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)

The IL-7R plays an essential role in gammadelta T cell development by inducing V-J recombination of the TCRgamma locus through STAT5. Although tyrosine residues in the intracellular domain of the mouse IL-7R alpha-chain (IL-7Ralpha) have been implicated in STAT5 activation, it is still unknown whether they are essential for gammadelta T cell development. In this study, we showed that those IL-7Ralpha tyrosine residues are not essential for gammadelta T cell development, because phenylalanine replacement of four intracellular tyrosine residues (IL-7R-FFFF) partially rescued gammadelta T cell development of IL-7Ralpha-/- progenitors. To examine signaling pathways activated by IL-7R-FFFF, we introduced a chimeric receptor consisting of the human IL-4R alpha-chain and mouse IL-7R-FFFF (4R/7R-FFFF) into an IL-7-dependent pre-B cell line and found that 4R/7R-FFFF induced TCRgamma germline transcription and STAT5 activation. Treatment of cells with MEK1/2 inhibitors significantly decreased levels of TCRgamma germline transcription and STAT5 tyrosine phosphorylation mediated by 4R/7R-FFFF, suggesting that MEK1/2 plays an alternative role in STAT5 activation by IL-7R. MEK1/2 associated with STAT5 and induced STAT5 tyrosine phosphorylation and DNA binding activity. Furthermore, MEK1 directly phosphorylated a STAT5 tyrosine residue in vitro. Finally, active MEK1 partially rescued TCRgamma germline transcription by IL-7R in a pre-T cell line. These results demonstrate that MEK1/2 induces TCRgamma germline transcription by phosphorylating STAT5 through IL-7R-FFFF and suggest a potential role for MAPK in IL-7R tyrosine-independent activation of STAT5.
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PMID:MEK1/2 induces STAT5-mediated germline transcription of the TCRgamma locus in response to IL-7R signaling. 1856 15

Expression of p21(Sdi1) downstream of p53 is essential for induction of cellular senescence, although cancer cell senescence can also occur in the p53 null condition. We report herein that senescence-associated phosphorylated extracellular signal-regulated protein kinases 1 and 2 (SA-pErk1/2) enhanced p21(Sdi1) transcription by phosphorylating Sp1 on Ser(59) downstream of protein kinase C (PKC) alpha. Reactive oxygen species (ROS), which was increased in cellular senescence, significantly activated both PKCalpha and PKCbetaI. However, PKCalpha, but not PKCbetaI, regulated ROS generation and cell proliferation in senescent cells along with activation of cdk2, proven by siRNAs. PKCalpha-siRNA also reduced SA-pErk1/2 expression in old human diploid fibroblast cells, accompanied with changes of senescence phenotypes to young cell-like. Regulation of SA-pErk1/2 was also confirmed by using catalytically active PKCalpha and its DN-mutant construct. These findings strongly suggest a new pathway to regulate senescence phenotypes by ROS via Sp1 phosphorylation between PKCalpha and SA-pErk1/2: employing GST-Sp1 mutants and MEK inhibitor analyses, we found that SA-pErk1/2 regulated Sp1 phosphorylation on the Ser(59) residue in vivo, but not threonine, in cellular senescence, which regulated transcription of p21(Sdi1) expression. In summary, PKCalpha, which was activated in senescent cells by ROS strongly activated Erk1/2, and the SA-pErk1/2 in turn phosphorylated Sp1 on Ser(59). Sp1-enhanced transcription of p21(Sdi1) resulted in regulation of cellular senescence in primary human diploid fibroblast cells.
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PMID:Phosphorylated extracellular signal-regulated protein kinases 1 and 2 phosphorylate Sp1 on serine 59 and regulate cellular senescence via transcription of p21Sdi1/Cip1/Waf1. 1931 49

