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)

Thrombopoietin (Tpo) is a cytokine regulating megakaryocyte maturation and platelet formation. We studied Tpo-induced signal transduction, and found that Tpo induces phosphorylation of adapter molecules. Shc and Vav, and of serine/threonine kinases Raf-1 and mitogen-activated protein (MAP) kinases. Further, Tpo induced activation of Ras, MAP kinase kinase, MAP kinase and Pim-1. Taken together with other observations, we concluded that Tpo induces the activation of at least two distinct signaling pathways, a specific Tyk2-JAK2/STAT1-STAT3-STAT5 signaling cascade and a common Shc/Vav/Ras/Raf-1/MAP kinase kinase/MAP kinase signaling cascade.
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PMID:Thrombopoietin induces activation of at least two distinct signaling pathways. 854 84

The K562 erythroleukemia cell line was used to study the molecular mechanisms regulating lineage commitment of hematopoietic stem cells. Phorbol esters, which initiate megakaryocyte differentiation in this cell line, caused a rapid increase in extracellular-signal-regulated kinase (ERK), which remained elevated for 2 h and returned to near-basal levels by 24 h. In the absence of extracellular stimuli, ERK could be activated by expression of constitutively active mutants of mitogen-activated protein (MAP) kinase kinase (MKK), resulting in cell adhesion and spreading, increased cell size, inhibition of cell growth, and induction of the platelet-specific integrin alphaIIb beta3, all hallmarks of megakaryocytic differentiation. In contrast, expression of wild-type MKK had little effect. In addition, constitutively active MKK suppressed the expression of an erythroid marker, alpha-globin, indicating the ability to suppress cellular responses necessary for alternative cell lineages. The MKK inhibitor PD98059 blocked MKK/ERK activation and cellular responses to phorbol ester, demonstrating that activation of MKK is necessary and sufficient to induce a differentiation program along the megakaryocyte lineage. Thus, the MAP kinase cascade, which promotes cell growth and proliferation in many cell types, instead inhibits cell proliferation and initiates lineage-specific differentiation in K562 cells, establishing a model system to investigate the mechanisms by which this signal transduction pathway specifies cell fate and developmental processes.
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PMID:Megakaryocytic differentiation induced by constitutive activation of mitogen-activated protein kinase kinase. 912 42

Thrombopoietin (Tpo) is a cytokine which stimulates megakaryocyte maturation. We found that Tpo is constitutively and ubiquitously expressed in all tissues examined, including bone marrow stromal cells, even in thrombocytopenia, thrombosis and steady-state condition in mice. Thus, platelet level in circulation is not regulated by Tpo gene expression. Furthermore, when the purified megakaryocytes were cocultured with the stromal cells, most of the megakaryocytes adhered to the stromal cells and remained unchanged, while free megakaryocytes induced proplatelet formation. Thus the stromal cells in bone marrow secrete Tpo and stimulate megakaryocytopoiesis, but the interaction of megakaryocytes with the stromal cells may suppress platelet formation. Study on signal transduction through Mp1 revealed that Tpo induces activation of JAK2 and Tyk2, which in turn activate STAT1, STAT3 and STAT5. Further, Tpo stimulates transcription factors GATA-1 and NF-E2, which induce differentiation markers, GPIIb/IIIa and Pm-1. In addition, Shc, Vav, Ras, Raf-1, MAPKK, MAPK and Pim-1 are also activated. Thus, Tpo activates a lineage-specific cascade as well as a specific JAK-STAT cascade and a common signaling cascade.
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PMID:Regulation of megakaryocytopoiesis by thrombopoietin and stromal cells. 920 16

Erythroid and megakaryocyte lineages are closely linked and may share a common bipotent progenitor. However, the mechanisms associated with cell lineage commitment are not fully understood. The K562 erythroleukemia cell line serves as a model to study the biochemical changes associated with erythroid and megakaryocyte (E/M) differentiation. We have previously established that PMA-induced megakaryocyte differentiation of K562 cells requires the activity of the MEK/MAPK pathway (Herrera et al Exp Cell Res 1998; 238: 407-414). Here, we show that the PMA-induced phenotypic changes of K562 cells such as polylobulation of the nucleus and Pyk2 expression are independent of MAPK activation. In addition, we also demonstrate that inhibition of the basal activity of the extracellular regulated kinase (ERK/MAPK) pathway enhances the erythroid phenotype of these cells. These results suggest that the MAPK pathway regulates the E/M lineage commitment of K562 cells.
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PMID:PMA-induced phenotypic changes in K562 cells: MAPK-dependent and -independent events. 984 25

