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)

Stimulation of the erythropoietin (EPO) receptor triggers a cascade of signaling events. We reported that EPO upregulates c-myc expression through 2 pathways in BaF3-EpoR cells--a phosphatidylinositol 3-kinase (PI3K) pathway operating on transcriptional initiation and a Raf-1-mitogen-activated protein kinase (MAPK) pathway affecting elongation. We now show that EPO induces phosphorylation of Raf-1 at serine 338 and within the carboxy-terminal domain, resulting in an electrophoretic mobility change (hyperphosphorylation). Importantly, MEK 1 inhibitor PD98059 blocked only the hyperphosphorylation of Raf-1 but not the phosphorylation at serine 338. This inhibition of Raf-1 hyperphosphorylation resulted in increased kinase activity of Raf-1 and increased phosphorylation of MEK, suggesting that the hyperphosphorylation of Raf-1 inhibits its MEK kinase activity. Deletion of the first 184 amino acids of Raf-1, which are involved in its interaction with Ras, had no effect on EPO-induced phosphorylation. Introducing the dominant-negative N17Ras or GAP had no effect on EPO-induced kinase activity of Raf-1 and ELK activation. N17Ras failed to inhibit ELK activation in another cell line-Rauscher murine erythroleukemia- which expresses the EPO receptor endogenously and differentiates in response to the hormone. These results indicate the presence of a Ras-independent mechanism for Raf-1 and MEK activation in these cells.
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PMID:Erythropoietin regulation of Raf-1 and MEK: evidence for a Ras-independent mechanism. 1502 17

Arsenic trioxide (ATO) induces differentiation and apoptosis of malignant cells in vitro and in vivo and has been used in the treatment of a variety of hematologic malignancies. We found that in NB4 acute promyelocytic and in K562 erythroleukemia cell lines treatment with the MEK1 inhibitors PD98059 and PD184352 greatly enhances apoptotic cell death induced by ATO alone. Combined treatment results in the induction of the p53AIP1 (p53-regulated apoptosis-inducing protein 1) gene in both cell lines. Because NB4 and K562 cell lines carry an inactive p53, we investigated the possible role of p73, a p53 paralogue that has been shown to regulate several p53 target genes including p21, Bax, and p53AIP1. We found that MEK1 inhibitors reduce the levels of dominant-negative (DeltaN) p73 proteins and promote the accumulation of endogenous p73alpha through its transcriptional activation and its tyrosine phosphorylation, resulting in p21 up-regulation and significant inhibition of cell growth. ATO reduces DeltaNp73 levels and promotes a p300-mediated acetylation of endogenous p73, thus favoring cell cycle arrest and apoptosis. Finally, the combined treatment with MEK1 inhibitors and ATO enhances the affinity of phosphoacetylated p73 for the p53AIP1 promoter in vivo, as determined by chromatin immunoprecipitation experiments, leading to p53AIP1 up-regulation and increased apoptosis.
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PMID:Treatment with arsenic trioxide (ATO) and MEK1 inhibitor activates the p73-p53AIP1 apoptotic pathway in leukemia cells. 1503 Dec 5

The ability of the human prostacyclin receptor (hIP) to regulate the activities of signal transducers and activators of transcription (STATs) has not yet been documented. In the present study, we have delineated the mechanism by which hIP induces STAT3 phosphorylations in human erythroleukemia (HEL) cells. Stimulation of endogenous hIP by its specific agonist, cicaprost, resulted in STAT3 Tyr705 and Ser727 phosphorylations in a time- and concentration-dependent manner. Cicaprost-induced STAT3 Tyr705 and Ser727 phosphorylations were resistant to pertussis toxin (PTX) treatment, suggesting that these responses were mediated through PTX-insensitive G proteins. In addition, extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK), but not p38 MAPK, were shown to be phosphorylated by cicaprost in a time- and concentration-dependent manner via PTX-insensitive G proteins. The levels of the interaction between STAT3, ERK and JNK were enhanced by cicaprost treatment. The involvement of Raf-1, MEK1/2 and JNK in cicaprost-induced phosphorylations of STAT3 was illustrated by the use of their selective inhibitors. In contrast, p38 MAPK did not appear to be required. Similar observations were obtained with STAT1 upon stimulation by cicaprost. Taken together, these results demonstrate for the first time that hIP activation by cicaprost can lead to STAT1 and STAT3 phosphorylations via signaling pathways involving PTX-insensitive G proteins, ERK and JNK.
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PMID:Prostacyclin receptor induces STAT1 and STAT3 phosphorylations in human erythroleukemia cells: a mechanism requiring PTX-insensitive G proteins, ERK and JNK. 1597 46

