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)

Orf virus, a member of the poxvirus family, produces a pustular dermatitis in sheep, goats, and humans. The lesions induced after infection with orf virus show extensive proliferation of vascular endothelial cells, dilation of blood vessels and dermal swelling. An explanation for the nature of these lesions may lie in the discovery that orf virus encodes an apparent homolog of the mammalian vascular endothelial growth factor (VEGF) family of molecules. These molecules mediate endothelial cell proliferation, vascular permeability, angiogenesis, and lymphangiogenesis via the endothelial cell receptors VEGFR-1 (Flt1), VEGFR-2 (KDR/Flk1), and VEGFR-3 (Flt4). The VEGF-like protein of orf virus strain NZ2 (ORFV2-VEGF) is most closely related in primary structure to VEGF. In this study we examined the biological activities and receptor specificity of the ORFV2-VEGF protein. ORFV2-VEGF was found to be a disulfide-linked homodimer with a subunit of approximately 25 kDa. ORFV2-VEGF showed mitogenic activity on bovine aortic and human microvascular endothelial cells and induced vascular permeability. ORFV2-VEGF was found to bind and induce autophosphorylation of VEGFR-2 and was unable to bind or activate VEGFR-1 and VEGFR-3, but bound the newly identified VEGF165 receptor neuropilin-1. These results indicate that, from a functional viewpoint, ORFV2-VEGF is indeed a member of the VEGF family of molecules, but is unique, however, in that it utilizes only VEGFR-2 and neuropilin-1.
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PMID:Vascular endothelial growth factor (VEGF)-like protein from orf virus NZ2 binds to VEGFR2 and neuropilin-1. 1007 38

Exposure to silica has been associated with progressive pulmonary inflammation and fibrosis. While the fibroblasts play an important role in the pathogenesis of silicosis, the direct interaction between silica and fibroblasts is poorly understood. We observed that silica particles stimulated intracellular ROS generation in Rat2 fibroblast, evidenced by DCFH oxidation. Silica-induced DCFH oxidation was inhibited by catalase and DPI, a flavoenzyme inhibitor. Additionally, the time course of elevation of the intracellular ROS was paralleled by the increases of MEK and ERK phosphorylation. Silica-induced ERK phosphorylation was also effectively attenuated by catalase and DPI. However, SOD enhanced the silica-induced ERK phosphorylation, indicating a role for H(2)O(2) in ERK activation. Furthermore, ERK and MEK phosphorylation are reproduced by H(2)O(2) treatment. Taken together, these results demonstrate that silica stimulates ROS production via flavoenzyme-dependent mechanism in Rat2 fibroblasts and the H(2)O(2), in turn, serves as a signal transduction element in activating MEK-ERK pathway.
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PMID:Silica-induced generation of reactive oxygen species in Rat2 fibroblast: role in activation of mitogen-activated protein kinase. 1047 90

The human epidermal growth factor receptors 2, 3, and 4 (HER2, HER3, and HER4, respectively) are frequently overexpressed in many human cancers, and therefore may be potential targets for receptor-mediated gene transfer. To evaluate this possibility, we constructed a series of HER-targeted gene transfer vehicles by covalently linking poly-L-lysine polymers (pLYS) to the epidermal growth factor-like domain of the HER ligand neuregulin-1 (NRG1(177-244)), a HER2 antibody (Ab), and the Fab fragment of the HER2 Ab. In vitro, pLYS modification of NRG1(177-244) decreased the affinity of the ligand for HER3 or HER4 homodimer receptors by 6- to 7-fold. DNA loading of the pLYS-modified NRG1(177-244) had a minimal additional affect on the affinity of the complex for its receptor. In cell lines engineered to solely express HER2, HER3, or HER4, each vehicle correctly targeted the receptors; the NRG1(177-244) construct transferred a luciferase gene only into cells expressing HER4, whereas the HER2 Ab and Fab constructs transferred the reporter gene only into cells expressing HER2. The most efficient gene transfer occurred using the intact HER2 Ab as a gene transfer vehicle, whereas the Fab fragment of the HER2 Ab was the least efficient, and NRG1(177-244) was intermediate. These studies suggest that the NRG receptor or HER2, a component of the receptor, can be pursued as targets for gene transfer.
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PMID:Neuregulin receptor-mediated gene transfer by human epidermal growth factor receptor 2-targeted antibodies and neuregulin-1. 1060 50