Thrombopoietin (TPO) plays a critical role in megakaryocyte proliferation and differentiation. Using various cultured cell lines, several recent studies have implicated the mitogen-activated protein kinase (MAPK) pathway in megakaryocyte differentiation. In the study reported here, we examined the role played by thrombopoietin-induced MAPK activity in a cytokine-dependent cell line (BAF3/Mpl) and in primary murine megakaryocytes. In both systems, extracellular signal-regulated protein kinase (ERK) 1 and 2 MAPK phosphorylation was rapidly induced by TPO stimulation. To identify the Mpl domain responsible for MAPK activation, BAF3 cells expressing truncated forms of the Mpl receptor were studied. Phosphorylation of ERKs did not require elements of the cytoplasmic signaling domain distal to Box 2 and was not dependent on phosphorylation of the adapter protein Shc. ERK activation in murine megakaryocytes was maximal at 10 minutes and was markedly decreased over the subsequent 3 hours. Next, the physiologic consequences of MAPK inhibition were studied. Using the MAPK kinase (MEK) inhibitor, PD 98059, blockade of MAPK activity substantially reduced TPO-dependent proliferation in BAF3/Mpl cells and markedly decreased mean megakaryocyte ploidy in cultures. To exclude an indirect effect of MAPK inhibition on stromal cells in whole bone marrow, CD41(+) cells were selected and then cultured in TPO. The number of polyploid megakaryocytes derived from the CD41-selected cells was also significantly reduced by MEK inhibition, as was their geometric mean ploidy. These studies show an important role for MAPK in TPO-induced endomitosis and underscore the value of primary cells when studying the physiologic effects of signaling pathways.
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PMID:Thrombopoietin-induced activation of the mitogen-activated protein kinase (MAPK) pathway in normal megakaryocytes: role in endomitosis. 1043 15

Phorbol 12-myristate 13-acetate (PMA)-induced differentiation of human erythroleukemic K562 cells is characterized by growth arrest, morphological change, and expression of megakaryocyte-specific proteins. We examined the possible involvement of cell cycle regulators with PMA-induced growth arrest and megakaryocytic differentiation of K562 cells. The concentrations of cyclin D1 and p21Waf1/Cip1 were dramatically increased, whereas those of cyclin B1 and cdc2 were decreased, by PMA treatment. The concentrations of most cyclin-dependent kinases (Cdk2, Cdk4, and Cdk6), however, were unchanged by PMA treatment. PD98059, a specific inhibitor of MEK1, partially prevented the increase in cyclin D1 caused by PMA and fully reversed the down-regulation of cyclin B1 protein seen in response to PMA treatment. Thus, it is demonstrated here that the PMA-mediated changes of cyclin D1 and B1 are the result of a persistent increase in extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) activity.
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PMID:ERK/MAPK pathway is required for changes of cyclin D1 and B1 during phorbol 12-myristate 13-acetate-induced differentiation of K562 cells. 1068 62

Megakaryocyte differentiation is often accompanied by the changes of gene expression pattern. Here we reported that the expression of DAB2, a putative adaptor protein in cell signaling, was induced at the protein and mRNA levels upon 12-O-tetradecanoylphorbol-13-acetate-mediated megakaryocyte differentiation of human chronic myeloid leukemic K562 cells. On the other hand, the differentiation agents DMSO and retinoic acid had no effect on DAB2 expression. Analysis of promoter activity with the human DAB2 luciferase reporter constructs suggested that the regulation is partially at the transcriptional level. The responsive sequences located within an 80-bp DAB2 promoter region. To determine the involvement of MEK1-p42/p44 MAPK pathway in mediating DAB2 gene expression, we have performed the following experiments and found that (i) there was sustained activation of p42/p44 MAPK, but not p38 MAPK, upon K562 cells differentiation; (ii) application of MEK1 inhibitor U0126 reduced the expression of DAB2 protein, mRNA and promoter activity, as well as cell differentiation; (iii) constitutively active MEK1 increased DAB2 promoter activity; and (iv) dominant negative ERK2 abolished constitutively active MEK1-induced DAB2 promoter activity. Taken together, our results indicate that DAB2 gene is induced upon megakaryocyte differentiation by the MEK1-p42/p44 MAPK pathway and may define a new role of DAB2 in hematopoietic cell differentiation.
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PMID:Induction of disabled-2 gene during megakaryocyte differentiation of k562 cells. 1143 82

