Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.24 (mitogen-activated protein kinase)
 95,810 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In PC12 cells, cAMP stimulates the MAP kinase pathway by an unknown mechanism. Firstly, we examined the role of calcium ion mobilization and of protein kinase C in cAMP-stimulated MAP kinase activation. We show that cAMP stimulates p44mapk independently of these events. Secondly, we studied the role of B-Raf in this process. We observed that NGF, PMA and cAMP induce the phosphorylation of B-Raf as well as an upward shift in its electrophoretic mobility. We show that B-Raf is activated following NGF and PMA treatment of PC12 cells, and that it can phosphorylate and activate MEK-1. However, cAMP inhibits B-Raf autokinase activity as well as its ability to phosphorylate and activate MEK-1. This inhibition is likely to be due to a direct effect since we found that PKA phosphorylates B-Raf in vitro. Further, we show that B-Raf binds to p21ras, but more important, this binding to p21ras is virtually abolished with B-Raf from PC12 cells treated with CPT-cAMP. Hence, these data indicate that the PKA-mediated phosphorylation of B-Raf hampers its interaction with p21ras, which is responsible for the PKA-mediated decrease in B-Raf activity. Finally, our work suggests that in PC12 cells, cAMP stimulates MAP kinase through the activation of an unidentified MEK kinase and/or the inhibition of a MEK phosphatase.
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PMID:Regulation of the MAP kinase cascade in PC12 cells: B-Raf activates MEK-1 (MAP kinase or ERK kinase) and is inhibited by cAMP. 783 30

Up-regulation of ERK (extracellular signal-regulated kinase or MAP kinase) and MEK (ERK kinase or MAPK kinase) expression after rat facial nerve injury was demonstrated by in situ hybridization histochemistry and immunohistochemistry. These two enzymes play roles in one of the major intracellular signal cascade pathways involving receptor tyrosine kinase common to growth factor receptors, and transcription factors. Significant increases in ERK1 mRNA levels were observed from day 3 after facial nerve transection, with the highest level of expression from 1 to 2 weeks after the operation. This high level of mRNA expression then decreased gradually to the normal level. ERK1-like immunoreactivity showed a similar time course to that of its mRNA expression; however, the decay profile was more prolonged. The up-regulation of MEK, the ERK kinase/MAPK kinase, was also detected by immunohistochemistry. The protein expression profiles were almost equivalent, but the MEK expression was slightly advanced, suggesting that the observed up-regulation of MEK was not due to that of ERK. The receptor tyrosine kinase signal transduction pathway via MEK-ERK located downstream of growth factor receptors seems vital as a regulator of the synthesis of molecules that play important roles in the recovery process following injury or/and regeneration.
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PMID:Up-regulation of ERK (MAP kinase) and MEK (MAP kinase kinase) transcription after rat facial nerve transection. 783 28

Exposure of mesangial cells to platelet-derived growth factor (PDGF) BB caused a significant stimulation of cell proliferation and protein synthesis, as measured by [3H]thymidine incorporation and [3H]leucine incorporation respectively. In contrast, cells treated with angiotensin II had no significant increase in [3H]thymidine incorporation, but demonstrated a marked increase in [3H]leucine incorporation. Furthermore, angiotensin II significantly increased total protein content per cell. These data show that, whereas PDGF-BB is a mitogen and stimulates mesangial-cell hyperplasia, angiotensin II causes hypertrophy of the cells without hyperplasia. Treatment of mesangial cells with PDGF and angiotensin II rapidly and dose-dependently stimulated mitogen-activated protein (MAP) kinase activity, as shown by an assay for activity in vitro using myelin basic protein as a substrate, and by immunoprecipitation of 32P-labelled cells with specific antibodies against the 42 kDa and 44 kDa mitogen-activated protein kinases p42mapk and p44mapk, respectively. Whereas stimulation with PDGF-BB caused a potent and sustained (for more than 30 min) phosphorylation and activation of p42mapk and p44mapk, as well as of the upstream activators MAP kinase kinase and c-Raf, the effect of angiotensin II was less potent, reaching a peak at 5-10 min and thereafter declining rapidly. In summary, these results suggest that PDGF-BB and angiotensin II differ in their potency and duration of activation of the MAP kinase cascade, which may explain why PDGF-BB is a potent mitogen for mesangial cells, whereas angiotensin II only triggers mesangial-cell hypertrophy.
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PMID:Platelet-derived growth factor and angiotensin II stimulate the mitogen-activated protein kinase cascade in renal mesangial cells: comparison of hypertrophic and hyperplastic agonists. 784 76

