by the ternary complex factors (TCFs), Elk-1, Sap-1a, and Net (Sap-2). Reporter gene assay using HeLa cells indicated that the Cctq gene promoter contains a cis-acting element of the CCGGAAGT sequence (CQE1) at -36 bp. The major CQE1-binding proteins in HeLa cell nuclear extract was recognized by anti-Elk-1 or anti-Sap-1a antibodies in electrophoretic mobility shift assay, and recombinant Elk-1, Sap-1a, or Net specifically recognized CQE1. The CQE1-dependent transcriptional activity in HeLa cells was virtually abolished by overexpression of the DNA binding domains of TCFs. Overexpression of full-length TCFs with Ras indicated that exogenous TCFs can regulate the CQE1-dependent transcription in a Ras-dependent manner. PD98059, an inhibitor of MAPK, significantly repressed the CQE1-dependent transcription. However, no serum response factor was detected by electrophoretic mobility shift assay using the CQE1 element. These results indicate that transcription of the Cctq gene is regulated by TCFs under the control of the Ras/MAPK pathway, probably independently of serum response factor.
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PMID:Transcriptional regulation of the cytosolic chaperonin theta subunit gene, Cctq, by Ets domain transcription factors Elk-1, Sap-1a, and Net in the absence of serum response factor. 1278 37

NFAT and SRF are important in the regulation of proliferation and cytokine production in lymphocytes. NFAT activation by the B cell receptor (BCR) occurs via the PLCgamma-Ca(2+)-calcineurin pathway, however how the BCR activates SRF is unclear. We show here that like NFAT, BCR regulation of SRF occurs via an Src-Syk-Tec-PLCgamma-Ca(2+) (Lyn-Syk-Btk-PLCgamma-Ca(2+)) pathway. However, SRF responds to lower Ca(2+) and is less dependent on IP(3)R expression than NFAT. Ca(2+)-regulated calcineurin plays a partial role in SRF activation, in combination with diacylglycerol (DAG), while is fully required for NFAT activation. Signals from the DAG effectors protein kinase C, Ras and Rap1, and the downstream MEK-ERK pathway are required for both SRF and NFAT; however, NFAT but not SRF is dependent on JNK signals. Both SRF and NFAT were also dependent on Rac, Rho, CDC42 and actin. Finally, we show that Ca(2+) is not required for ERK activation, but instead for its association with nuclear areas of the cell. These data suggest that combinatorial assembly of signaling pathways emanating from the BCR differentially regulate NFAT and SRF, to activate gene expression.
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PMID:Differential regulation of NFAT and SRF by the B cell receptor via a PLCgamma-Ca(2+)-dependent pathway. 1291 15

The transcription factor Elk-1 belongs to the ternary complex factor (TCF) subfamily of Ets proteins. TCFs interact with serum response factor to bind jointly to serum response elements in the promoters of immediate-early genes (IEGs). TCFs mediate the rapid transcriptional response of IEGs to various extracellular stimuli which activate mitogen-activated protein kinase signaling. To investigate physiological functions of Elk-1 in vivo, we generated Elk-1-deficient mice by homologous recombination in embryonic stem cells. These animals were found to be phenotypically indistinguishable from their wild-type littermates. Histological analysis of various tissues failed to reveal any differences between Elk-1 mutant and wild-type mice. Elk-1 deficiency caused no changes in the proteomic displays of brain or spleen extracts. Also, no immunological defects could be detected in mice lacking Elk-1, even upon infection with coxsackievirus B3. In mouse embryonic fibroblasts, Elk-1 was dispensable for c-fos and Egr-1 transcriptional activation upon stimulation with serum, lysophosphatidic acid, or tetradecanoyl phorbol acetate. However, in brains of Elk-1-deficient mice, cortical and hippocampal CA1 expression of c-fos, but not Egr-1 or c-Jun, was markedly reduced 4 h following kainate-induced seizures. This was not accompanied by altered patterns of neuronal apoptosis. Collectively, our data indicate that Elk-1 is essential neither for mouse development nor for adult life, suggesting compensatory activities by other TCFs.
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PMID:Mice deficient for the ets transcription factor elk-1 show normal immune responses and mildly impaired neuronal gene activation. 1467 63