Thrombopoietin (TPO) plays a critical role not only in proliferation and differentiation of megakaryocytes but also in erythroid differentiation. We have investigated whether the different pathway of mitogen-activated protein kinase (MAPK) after TPO stimulation may discriminate megakaryocyte and erythroid differentiation. In this study, we have used human CD34+ hematopoietic progenitor cells (HPCs) from cord blood (CB) in serum-free liquid culture supplemented with TPO, to compare the respective effects of specific inhibitors of MAPK kinase (MEK) (PD98059) and p38 MAP kinase (p38) (SB203580) on megakaryocyte and erythroid development. PD98059, but not SB203580, significantly suppressed TPO-induced megakaryocyte differentiation when examined by the expression of CD41 and polyploidy assay. In the presence of SB203580, CD34+/CD36+ erythroid progenitors clearly decreased, whereas they increased when cultured with PD98059. These results indicate that activation of extracellular-signal-regulated kinase (ERK) is required for TPO-induced megakaryocyte differentiation and that p38 is required for TPO-induced erythroid differentiation.
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PMID:Requirement of thrombopoietin-induced activation of ERK for megakaryocyte differentiation and of p38 for erythroid differentiation. 1144 31

Extracellular signal-regulated kinases 1 and 2 (ERK1/2) are a group of kinases that play an important role in proliferation and differentiation. In megakaryocyte-like human erythroleukemia (HEL) cells, ERK2 was found to be predominantly expressed and strongly activated by prostaglandin (PG) E(2), thrombin, and epinephrine. On the other hand, adenosine, ADP, ATP, and UTP did not significantly increase ERK1/2 phosphorylation. However, of the agonists tested, only ADP was able to stimulate thymidine uptake. Pretreatment with pertussis toxin abolished the PGE(2) response but had less of an effect on thrombin. PGE(2)- and thrombin-induced ERK1/2 activation was mimicked by 4-beta-phorbol-12-myristate-13-acetate and ionomycin and blocked by mitogen-activated protein kinase kinase inhibitor 1,4 diamino-2,3-dicyano-1,4-bis[2-aminophenylthio]butadiene but displayed differential sensitivity to protein kinase C inhibitor bisindolylmaleimide I and Ca(2+) chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid. Analogs of cAMP or agents that stimulate cAMP production were either weak or ineffective activators. Further studies indicate that the effect of thrombin was blocked by the phosphoinositide 3-kinase inhibitor wortmannin but not by agents inhibiting tyrosine kinase activity. On the contrary, herbimycin, but not wortmannin, attenuated the effect of PGE(2). Collectively, these results indicate that ERK1/2 are selectively activated by G protein-coupled receptors and not functionally associated with proliferation in HEL cells. ERK1/2 activation in response to PGE(2) and thrombin is mediated by distinctive types of G proteins and is differentially regulated by multiple pathways, including calcium mobilization, protein kinase C, phosphoinositide 3-kinase, and tyrosine kinases.
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PMID:Extracellular signal-regulated kinases and g protein-coupled receptors in megakaryocytic human erythroleukemia cells: selective activation, differential regulation, and dissociation from mitogenesis. 1175 34

Thrombopoietin stimulates extracellular signal-related kinase 1/2 (ERK1/2) phosphorylation in megakaryocytes, and the classic mitogen-activated protein (MAP) kinase (Raf/mitogen-induced extracellular kinase [MEK]/ERK) pathway has been implicated directly and indirectly to play a critical role in megakaryocytopoiesis. However, the involvement of specific Raf family members in megakaryocytopoiesis is unknown. raf-1(-/-) mice were therefore used to directly determine the role of Raf-1 in megakaryocytopoiesis. Surprisingly, raf-1(-/-) mice have a modestly higher platelet count than their raf-1(+/+) littermates. Nonetheless, the absence of Raf-1 does not alter thrombopoietin-induced expansion of primary megakaryocyte-lineage cells, the development of apoptotic megakaryocytes in the presence or absence of thrombopoietin, or the development of megakaryocyte DNA ploidy distribution. Moreover, raf-1(-/-) megakaryocytes do not have a compensatory increase in A-Raf or B-Raf expression, and thrombopoietin-induced ERK1/2 phosphorylation is similar in raf-1(-/-) and raf-1(+/+) megakaryocytes. These unexpected findings demonstrate that Raf-1 is dispensable for megakaryocytopoiesis, and for thrombopoietin-induced ERK1/2 activation in primary megakaryocyte-lineage cells.
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PMID:Raf-1 is not required for megakaryocytopoiesis or TPO-induced ERK phosphorylation. 1457 68


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