Platelet-derived growth factor (PDGF) B-chain and PDGF receptor beta (PDGFR beta) are essential for glomerulogenesis. Mice deficient in PDGF B-chain or PDGFR beta exhibit an abnormal glomerular phenotype characterized by total lack of mesangial cells. In this study, we localized PDGFR beta in the developing rat kidney and explored the biological effects of PDGF in metanephric mesenchymal cells in an attempt to determine the mechanism by which PDGF regulates mesangial cell development. Immunohistochemical and in situ hybridization studies of rat embryonic kidneys reveal that PDGFR beta localizes to undifferentiated metanephric mesenchyme and is later expressed in the cleft of the comma-shaped and S-shaped bodies and in more mature glomeruli in a mesangial distribution. We also isolated and characterized cells from rat metanephric mesenchyme. Metanephric mesenchymal cells express vimentin and alpha-smooth muscle actin but not cytokeratin. These cells also express functional PDGFR beta, as demonstrated by autophosphorylation of the receptor as well as activation of phosphatidylinositol 3 kinase in response to PDGF B-chain homodimer. PDGF B-chain also induces migration and proliferation of metanephric mesenchymal cells. Taken together with the fact that PDGF B-chain is expressed in the glomerular epithelium and mesangial area, as demonstrated in the human embryonic kidney, we suggest that PDGF B-chain acts in a paracrine fashion to stimulate the migration and proliferation of mesangial cell precursors from undifferentiated metanephric mesenchyme to the mesangial area. PDGF B-chain also likely stimulates proliferation of mesangial cell precursors in an autocrine fashion once these cells migrate to the glomerular tuft.
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PMID:Platelet-derived growth factor receptor beta regulates migration and DNA synthesis in metanephric mesenchymal cells. 1073 1

Platelet-derived growth factor (PDGF) is a potent mitogen for mesenchymal cells. The PDGF B-chain (c-sis proto-oncogene) homodimer (PDGF BB) and v-sis, its viral counterpart, activate both alpha- and beta-receptor subunits (alpha-PDGFR and beta-PDGFR) and mediate anchorage-independent growth in NIH3T3 cells. In contrast, the PDGF A chain homodimer (PDGF AA) activates alpha-PDGFR only and fails to induce phenotypic transformation. In the present study, we investigated alpha- and beta-PDGFR specific signaling pathways that are responsible for the differences between the transforming ability of PDGF AA and BB. To study PDGF BB activation of beta-PDGFR, we established NIH3T3 clones in which alpha-PDGFR signaling is inhibited by a dominant-negative alpha-PDGFR, or an antisense construct of alpha-PDGFR. Here, we demonstrate that beta-PDGFR activation alone is sufficient for PDGF BB-mediated anchorage-independent cell growth. More importantly, inhibition of alpha-PDGFR signaling enhanced PDGF BB-mediated phenotypic transformation, suggesting that alpha-PDGFR antagonizes beta-PDGFR-induced transformation. While both alpha- and beta-receptors effectively activate ERKs, alpha-PDGFR, but not beta-PDGFR, activates stress-activated protein kinase-1/c-Jun NH(2)-terminal kinase-1 (JNK-1). Inhibition of JNK-1 activity using a dominant-negative JNK-1 mutant markedly enhanced PDGF BB-mediated anchorage-independent cell growth, demonstrating an antagonistic role for JNK-1 in PDGF-induced transformation. Consistently, overexpression of wild-type JNK-1 reduced PDGF BB-mediated transformation. Taken together, the present study showed that alpha- and beta-PDGFRs differentially regulate Ras-mitogen-activated protein kinase pathways critical for regulation of cell transformation, and transformation suppressing activity of alpha-PDGFR involves JNK-1 activation.
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PMID:Platelet-derived growth factor (PDGF) receptor-alpha activates c-Jun NH2-terminal kinase-1 and antagonizes PDGF receptor-beta -induced phenotypic transformation. 1077 15