The three ternary complex factors (TCFs) Elk-1, Net and Sap-1 form a subfamily of the E twenty-six (Ets) domain transcription factors. Their characteristic property is the ability to form a ternary nucleoprotein complex with the serum response factor (SRF) over the serum response element (SRE) of the c-fos promoter. The molecular mechanisms that underlie the function and regulation of these factors have been extensively studied and the TCFs are a paradigm for the study of transcriptional regulation in response to extracellular signalling through the mitogen-activated protein (MAP) kinase pathway. As final effectors of multiple signalling pathways and components of protein complexes on immediate early promoters, they represent key elements in the complex and dynamic regulation of gene expression. This review summarises the molecular, structural and biochemical studies that have led to the understanding of the functional domains of the TCFs, ternary complex formation, transcriptional regulation, protein partners and target genes in cell lines. Finally, the emerging studies of the biological roles of the TCFs in vivo will be discussed.
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PMID:Ets ternary complex transcription factors. 1469 67

We have taken an integrated approach in which expression profiling has been combined with the use of small molecule inhibitors and computational analysis of transcription factor binding sites to characterize regulatory sequences of genes that are targets of specific signaling pathways in growth factor-stimulated human cells. T98G cells were stimulated with platelet-derived growth factor (PDGF) and analyzed by DNA microarrays, which identified 74 immediate-early gene transcripts. Cells were then treated with inhibitors to identify subsets of genes that are targets of the phosphatidylinositol 3-kinase (PI3K) and MEK/ERK signaling pathways. Four groups of PDGF-induced genes were defined: independent of PI3K and MEK/ERK signaling, dependent on PI3K signaling, dependent on MEK/ERK signaling, and dependent on both pathways. The upstream regions of all genes in the four groups were scanned using TRANSFAC for putative cis-elements as compared with a background set of non-induced genes. Binding sites for 18 computationally predicted transcription factors were over-represented in the four groups of co-expressed genes compared with the background sequences (p < 0.01). Many of the cis-elements identified were conserved in orthologous mouse genes, and many of the predicted elements and their cognate transcription factors were consistent with previous experimental data. In addition, chromatin immunoprecipitation assays experimentally verified nine predicted SRF binding sites in T98G cells, including a previously unknown SRF site upstream of DUSP5. These results indicate that groups of human genes regulated by discrete intracellular signaling pathways share common cis-regulatory elements.
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PMID:Identification of transcription factor binding sites upstream of human genes regulated by the phosphatidylinositol 3-kinase and MEK/ERK signaling pathways. 1476 1

Elk-1 is a member of the TCF subfamily of Ets proteins. TCFs interact with SRF at serum response elements (SREs) of immediate early genes (IEGs), such as c-fos and Egr-1, thereby mediating IEG induction upon extracellular stimulation. We previously generated an Elk-1 null allele (Elk1-137) in murine embryonic stem (ES) cells by homologous recombination. In Elk1-137, the Elk-1 gene was replaced by a Hygromycin B phosphotransferase - Thymidine Kinase (HygTk) fusion gene, flanked by two nonidentical Flp recombinase recognition (FRT) sites (Cesari et al., [2004] Mol Cell Biol, in press) to allow for the subsequent generation of alternative alleles of interest by recombinase-mediated cassette exchange (RMCE). Elk1-deficient mice derived from Elk-1((137/0)) ES cells are viable and do not reveal strong phenotypical abnormalities, apart from male sterility. However, the Elk-1 locus contains the Tk cassette, which has previously been related to this defect. Therefore, in our first experiment involving the technique of Flp RMCE we chose to remove the HygTk cassette in Elk-1((137/0)) ES cells and to generate Elk-1((RMCE16/0)) and Elk-1((RMCE16/RMCE16)) mice. In so doing, we provide evidence that the sterility of Elk1((137/0)) mice was not due to the absence of Elk-1 but rather the presence of HygTk. This is the first report of mice derived from ES cells which were subjected to Flp RMCE and thus proves that RMCE is a powerful tool for the genetic engineering of previously tagged loci in the mouse genome.
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PMID:Elk-1 knock-out mice engineered by Flp recombinase-mediated cassette exchange. 1499 71