Members of the mitogen-activated protein kinase (MAPK) family, including Jun amino-terminal kinase (JNK) and extracellular signal-related kinase (ERK), play an important role in the proliferation of erythroid cells in response to erythropoietin (Epo). Erythroid cells infected with the Friend spleen focus-forming virus (SFFV) proliferate in the absence of Epo and show constitutive activation of Epo signal transduction pathways. We previously demonstrated that the ERK pathway was constitutively activated in Friend SFFV-infected erythroid cells, and in this study JNK is also shown to be constitutively activated. Pharmacological inhibitors of both the ERK and JNK pathways stopped the proliferation of primary erythroleukemic cells from Friend SFFV-infected mice, with little induction of apoptosis, and furthermore blocked their ability to form Epo-independent colonies. However, only the JNK inhibitor blocked the proliferation of erythroleukemia cell lines derived from these mice. The JNK inhibitor caused significant apoptosis in these cell lines as well as an increase in the fraction of cells in G(2)/M and undergoing endoreduplication. In contrast, the growth of erythroleukemia cell lines derived from Friend murine leukemia virus (MuLV)-infected mice was inhibited by both the MEK and JNK inhibitors. JNK is important for AP1 activity, and we found that JNK inhibitor treatment reduced AP1 DNA-binding activity in primary erythroleukemic splenocytes from Friend SFFV-infected mice and in erythroleukemia cell lines from Friend MuLV-infected mice but did not alter AP1 DNA binding in erythroleukemia cell lines from Friend SFFV-infected mice. These data suggest that JNK plays an important role in cell proliferation and/or the survival of erythroleukemia cells.
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PMID:Activation of the Jun N-terminal kinase pathway by friend spleen focus-forming virus and its role in the growth and survival of friend virus-induced erythroleukemia cells. 1618 78

The effect of ERK, p38, and JNK signaling on p53-dependent apoptosis and cell cycle arrest was investigated using a Friend murine erythroleukemia virus (FVP)-transformed cell line that expresses a temperature-sensitive p53 allele, DP16.1/p53ts. In response to p53 activation at 32 degrees C, DP16.1/p53ts cells undergo p53-dependent G(1) cell cycle arrest and apoptosis. As a result of viral transformation, these cells express the spleen focus forming env-related glycoprotein gp55, which can bind to the erythropoietin receptor (EPO-R) and mimics many aspects of EPO-induced EPO-R signaling. We demonstrate that ERK, p38 and JNK mitogen-activated protein kinases (MAPKs) are constitutively active in DP16.1/p53ts cells. Constitutive MEK activity contributes to p53-dependent apoptosis and phosphorylation of p53 on serine residue 15. The pro-apoptotic effect of this MAPK kinase signal likely reflects an aberrant Ras proliferative signal arising from FVP-induced viral transformation. Inhibition of MEK alters the p53-dependent cellular response of DP16.1/p53ts from apoptosis to G(1) cell cycle arrest, with a concomitant increase in p21(WAF1), suggesting that the Ras/MEK pathway may influence the cellular response to p53 activation. p38 and JNK activity in DP16.1/p53ts cells is anti-apoptotic and capable of limiting p53-dependent apoptosis at 32 degrees C. Moreover, JNK facilitates p53 protein turnover, which could account for the enhanced apoptotic effects of inhibiting this MAPK pathway in DP16.1/p53ts cells. Overall, these data show that intrinsic MAPK signaling pathways, active in transformed cells, can both positively and negatively influence p53-dependent apoptosis, and illustrate their potential to affect cancer therapies aimed at reconstituting or activating p53 function.
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PMID:The involvement of MAPK signaling pathways in determining the cellular response to p53 activation: cell cycle arrest or apoptosis. 1633 May 47