Epidermal growth factor (EGF) induces a rapid increase in the phosphorylation of extracellular signal-regulated kinases (ERKs) in the human osteosarcoma osteoblastic cell line G292 and in primary cultures of rat osteoblastic cells. This phosphorylation is transient and time-dependent. Maximal stimulation is attained within 1 min in G292 and within 5 min in rat osteoblastic cells. Enzymatic activity in G292 cells is also induced rapidly after EGF stimulation. Western blot analysis revealed that enhancement of the phosphorylation of ERKs in the EGF-stimulated cells is not due to an increase in ERK protein, since EGF-treatment does not lead to an increase in the absolute amount of ERKs present even after 2 days of stimulation. The pattern of expression of the ERKs observed in the two cell types differs in the apparent molecular weights observed. The most slowly migrating immunoreactive protein (approximately 45 kDa) in normal rat osteoblastic cells is ERK1, identified by an ERK1-selective antiserum. The same antiserum reacts only weakly with one of the ERK proteins (44 kDa) blotted from the human osteosarcoma cell line G292. Phorbol 12-myristate 13-acetate (PMA) is also capable of inducing ERK phosphorylation, albeit to a lasser degree. The combination of PMA and EGF does not produce a greater response than EGF alone. The role of protein kinase C (PKC) in the EGF-stimulated ERK signaling pathway was further examined by inhibition of PKC with the staurosporine analog, CGP41251, and by down-regulation of PKC via chronic treatment with PMA. Chronic PMA treatment results in a partial inhibition of the EGF-mediated phosphorylation. CGP41251 completely abolishes the increased ERK activity produced by PMA, but the effect of EGF in this regard is potentiated. We conclude that PKC and EGF act through parallel pathways to stimulate ERK phosphorylation and activity. The inhibitor studies, in addition, indicate that activation of PKC may moderate the actions of the EGF pathway via a tonic inhibitory feedback.
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PMID:EGF-mediated phosphorylation of extracellular signal-regulated kinases in osteoblastic cells. 786 Jun 43

We have examined porcine granulosa cells (pGCs) for the presence of immunodetectable mitogen-activated protein (MAP) kinases (extracellular signal-regulated kinases, ERK) and have further studied the effects of epidermal growth factor (EGF) on the activation of these kinases. Cell lysates prepared from untreated monolayer cultures of pGCs were subjected to Western immunoblotting analysis using monoclonal antibodies to ERK1, ERK2 and pan-specific ERK. MAP kinases were detected having mol wts of 87K (ERK87), 54K (ERK54), 44K (ERK1), and 42K (ERK2). Treatment of pGCs with increasing concentrations (1-10 ng/ml) of EGF for 10 min resulted in electrophoretic mobility shifts of ERK1 and ERK2 suggesting hyperphosphorylation. Immunoprecipitation with an antiphosphotyrosine antibody (PY20), followed by Western analysis using pan-ERK, revealed a marked concentration-dependent increase in tyrosine phosphorylation of ERK2 in response to EGF treatment. The mobility shift and tyrosine phosphorylation of ERK2 was observed as early as 1 min after treatment with 10 ng/ml EGF. In-gel myelin basic protein (MBP) kinase assays revealed significant MBP kinase activity associated with ERK1 and ERK2 in total cell lysates and ERK2 in PY20 immunoprecipitates. Although ERK1 displayed a moderate mobility shift in response to EGF, tyrosine phosphorylation of this MAP kinase was not appreciably increased by EGF. Furthermore, PY20 immunoprecipitates demonstrated minimal MBP kinase associated with ERK1 in response to EGF treatment. Electrophoretic migration, tyrosine phosphorylation, and MBP kinase activity of the ERK54 and ERK87 was not effected regardless of EGF concentration or duration of treatment. These data demonstrate for the first time that pGCs contain immunodetectable MAP kinases. EGF, in a concentration- and time-dependent manner, increases tyrosine phosphorylation and MBP kinase activity (i.e. activation) of ERK2, and to a lesser degree ERK1, suggesting that the activation of MAP kinase may mediate the mitogenic action of EGF in pGCs.
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PMID:Effects of epidermal growth factor on the tyrosine phosphorylation of mitogen-activated protein kinases in monolayer cultures of porcine granulosa cells. 786 73