BAD is a proapoptotic member of the Bcl-2 protein family that is regulated by phosphorylation in response to survival factors. Although much attention has been devoted to the identification of phosphorylation sites in murine BAD, little data are available with respect to phosphorylation of human BAD protein. Using mass spectrometry, we identified here besides the established phosphorylation sites at serines 75, 99, and 118 several novel in vivo phosphorylation sites within human BAD (serines 25, 32/34, 97, and 124). Furthermore, we investigated the quantitative contribution of BAD targeting kinases in phosphorylating serine residues 75, 99, and 118. Our results indicate that RAF kinases represent, besides protein kinase A, PAK, and Akt/protein kinase B, in vivo BAD-phosphorylating kinases. RAF-induced phosphorylation of BAD was reduced to control levels using the RAF inhibitor BAY 43-9006. This phosphorylation was not prevented by MEK inhibitors. Consistently, expression of constitutively active RAF suppressed apoptosis induced by BAD and the inhibition of colony formation caused by BAD could be prevented by RAF. In addition, using the surface plasmon resonance technique, we analyzed the direct consequences of BAD phosphorylation by RAF with respect to association with 14-3-3 and Bcl-2/Bcl-X(L) proteins. Phosphorylation of BAD by active RAF promotes 14-3-3 protein association, in which the phosphoserine 99 represented the major binding site. Finally, we show here that BAD forms channels in planar bilayer membranes in vitro. This pore-forming capacity was dependent on phosphorylation status and interaction with 14-3-3 proteins. Collectively, our findings provide new insights into the regulation of BAD function by phosphorylation.
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PMID:Identification of novel in vivo phosphorylation sites of the human proapoptotic protein BAD: pore-forming activity of BAD is regulated by phosphorylation. 1966 65

Drugs of abuse induce behavioral neuroadaptations whose molecular mechanisms, partly known, are crucial to understanding drug addictions. The multifunctional adaptor Fas-associated protein with death domain (FADD) was recently associated with the induction of neuroplasticity. This study investigated the modulation of FADD and MAP kinase signaling, as well as their interactions with PEA-15 (phosphoprotein enriched in astrocytes-15 kDa) and Akt1 pathways, during the expression of unconditioned morphine-induced psychomotor sensitization. In morphine-pretreated rats (10mg/kg during 5 days), a challenge dose of the opiate induced a robust psychomotor sensitization at early withdrawal (3 days, SW 3), but not after a prolonged abstinence period (14 days), which was coincident with an accelerated dopamine turnover in the striatum. Marked concomitant increases in the content of p-FADD (48%) and the activation of MEK-ERK (46-79%) were quantified during the short-term expression of morphine sensitization (SW 3, in the absence of morphine challenge). At SW 3, p-PEA-15, a FADD-ERK binding partner, was also upregulated (51%) as well as the activation of its phosphorylating Akt1 kinase (49%). Notably, the MEK inhibitor SL 327 attenuated (58%) the expression of morphine-induced psychomotor sensitization (SW 3) and fully prevented the upregulation of p-FADD, p-PEA-15 and p-Akt1 at SW 3. The results indicate that the activation of MEK/ERK, the upregulation of p-FADD and that of the linking partners PEA-15/Akt1 have a major role in mediating the short-lasting expression of unconditioned psychomotor sensitization induced by morphine in rats.
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PMID:The time course of unconditioned morphine-induced psychomotor sensitization mirrors the phosphorylation of FADD and MEK/ERK in rat striatum: role of PEA-15 as a FADD-ERK binding partner in striatal plasticity. 1975 90

The mitogen-activated protein kinases MEK/ERK pathway regulates fundamental processes in malignant cells and represents an attractive target in the development of new cancer treatments especially for human hepatocarcinoma highly resistant to chemotherapy. Although gene extinction experiments have suggested distinct roles for these proteins, the MEK/ERK cascade remains widely considered as exhibiting an overlap of functions. To investigate the functionality of each kinase in tumorigenesis, we have generated stably knock-down clones for MEK1/2 and ERK1/2 isoforms in the human hepatocellular carcinoma line HuH7. Our results have shown that RNAi strategy allows a specific disruption of the targeted kinases and argued for the critical function of MEK1 in liver tumor growth. Transient and stable extinction experiments demonstrated that MEK1 isoform acts as a major element in the signal transduction by phosphorylating ERK1 and ERK2 after growth factors stimulation, whereas oncogenic level of ERK1/2 phosphorylation appears to be MEK1 and MEK2 dependent in basal condition. In addition, silencing of MEK1 or ERK2 abolished cell proliferation and DNA replication in vitro as well as tumor growth in vivo after injection in rodent. In contrast, targeting MEK2 or ERK1 had no effect on hepatocarcinoma progression. These results strongly corroborate the relevance of targeting the MEK cascade as attested by pharmacologic drugs and support the potential application of RNAi in future development of more effective cancer therapies. Our study emphasizes the importance of the MEK/ERK pathway in human hepatocarcinoma cell growth and argues for a crucial role of MEK1 and ERK2 in this regulation.
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PMID:RNAi-mediated MEK1 knock-down prevents ERK1/2 activation and abolishes human hepatocarcinoma growth in vitro and in vivo. 1981 36