Certain point mutations within the hydrophobic transmembrane domains of class I receptor tyrosine kinases have been associated with oncogenic transformation in vitro and in vivo [Gullick, J., and Srinivasan, R. (1998) Breast Cancer Res. Treat. 52, 43-53]. An important example is the replacement of a single (hydrophobic) valine by (charged) glutamate in the rat protein, Neu, and in the homologous human protein, ErbB-2. It has been suggested that the oncogenic nature of this Val-->Glu substitution may derive from alteration of the transmembrane domain's ability to take part in direct side-to-side associations. In the present work, we examined the basis of this phenomenon by studying transmembrane portions of ErbB-2 in fluid bilayer membranes. An expression system was designed to produce such peptides from the wild-type ErbB-2, and from an identical region of the transforming mutant in which Val(659) is replaced by Glu. All peptides were 50-mers, containing the appropriate transmembrane domain plus contiguous stretches of amino acids from the cytoplasmic and extracellular domains. Deuterium heteronuclear probes were incorporated into alanine side chains (thus, each alanine -CH(3) side chain became -CD(3)). Given the presence of natural alanine residues at positions 648 and 657 within ErbB-2, this approach afforded heteronuclear probes within the motif Ser(656)AlaValValGlu(660), thought to be important for homodimer formation, and nine residues upstream of this site. Further peptides were produced, by site-directed mutagenesis, to confirm spectral assignments and to provide an additional probe location at position 670 (11 residues downstream of the motif region). On SDS-polyacrylamide gels, the transmembrane peptides migrated as predominant monomers in equilibrium with smaller populations of homodimers/-oligomers. CD spectra of both wild-type and transforming mutant peptides were consistent with the transmembrane portions being basically alpha-helical. (2)H NMR spectra of each transmembrane peptide were obtained in fluid phospholipid bilayers of 1-palmitoyl-2-oleoylphosphatidylcholine (POPC) from 35 to 65 degrees C. Results were consistent with the concept that the glutamic acid residue characterizing the mutant is uncharged at neutral pH. Narrowed spectral components from species rotating rapidly and symmetrically within the membrane appeared to represent monomeric peptide. Mutation of Val(659) to Glu within the hydrophobic domain induced changes in side chain angulation of at least 6-8 degrees at Ala(657) (i.e., within the five amino acid motif thought to be involved in homodimer formation), and downstream of this site to residue 670. There was little evidence of effect at the upstream site (Ala(648)) at the membrane surface. This result argues that the transforming mutation is associated with significant intramolecular rearrangement of the monomeric transmembrane helix-extending over some four helix turns-which could influence its lateral associations. In addition, temperature effects on spectral quadrupole splittings suggested that there is greater peptide backbone flexibility for the wild-type transmembrane region.
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PMID:Val(659)-->Glu mutation within the transmembrane domain of ErbB-2: effects measured by (2)H NMR in fluid phospholipid bilayers. 1082 74

Neuropilin-1 (NP-1) was first identified as a semaphorin receptor involved in neuron guidance. Subsequent studies demonstrated that NP-1 also binds an isoform of vascular endothelial growth factor (VEGF) as well as several VEGF homologs, suggesting that NP-1 may also function in angiogenesis. Here we report in vitro binding experiments that shed light on the interaction between VEGF165 and NP-1, as well as a previously unknown interaction between NP-1 and one of the VEGF receptor tyrosine kinases, VEGFR1 or Flt-1. BIAcore analysis demonstrated that, with the extracellular domain (ECD) of NP-1 immobilized at low density, VEGF165 bound with low affinity (K(d) = 2 microm) and fast kinetics. The interaction was dependent on the heparin-binding domain of VEGF165 and increased the affinity of VEGF165 for its signaling receptor VEGFR2 or kinase insert domain-containing receptor. The affinity of VEGF165 for the NP-1 ECD was greatly enhanced either by increasing the density of immobilized NP-1 (K(d) = 113 nm) or by the addition of heparin (K(d) = 25 nm). We attribute these affinity enhancements to avidity effects mediated by the bivalent VEGF165 homodimer or multivalent heparin. We also show that the NP-1 ECD binds with high affinity (K(d) = 1.8 nm) to domains 3 and 4 of Flt-1 and that this interaction inhibits the binding of NP-1 to VEGF165. Based on these results, we propose that NP-1 acts as a coreceptor for various ligands and that these functions are dependent on the density of NP-1 on the cell membrane. Furthermore, Flt-1 may function as a negative regulator of angiogenesis by competing for NP-1.
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PMID:The interaction of neuropilin-1 with vascular endothelial growth factor and its receptor flt-1. 1084 81