Induction of the human c-fos proto-oncogene by mitogens depends on the formation of a ternary complex by p62TCF with the serum response factor (SRF) and the serum response element (SRE). We demonstrate that Elk-1, a protein closely related to p62TCF in function, is a nuclear target of two members of the MAP kinase family, ERK1 and ERK2. Phosphorylation of Elk-1 increases the yield of ternary complex in vitro. At least five residues in the C-terminal domain of Elk-1 are phosphorylated upon growth factor stimulation of NIH3T3 cells. These residues are also phosphorylated by purified ERK1 in vitro, as determined by a combination of phosphopeptide sequencing and 2-D peptide mapping. Conversion of two of these phospho-acceptor sites to alanine impairs the formation of ternary complexes by the resulting Elk-1 proteins. Removal of these serine residues also drastically diminishes activation of the c-fos promoter in epidermal growth factor-treated cells. Analogous mutations at other sites impair activation to a lesser extent without affecting ternary complex formation in vitro. Our results indicate that phosphorylation regulates ternary complex formation by Elk-1, which is a prerequisite for the manifestation of its transactivation potential at the c-fos SRE.
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PMID:ERK phosphorylation potentiates Elk-1-mediated ternary complex formation and transactivation. 788 42

The mitogen-induced early-response gene, PAC-1, encodes a nuclear 32-kDa tyrosine-threonine dual specificity phosphatase, which has been shown to specifically dephosphorylate the mitogen activated protein (MAP) kinases, ERK1 and ERK2. Here, we describe the structure and sequence of the murine PAC-1 gene. Transcription starts at three major sites located between 80 and 90 nucleotides upstream of the murine PAC-1 initiation codon within a highly G/C-rich region. The gene comprises three exons, with exon 1 encoding the unique N-terminal half of the protein, while exons 2 and 3 encode the C-terminus that is homologous to the closely related phosphatases, 3CH134 and VH1. The conserved catalytic domain common to all tyrosine phosphatases is encompassed by exon 3. The organization of the murine PAC-1 gene suggests that the PAC-1 N-terminus, which may serve a regulatory function, has evolved as a separate domain from the C-terminal catalytic domain.
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PMID:Structure of the gene encoding the murine dual specificity tyrosine-threonine phosphatase PAC1. 789 76

We have previously shown that hypoxia causes the activation of nuclear factor-kappa B (NF-kappa B), and the phosphorylation of its inhibitory subunit, I kappa B alpha, on tyrosine residues. With the use of dominant negative mutants of Ha-Ras and Raf-1, we investigated some of the early signaling events leading to the activation of NF-kappa B by hypoxia. Both dominant negative alleles of Ha-Ras and Raf-1 inhibited NF-kappa B induction by hypoxia, suggesting that the hypoxia-induced pathway of NF-kappa B induction is dependent on Ras and Raf-1 kinase activity. Furthermore, although conditions of low oxygen can also activate mitogen-activated protein kinases (ERK1 and ERK2), these kinases do not appear to be involved in regulating NF-kappa B by low oxygen conditions, as dominant negative mutants of mitogen-activated protein kinase do not inhibit NF-kappa B activation by hypoxia. Since Ras and Raf-1 have been previously shown to work downstream from membrane-associated tyrosine kinases such as Src, we determined if the Src membrane-associated kinase was also activated by low oxygen conditions. We detected an increase in Src proto-oncogene activity within 15-30 min of cellular exposure to hypoxia. We postulate that Src activation by hypoxia may be one of the earliest events that precedes Ras activation in the signaling cascade which ultimately leads to the phosphorylation and dissociation of the inhibitory subunit of NF-kappa B, I kappa B alpha.
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PMID:Hypoxic activation of nuclear factor-kappa B is mediated by a Ras and Raf signaling pathway and does not involve MAP kinase (ERK1 or ERK2). 792 53