Infection of erythroid progenitor cells by Friend spleen focus-forming virus (SFFV) leads to acute erythroid hyperplasia and eventually to erythroleukemia in susceptible strains of mice. The viral envelope protein, SFFV gp55, forms a complex with the erythropoietin receptor (EpoR) and a short form of the receptor tyrosine kinase Stk (sf-Stk), activating both and inducing Epo-independent proliferation. Recently, we discovered that coexpression of SFFV gp55 and sf-Stk is sufficient to transform NIH 3T3 and primary fibroblasts. In the current study, we demonstrate that sf-Stk and its downstream effectors are critical to this transformation. Unlike SFFV-derived erythroleukemia cells, which depend on PU.1 expression for maintenance of the transformed phenotype, SFFV gp55-sf-Stk-transformed fibroblasts are negative for PU.1. Underscoring the importance of sf-Stk to fibroblast transformation, knockdown of sf-Stk abolished the ability of these cells to form anchorage-independent colonies. Like SFFV-infected erythroid cells, SFFV gp55-sf-Stk-transformed fibroblasts express high levels of phosphorylated MEK, ERK, phosphatidylinositol 3-kinase (PI3K), Gab1/2, Akt, Jun kinase (JNK), and STAT3, but unlike virus-infected erythroid cells they fail to express phosphorylated STATs 1 and 5, which may require involvement of the EpoR. In addition, the p38 mitogen-activated protein kinase (MAPK) stress response is suppressed in the transformed fibroblasts. Inhibition of either JNK or the PI3K pathway decreases both monolayer proliferation and anchorage-independent growth of the transformed fibroblasts as does the putative kinase inhibitor luteolin, but inhibition of p38 MAPK has no effect. Our results indicate that sf-Stk is a molecular endpoint of transformation that could be targeted directly or with agents against its downstream effectors.
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PMID:The tyrosine kinase sf-Stk and its downstream signals are required for maintenance of friend spleen focus-forming virus-induced fibroblast transformation. 1795 67

Human prostacyclin receptor (hIP) stimulates STAT3 via pertussis toxin-insensitive G proteins in human erythroleukemia (HEL) cells. Since hIP can utilize G(s) and G(q) proteins for signal transduction and that both G proteins can induce STAT3 phosphorylation and activation via complex signaling networks, we sought to determine if one of them is predominant in mediating the hIP signal. Stimulation of STAT3 Tyr(705) and Ser(727) phosphorylations by the IP-specific agonist, cicaprost, was sensitive to inhibition of protein kinase A, phospholipase Cbeta, protein kinase C, calmodulin-dependent protein kinase II and Janus kinase 2/3. Unlike Galpha(16)-mediated regulation of STAT3 in the same cells, cicaprost-induced STAT3 Tyr(705) phosphorylation was resistant to inhibition of Src and MEK while STAT3 Ser(727) phosphorylation distinctly required phosphatidylinositol-3 kinase. This unique inhibitor-sensitivity pattern of STAT3 phosphorylation was reproduced in HEL cells by stimulating the G(16)-coupled C5a receptor in the presence of dibutyryl-cAMP, suggesting that the change in inhibitor-sensitivity was due to activation of the G(s) pathway. This postulation was confirmed by expressing constitutively active Galpha(16)QL and Galpha(s)QL in human embryonic kidney 293 cells and the inhibitor-sensitivity of Galpha(16)QL-induced STAT3 phosphorylations could be converted by the mere presence of Galpha(s)QL to resemble that obtained with cicaprost in HEL cells. In addition, the restoration of the Galpha(16)-mediated inhibitor-sensitivity upon cicaprost induction in Galpha(s)-knocked down HEL cells again verified the pivotal role of G(s) signal. Taken together, our observations illustrate that co-stimulation of G(s) and G(q) can result in the fine-tuning of STAT3 activation status, and this may provide the basis for cell type-specific responses following activation of hIP.
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PMID:Prostacyclin receptor-induced STAT3 phosphorylation in human erythroleukemia cells is mediated via Galpha(s) and Galpha(16) hybrid signaling. 1875 67

