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

Global gene expression during the induction of ion pair-mediated apoptosis was evaluated by an apoptosis microarray system. Human bone marrow stromal cells were cultured in the presence of 10(-6) M dexamethasone to promote osteogenesis. After 28 days, these cells expressed elevated alkaline phosphatase activity and maintained Cbfa1 expression even when challenged with an apoptogen. Apoptosis was initiated by treating cells with 3 mM Ca(2+) and 5 mM Pi for 4 h. 32P-Labeled mRNA was hybridized to a human apoptosis microarray containing 205 cDNA fragments. We found that apoptosis influenced the expression of 15 genes mainly involved in cell cycle and cell signaling. These genes included IGFBPs and ERK1, known to play a role in cell survival; GST and GST mu, required for maintenance of thiol redox; TNFR1, a gene product that initiates cell death; and finally, BAD, a gene that encodes a proapoptotic protein. Real-time PCR analysis showed that the expression of ERK1, TNFR1, and GST was modulated by 1.89-, 2.66-, and 1.6 fold after 4 h and by 1-, 1.91-, and 1.5 fold, respectively, after 8 h treatment with the ion pair. In addition, we also measured the expression of Bcl-2 and Bax by quantitative RT-PCR. We noted that these two genes were increased 3.07 and 2.99 fold, respectively, after 8 h treatment with the apoptogen. Results of this study suggest that the ion pair influenced ERK1 and TNFR1 signaling pathways and affected thiol metabolism, whereas Bcl-2 and Bax were expressed at late stages of the death process.
 ...
PMID:Detection of apoptotic gene expression in human osteoblast-like cells by cDNA microarrays. 1292 26

A synthetic K vitamin analogue, 2-(2-mercaptothenol)-3-methyl-1,4-naphthoquinone or Cpd 5, was previously found to be a potent inhibitor of cell growth [Nishikawa et al., (1995) J. Biol. Chem. 270, 28304-28310]. The mechanisms of cell growth were hypothesized to include the inactivation of cellular protein tyrosine phosphatases, especially the Cdc25 family [Tamura et al. (2000) Cancer Res. 60, 1317-1325]. In this study, we synthesized PD 49, a new biotin containing Cpd 5 derivative, to search for evidence of direct interaction of these arylating analogues with Cdc25A, Cdc25B, and Cdc25C phosphatases. PD 49 was shown to directly bind to GST-Cdc25A, GST-Cdc25B, their catalytic fragments, and GST-Cdc25C. The binding could be competed with excess glutathione or Cpd 5, and a cysteine-to-serine mutation of the catalytic cysteine abolished binding. This was consistent with an involvement in binding of cysteine in the catalytic domain. This interaction between PD 49 and Cdc25 also occurred in lysates of treated cells. PD 49 also bound to protein phosphatases other than Cdc25. We found that the new analogue also inhibited Hep3B human hepatoma cell growth. This growth inhibition involved ERK1/2 phosphorylation and was inhibited by a MEK antagonist. The results demonstrate a direct interaction and binding between this growth-inhibiting K vitamin derivative with both purified as well as with cellular Cdc25A, Cdc25B, and Cdc25C.
 ...
PMID:Binding and inhibition of Cdc25 phosphatases by vitamin K analogues. 1295 Jan 76

Receptor activator of NF-kappaB ligand (RANKL) is essential for differentiation and function of osteoclasts. The negative signaling pathways downstream of RANKL are not well characterized. By retroviral transduction of RAW264.7 cells with a dominant negative Src homology 2 domain-containing phosphatase-1 (SHP-1)(C453S), we studied the role of tyrosine phosphatase SHP-1 in RANKL-induced osteoclastogenesis. Over-expression of SHP-1(C453S) significantly enhanced the number of tartrate-resistant acid phosphatase-positive multinuclear osteoclast-like cells in response to RANKL in a dose-dependent manner. RANKL induced the recruitment of SHP-1 to a complex containing TNFR-associated factor (TRAF)6. GST pull down experiments indicated that the association of SHP-1 with TRAF6 is mediated by SHP-1 lacking the two Src homology 2 domains. RANKL-stimulated IkappaB-alpha phosphorylation, IkappaB-alpha degradation and DNA binding ability of NF-kappaB were increased after over-expression of SHP-1(C453S). However, RANKL-induced phosphorylation of mitogen-activated protein kinases, extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38 mitogen-activated protein kinase, was unchanged. In addition, SHP-1 regulated RANKL-stimulated tyrosine phosphorylation of p85 subunit of phosphatidylinositol 3 kinase and the phosphorylation of Akt. Increased numbers of osteoclasts contribute to severe osteopenia in Me(v)/Me(v) mice due to mutation of SHP-1. Like RAW264.7 cells expressing SHP-1(C453S), the bone marrow macrophages of Me(v)/Me(v) mice generated much more osteoclast-like cells than that of littermate controls in response to RANKL. Furthermore compared with controls, RANKL induces enhanced association of TRAF6 and RANK in both RAW264.7 cells expressing SHP-1(C453S) and bone marrow macrophages from Me(v)/Me(v) mice. Therefore, SHP-1 plays a role in signals downstream of RANKL by recruitment to the complex containing TRAF6 and these observations may help to understand the mechanism of osteoporosis in Me(v)/Me(v) mice.
 ...
PMID:Receptor activator of NF-kappa B ligand stimulates recruitment of SHP-1 to the complex containing TNFR-associated factor 6 that regulates osteoclastogenesis. 1450 Jun 59