While testing purines related to the non-specific protein kinase inhibitors N6-dimethylaminopurine and N6-(delta 2-isopentenyl)adenine as potential inhibitors of the p34cdc2/cyclin B kinase, we discovered a compound with high specificity, 2-(2-hydroxyethylamino)-6- benzylamino-9-methylpurine (olomoucine). Kinetic analysis of kinase inhibition reveals that olomoucine behaves as a competitive inhibitor for ATP and as a non-competitive inhibitor for histone H1 (linear inhibition for both substrates). The kinase specificity of this inhibition was investigated for 35 highly purified kinases (including p34cdk4/cyclin D1, p40cdk6/cyclin D3, cAMP-dependent and cGMP-dependent kinases, eight protein kinase C isoforms, calmodulin-dependent kinase II, myosin light-chain kinase, mitogen-activated S6 kinase, casein kinase 2, double-stranded RNA-activated protein kinase, AMP-stimulated kinase, eight tyrosine kinases). Most kinases are not significantly inhibited. Only the cell-cycle regulating p34cdc2/cyclin B, p33cdk2/cyclin A and p33cdk2/cyclin E kinases, the brain p33cdk5/p35 kinase and the ERK1/MAP-kinase (and its starfish homologue p44mpk) are substantially inhibited by olomoucine (IC50 values are 7, 7, 7, 3 and 25 microM, respectively). The cdk4/cyclin D1 and cdk6/cyclin D3 kinases are not significantly sensitive to olomoucine (IC50 values greater than 1 mM and 150 microM, respectively). N6-(delta 2-Isopentenyl)adenine is confirmed as a general kinase inhibitor with IC50 values of 50-100 microM for many kinases. The purine specificity of cyclin-dependent kinase inhibition was investigated: among 81 purine derivatives tested, only C2, N6 and N9-substituted purines exert a strong inhibitory effect on the p34cdc2/cyclin B kinase. An essentially similar sensitivity to this olomoucine family of compounds was observed for the brain-specific cdk5/p35 kinase. Structure/activity relationship studies allow speculation on the interactions of olomoucine and its analogues with the kinase catalytic subunit. Olomoucine inhibits in vitro M-phase-promoting factor activity in metaphase-arrested Xenopus egg extracts, inhibits in vitro DNA synthesis in Xenopus interphase egg extracts and inhibits the licensing factor, an essential replication factor ensuring that DNA is replicated only once in each cell cycle. Olomoucine inhibits the starfish oocyte G2/M transition in vivo. Through its unique selectivity olomoucine provides an anti-mitotic reagent that may preferentially inhibit certain steps of the cell cycle.
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PMID:Inhibition of cyclin-dependent kinases by purine analogues. 792 96

c-Jun transcriptional activity is stimulated by phosphorylation at two N-terminal sites: Ser-63 and -73. Phosphorylation of these sites is enhanced in response to a variety of extracellular stimuli, including growth factors, cytokines, and UV irradiation. New members of the mitogen-activated protein (MAP) kinase group of signal-transducing enzymes, termed JNKs, bind to the activation domain of c-Jun and specifically phosphorylate these sites. However, the N-terminal sites of c-Jun were also suggested to be phosphorylated by two other MAP kinases, ERK1 and ERK2. Despite these reports, we find that unlike the JNKs, ERK1 and ERK2 do not phosphorylate the N-terminal sites of c-Jun in vitro; instead they phosphorylate an inhibitory C-terminal site. Furthermore, the phosphorylation of c-Jun in vivo at the N-terminal sites correlates with activation of the JNKs but not the ERKs. The ERKs are probably involved in the induction of c-fos expression and thereby contribute to the stimulation of AP-1 activity. Our study suggests that two different branches of the MAP kinase group are involved in the stimulation of AP-1 activity through two different mechanisms.
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PMID:c-Jun N-terminal phosphorylation correlates with activation of the JNK subgroup but not the ERK subgroup of mitogen-activated protein kinases. 793 87


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