Galphaq plays a major role in platelet signal transduction, but little is known regarding its transcriptional regulation. We have reported that Galphaq is upregulated during phorbol 12-myristate 13-acetate (PMA)-induced megakaryocytic transformation of human erythroleukemia (HEL) cells and regulated by EGR-1, an early growth transcription factor. These studies focused on the initial 238 bp of the 5' upstream region of the Galphaq gene. In the present studies we characterize a minimal region -1042/-1037 bp from ATG in the 5' upstream of the Galphaq promoter that is associated with PMA responsiveness. In luciferase reporter gene studies in HEL cells, Galphaq 5' upstream promoter sequence -1042/-1 showed an about four-fold increased activity in PMA-treated compared to untreated cells. Deletion of 6-nt -1042/-1037 eliminated the difference. Gel-shift studies on Galphaq probe (-1042/-1012 bp) revealed binding of EGR-1 with PMA-treated but not untreated nuclear extracts, and this was dependent on the sequence -1042/-1037. Silencing of endogenous EGR-1 inhibited Galphaq induction by PMA. MEK/ERK inhibitor U0126 blocked PMA effect on promoter activity of the -1042/-1 construct. In conclusion, EGR-1 binding to sequence -1042/-1037 bp in Galphaq promoter mediates the induction of Galphaq gene by PMA via the MEK/ERK signaling pathway. These studies provide the first evidence of a PMA-responsive element in Galphaq promoter, and new insights into regulation of Galphaq gene by EGR-1.
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PMID:Phorbol 12-myristate 13-acetate (PMA) responsive sequence in Galphaq promoter during megakaryocytic differentiation. Regulation by EGR-1 and MAP kinase pathway. 1898 26

Muscarinic acetylcholine receptors (mAChRs) are members of the superfamily of G protein coupled receptors (GPCRs). Muscarinic receptors are relatively abundant in the central nervous system and in the peripheral parasympathetic nervous system. Several studies have suggested that muscarinic receptors also mediate some cellular events in hematopoietic cells. K562 erythroleukemia cells contain muscarinic receptors M2, M3 and M4, and activation of muscarinic receptors changes cell proliferation. We examined the effects of several compounds on cell proliferation in K562 erythroleukemia cells. These included a muscarinic receptor agonist carbachol (CCh), a protein kinase inhibitor staurosporine; the phospholipase C inhibitor U73122, the MEK 1-2 inhibitor UO126, the PI3-kinase inhibitor wortmannin, the Ca(2+) chelators BAPTA/AM and 2-aminoethoxy-diphenylborate (2APB). In addition, we also investigated muscarinic receptor mediated protein kinase C (PKC) expression in K562 cells. CCh caused a decrease in DNA synthesis in K562 cells supplemented with 1% fetal bovine serum after starvation. Pre-treatment of K562 cells with U73122 and BAPTA/AM antagonized the inhibitory effect of CCh, suggesting that phospholipase C and intracellular calcium are involved in CCh-mediated inhibition of proliferation in K562 cells. Our data also suggest that the regulatory roles of protein kinase C and the MAPK/ERK pathways in K562 cell proliferation are independent of cholinergic activation.
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PMID:The role of intracellular pathways in the proliferation of human K562 cells mediated by muscarinic receptors. 2380 Jul 97

There is increasing evidence showing that many extracellular cues modulate pre-mRNA alternative splicing, through different signaling pathways. We here show that 4.1R exon 16 splicing is altered in response to specific signals. The switch from erythroblastic isoform lacking exon 16 to mature erythrocytic isoform containing this exon is tightly regulated during late erythroid differentiation, and blocage of this splicing switch in erythroleukemia cells is seen as a consequence of the deregulation of important regulatory pathways. We support that combined inhibition of PI3K and activation of p38 signaling pathways impinge on erythroid 4.1R pre-mRNA alternative splicing switch, and on cell differentiation as witnessed by hemoglobin production. By contrast, MEK/ERK signaling appeared not to affect neither cell hemoglobin production nor erythroid 4.1R pre-mRNA splicing. We also found that the signal-induced alternative splicing is not typically distinctive of EPO-non-responsive cells, but operates in EPO-responsive cells as well. Pre-mRNA splicing is a major regulatory mechanism at the crossroad between transcription and translation. We here provide evidence that inhibition of PI3K activates the splicing switch in a promoter-dependent manner, whereas p38 activation induces this event in a promoter-independent fashion. Our data further support that constitutive activation of EPO-R by the viral protein gp55 and the short form of the tyrosine kinase receptor Stk, transduces PI3K proliferation signal, but not MAPK p38 differentiation signal. Concurrently, this work lend credence to the concept that DMSO triggers transient activation of p38 signaling and irreversible inhibition of PI3K/AKT signaling pathway, hence uncovering an old conundrum regarding the mechanism by which DMSO induces erythroleukemia cell differentiation.
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PMID:Combined inhibition of PI3K and activation of MAPK p38 signaling pathways trigger erythroid alternative splicing switch of 4.1R pre-mRNA in DMSO-induced erythroleukemia cells. 2399 58


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