The properties of two VEGF receptors, Flt-1 and KDR, in the signal transduction of VEGF in human umbilical vein endothelial cells (HUVECs) were investigated by using two newly developed blocking monoclonal antibodies (mAbs) against Flt-1 and KDR. VEGF stimulated the expression of transcription factor Ets-1 as well as matrix metalloproteinase-1 (MMP-1) and Flt-1 in HUVECs. The KDR/Flt-1 heterodimer and the KDR homodimer mediate the expression of Ets-1, MMP-1, and Flt-1. VEGF also stimulated DNA synthesis and migration of HUVECs. DNA synthesis is mediated by the same signaling system as the expression of Ets-1. In contrast, cell migration is regulated by two distinct signaling systems. The Flt-1 homodimer is required for actin reorganization. The KDR/Flt-1 heterodimer and the KDR homodimer are required for the assembly of vinculin in focal adhesion plaque by regulating the phosphorylation of focal adhesion kinase (FAK) and paxillin.
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PMID:Properties of two VEGF receptors, Flt-1 and KDR, in signal transduction. 1086 39

The epidermal growth factor (EGF)-like family of growth factors elicits cellular responses by stimulating the dimerization, autophosphorylation, and tyrosine kinase activities of the ErbB family of receptor tyrosine kinases. Although several different EGF-like ligands are capable of binding to a single ErbB family member, it is generally thought that the biological and biochemical responses of a single receptor dimer to different ligands are indistinguishable. To test whether an ErbB receptor dimer is capable of discriminating among ligands we have examined the effect of four EGF-like growth factors on signaling through the ErbB4 receptor homodimer in CEM/HER4 cells, a transfected human T cell line ectopically expressing ErbB4 in an ErbB-null background. Despite stimulating similar levels of gross receptor tyrosine phosphorylation, the EGF-like growth factors betacellulin, neuregulin-1beta, neuregulin-2beta, and neuregulin-3 exhibited different biological potencies in a cellular growth assay. Moreover, the different ligands induced different patterns of recruitment of intracellular signaling proteins to the activated receptor and induced differential usage of intracellular kinase signaling cascades. Finally, two-dimensional phosphopeptide mapping of ligand-stimulated ErbB4 revealed that the different growth factors induce different patterns of receptor tyrosine phosphorylation. These results indicate that ErbB4 activation by growth factors is not generic and suggest that individual ErbB receptors can discriminate between different EGF-like ligands within the context of a single receptor dimer. More generally, our observations significantly modify our understanding of signaling through receptor tyrosine kinases and point to a number of possible models for ligand-mediated signal diversification.
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PMID:Ligand discrimination in signaling through an ErbB4 receptor homodimer. 1086 24

When quiescent endothelial cells (ECs) are exposed to angiogenic factor such as VEGF; ECs express proteases to degrade extracellular matrices, migrate, proliferate and form new vessels. However, the molecular mechanism of these events is not fully characterized yet. We are studying the signal transduction and transcriptional regulation of angiogenesis. We investigated the properties of two VEGF receptors, Flt-1 and KDR, by using two newly developed blocking monoclonal antibodies (mAbs), i.e., anti-human Flt-1 mAb and anti-human KDR mAb. VEGF elicited induction of transcription factor Ets-1 in human umbilical vein endothelial cells (HUVECs). This induction was mediated by the KDR/Flt-1 heterodimer and the KDR homodimer. The role of transcription factor Ets-1 in angiogenesis was further clarified. We established both high and low Ets-1 expressing EC lines, and compared angiogenic properties of these cell lines with a parental murine EC line, MSS31. The growth rate was almost identical among three cell lines. It appeared that gene expressions of matrix metalloproteinases (MMP-1, MMP-3, and MMP-9) as well as integrin beta 3 were correlated with the level of Ets-1 expression. As a result, the invasiveness was enhanced in high Ets-1 expressing cells and reduced in low Ets-1 expressing cells compared with parental cells, and high Ets-1 expressing cells made more tube-like structures in type 1 collagen gel. These results indicate that Ets-1 is a principle transcription factor converting ECs to the angiogeneic phenotype.
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PMID:Signal transduction and transcriptional regulation of angiogenesis. 1094 59


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