Extracellular signal-regulated kinase (ERK)1/2 signalling plays a critical role in synaptic and structural plasticity. Recent preclinical and human brain studies suggest that depression and suicidal behaviour are associated with aberrant ERK1/2 signalling. MEK, is a dual-specificity kinase, which is the immediate upstream regulator of ERK1/2. Two isoforms of MEK (MEK1 and MEK2) exist. By phosphorylating at Ser and Thr residues, MEK activates ERK1/2, which then phosphorylates cytoplasmic and nuclear substrates. On the other hand, MEK itself is regulated through phosphorylation by upstream Raf kinases. Recently, we demonstrated that activation of ERK1/2 and B-Raf was attenuated in the brains of suicide subjects. To further investigate the regulation of ERK1/2 signalling, we examined the expression and activation of MEKs, the interaction of MEK with ERKs, MEK-mediated activation of ERK1/2, and ERK1/2-mediated activation of nuclear substrate Elk-1 in the prefrontal cortex and hippocampus of suicide subjects. In addition, in order to investigate whether MEK is regulated by B-Raf, we examined the B-Raf and MEK interaction. No significant changes were observed in expression levels of MEK1 or MEK2; however, the catalytic activity of only MEK1 (not MEK2) was decreased in both the prefrontal cortex and hippocampus of suicide subjects. The interaction of MEK1 with ERK1 and ERK2 was increased along with decreased phosphorylation and catalytic activity of ERK1/2. In addition, we found decreased phosphorylation of MEK1 and less interaction of B-Raf with MEK1. Our results demonstrate abnormalities in MEK1 at multiple levels and suggest that these abnormalities in MEK1 are crucial for aberrant ERK1/2 signalling in suicide brain.
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PMID:Aberrant extracellular signal-regulated kinase (ERK)1/2 signalling in suicide brain: role of ERK kinase 1 (MEK1). 1983 59

During infection, viruses hijack various host cell components and programs for their amplification, among which is the canonical ERK signaling pathway, mainly consisting of three tiered serine/threonine kinases, Raf, MEK and ERK. MEK1 and MEK2 are two isoforms of the kinase operating immediately upstream of ERK, and connecting Raf and ERK by phosphorylating ERK. Previous studies have suggested that different isoforms of MEK have distinct biological functions, although their in vitro kinase function may be redundant. However, little is known about the isoform-specific effects of these kinases on viral propagation. In this study, we showed that herpes simplex virus type 2 (HSV-2) infection of human embryonic kidney (HEK) 293 cells induced a sustained activation of ERK1/2. Inhibition of this ERK activation by U0126, a specific inhibitor of MEK1/2, severely impaired virus production. A similar reduction of virus production was also seen following transfection of cells with siRNAs for MEK1/2. Interestingly, a specific knockdown of MEK1 with siRNAs caused a marked inhibition of viral titers, viral proteins and virus-induced cytopathic effect (CPE), whereas silencing MEK2 had little effect. Therefore, our results demonstrate that MEK1 and MEK2 act differently and that HSV-2 hijacks host MEK1 for its own amplification. To our knowledge, this is the first report showing inhibition of HSV-2 replication by targeting human MEK1. This study also suggests that MEK1 could be a potential target for anti-HSV-2 therapy, which may minimize damage to the host cells engendered by targeting both MEK1 and MEK2.
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PMID:Distinct effects of knocking down MEK1 and MEK2 on replication of herpes simplex virus type 2. 2017 1