Gab2 (Grb2-associated binder-2), a member of the IRS (insulin receptor substrate)/Gab family of adapter proteins, undergoes tyrosine phosphorylation in response to cytokine or growth factor stimulation and serves as a docking platform for many signal transduction effectors, including the tyrosine phosphatase SHP-2 [SH2 (Src homology 2)-domain-containing tyrosine phosphatase]. Here, we report that, following IL-2 (interleukin-2) stimulation of human T lymphocytes, SHP-2 binds tyrosine residues 614 and 643 of human Gab2 through its N- and C-terminal SH2 domains respectively. However, the sole mutation of Tyr-614 into phenylalanine is sufficient to prevent Gab2 from recruiting SHP-2. Expression of the Gab2 Tyr-614-->Phe (Y614F) mutant, defective in SHP-2 association, prevents ERK (extracellular-signal-regulated kinase) activation and expression of a luciferase reporter plasmid driven by the c-fos SRE (serum response element), indicating that interaction of SHP-2 with Gab2 is required for ERK activation in response to IL-2. Further investigation of IL-2-dependent induction of SRE showed that expression of a constitutively active mutant of the RhoA GTPase synergizes with IL-2 for SRE-driven transcription, whereas a dominant-negative mutant reduces the IL-2 response. Thus, in response to IL-2, full induction of the SRE requires ERK-dependent as well as Rho-dependent signals that target the Ets-box and the CArG-box respectively. We also report that the synergy between Gab2/SHP-2 and RhoA for IL-2-dependent CArG-box-driven transcription depends upon MEK (mitogen-activated protein kinase/ERK kinase) activation, and is likely to involve regulation of the serum response factor co-activator MAL. Our studies thus provide new insights into the role of Gab2 and SHP-2 in IL-2 signal transduction.
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PMID:Interaction of the tyrosine phosphatase SHP-2 with Gab2 regulates Rho-dependent activation of the c-fos serum response element by interleukin-2. 1517 Mar 89

Sphingosine 1-phosphate (S1P) is a lipid agonist that regulates smooth muscle cell (SMC) and endothelial cell functions by activating several members of the S1P subfamily of G-protein-coupled Edg receptors. We have shown previously that SMC differentiation is regulated by RhoA-dependent activation of serum response factor (SRF). Because S1P is a strong activator of RhoA, we hypothesized that S1P would stimulate SMC differentiation. Treatment of primary rat aortic SMC cells with S1P activated RhoA as measured by precipitation with a glutathione S-transferase-rhotekin fusion protein. In SMC and 10T1/2 cells, S1P treatment up-regulated the activities of several transiently transfected SMC-specific promoters, and these effects were inhibited by the Rho-kinase inhibitor, Y-27632. S1P also increased smooth muscle alpha-actin protein levels in SMC but had no effect on SRF binding to the smooth muscle alpha-actin CArG B element. Quantitative reverse transcriptase-PCR showed that S1P treatment of SMC or 10T1/2 cells did not increase the mRNA level of either of the recently identified SRF co-factors, myocardin or myocardin-related transcription factor-A (MRTF-A). MRTF-A protein was expressed highly in SMC and 10T1/2 cultures, and importantly the effects of S1P were inhibited by a dominant negative form of MRTF-A indicating that S1P may regulate the transcriptional activity of MRTF-A. Indeed, S1P treatment increased the nuclear localization of FLAG-MRTF-A, and the effect of MRTF-A overexpression on smooth muscle alpha-actin promoter activity was inhibited by dominant negative RhoA. S1P also stimulated SMC growth by activating the early growth response gene, c-fos. This effect was not attenuated by Y-27632 but could be inhibited by the MEK inhibitor, UO126. S1P enhanced SMC growth through ERK-mediated phosphorylation of the SRF co-factor, Elk-1, as measured by gel shift and Elk-1 activation assays. Taken together these results demonstrate that S1P activates multiple signaling pathways in SMC and regulates proliferation by ERK-dependent activation of Elk-1 and differentiation by RhoA-dependent activation of MRTF-A.
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PMID:Sphingosine 1-phosphate stimulates smooth muscle cell differentiation and proliferation by activating separate serum response factor co-factors. 1529 66


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