Apoptosis signal-regulating kinase 1 (ASK1) is a serine/threonine kinase that responds to a plethora of stress-inducing signals. In turn, activation of ASK1 is associated with a number of human pathological conditions, including neurodegenerative disease, inflammation, and heart failure. In response to oxidative stress, ASK1 activates the cell death-associated p38 MAPK pathway by phosphorylating MKK6. Here, we investigated the regulation of oxidative stress-induced ASK1-catalyzed phosphorylation of MKK6. MKK6 phosphorylation levels increased immediately after H(2)O(2) treatment in intact cells and decreased following treatment for 30 min. When expressed in HEK293T cells, ASK1 was reproducibly purified within a high-molecular mass complex ( approximately 1500 kDa) known as the ASK1 signalosome. Measurement of the in vitro kinetic parameters revealed that the catalytic efficiency (k(cat)/K(m)) of ASK1 was 4000-fold greater in cells treated with H(2)O(2) for 3 min than in untreated cells. Interestingly, although the K(m(ATP)) values were found to be unchanged, the K(m(MKK6)) was dramatically decreased ( approximately 1000-fold). The increased affinity was specific for MKK6 and short-lived, as the K(m(MKK6)) returned to basal levels 30 min after treatment. Consistently, endogenous MKK6 was found within the ASK1 signalosome in intact cells and in addition copurified with ASK1 following treatment for 3 min. In contrast, proteins modulating ASK1 activity and degradation were found to interact with the ASK1 signalosome once MKK6 activation was completed. Taken together, these data suggest that oxidative stress rapidly increases ASK1 catalytic efficiency for MKK6 phosphorylation by increasing MKK6 binding affinity within the ASK1 signalosome prior to induction of inactivation and degradation of the complex.
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PMID:Mechanism of oxidative stress-induced ASK1-catalyzed MKK6 phosphorylation. 2036 19

Infection of a cell by lentiviruses, such as human immunodeficiency virus type 1 or feline immunodeficiency virus, results in the formation of a reverse transcription complex, the pre-integration complex (PIC), where viral DNA is synthesized. In non-dividing cells, efficient nuclear translocation of the PIC requires the presence of the inner nuclear lamina protein emerin (EMD). Here, we demonstrate that EMD phosphorylation is induced early after infection in primary non-dividing cells. Furthermore, we demonstrate that EMD phosphorylation is dependent on virion-associated mitogen-activated protein kinase (MAPK). Specific inhibition of MAPK activity with kinase inhibitors markedly reduced EMD phosphorylation and resulted in decreased integration of the proviral DNA into chromatin. Similarly, when a MEK1 kinase-inactive mutant was expressed in virus-producer cells, virus-induced phosphorylation of EMD was impaired and viral integration reduced during the subsequent infection. Expression of constitutively active MEK1 kinase in producer cells did not result in modulation of EMD phosphorylation or viral integration during subsequent infection. These studies demonstrate that, in addition to phosphorylating components of the PICs at an early step of infection, virion-associated MAPK plays a role in facilitating cDNA integration after nuclear translocation through phosphorylation of target-cell EMD.
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PMID:Lentivirus-associated MAPK/ERK2 phosphorylates EMD and regulates infectivity. 2046 47

The MEK-ERK pathway plays a role in DNA damage response (DDR). This has been thoroughly studied by modulating MEK activation. However, much less has been done to directly examine the contributions of ERK1 and ERK2 kinases to DDR. Etoposide induces G2/M arrest in a variety of cell lines, including MCF7 cells. DNA damage-induced G2/M arrest depends on the activation of the protein kinase ataxia-telangiectasia mutated (ATM). ATM subsequently activates CHK2 by phosphorylating CHK2 threonine 68 (T68) and CHK2 inactivates CDC25C via phosphorylation of its serine 216 (S216), resulting in G2/M arrest. To determine the contribution of ERK1 and ERK2 to etoposide-induced G2/M arrest, we individually knocked-down ERK1 and ERK2 in MCF7 cells using specific small interfering RNA (siRNA). Knockdown of either kinases significantly reduced ATM activation in response to etoposide treatment, and thereby attenuated phosphorylation of the ATM substrates, including the S139 of H2AX (gammaH2AX), p53 S15, and CHK2 T68. Consistent with these observations, knockdown of either ERK1 or ERK2 reduced etoposide-induced CDC25C S216 phosphorylation and significantly compromised etoposide-induced G2/M arrest in MCF7 cells. Taken together, we demonstrated that both ERK1 and ERK2 kinases play a role in etoposide-induced G2/M arrest by facilitating activation of the ATM pathway. These observations suggest that a cellular threshold level of ERK kinase activity is required for the proper checkpoint activation in MCF7 cells.
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PMID:Both ERK1 and ERK2 kinases promote G2/M arrest in etoposide-treated MCF7 cells by facilitating ATM activation. 2063 59


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