Phosphatase in activated T cells (PAC-1) is a mitogen-induced early responsive gene. It encodes a 32 kDa tyrosine-threonine dual specificity phosphatase. Constitutive expression of PAC-1 leads to an inhibition of MAP kinase activity in vivo. Such constitutive expression was reported in HTLV-1 infected cell lines. In the present study, we observed the constitutive over-expression of two transcripts related to PAC-1 in large granular lymphocyte (LGL) leukemia. By screening a LGL leukemia cDNA library using the 3' end of a PAC-1 probe, we obtained a clone (clone 8) which retains one and one half introns, excludes two exons, and matches one hundred percent with a DNA sequence on chromosome 2. The deduced amino acid sequence of the predicted protein contains 170 amino acids and is 144 amino acids shorter than PAC-1. When we expressed this protein in Escherichia coli as a GST-fusion protein, a 45 kDa (19 kDa PAC-1 variant+26 kDa GST protein) protein was obtained. The expressed protein was purified to near homogeneity by using a glutathione affinity column. The purified protein did not have any intrinsic phosphatase activity when assayed in vitro. But when this purified protein was added to a phosphatase assay system in combination with a recombinant dual specificity phosphatase, CL100, enhanced phosphatase activity was observed. The significance of the constitutive over-expression and its physiological role of this protein remain to be established in leukemic LGL.
 ...
PMID:Characterization of a variant of PAC-1 in large granular lymphocyte leukemia. 1468 Sep 39

Bim, a "BH3-only" protein, is expressed de novo following withdrawal of serum survival factors and promotes cell death. We have shown previously that activation of the ERK1/2 pathway promotes phosphorylation of Bim(EL), targeting it for degradation via the proteasome. However, the nature of the kinase responsible for Bim(EL) phosphorylation remained unclear. We now show that Bim(EL) is phosphorylated on at least three sites in response to activation of the ERK1/2 pathway. By using the peptidylprolyl isomerase, Pin1, as a probe for proline-directed phosphorylation, we show that ERK1/2-dependent phosphorylation of Bim(EL) occurs at (S/T)P motifs. ERK1/2 phosphorylates Bim(EL), but not Bim(S) or Bim(L), in vitro, and mutation of Ser(65) to alanine blocks the phosphorylation of Bim(EL) by ERK1/2 in vitro and in vivo and prevents the degradation of the protein following activation of the ERK1/2 pathway. We also find that ERK1/2, but not JNK, can physically associate with GST-Bim(EL), but not GST-Bim(L) or GST-Bim(S), in vitro. ERK1/2 also binds to full-length Bim(EL) in vivo, and we have localized a potential ERK1/2 "docking domain" lying within a 27-amino acid stretch of the Bim(EL) protein. Our findings provide new insights into the post-translational regulation of Bim(EL) and the role of the ERK1/2 pathway in cell survival signaling.
 ...
PMID:Extracellular signal-regulated kinases 1/2 are serum-stimulated "Bim(EL) kinases" that bind to the BH3-only protein Bim(EL) causing its phosphorylation and turnover. 1468 Dec 25

It has been shown that glutathione S-transferase pi (GSTpi) interacts with and suppresses the activity of c-Jun NH(2)-terminal kinase (JNK). GST-deficient mice (GSTpi(-/-)) have higher levels of circulating white blood cells, with similar proportions of lymphocytes, monocytes, and granulocytes. Interestingly, a selective expansion of splenic B lymphocytes was observed in GSTpi(-/-) animals but no change in T lymphocytes or natural killer cells. A peptidomimetic inhibitor of GSTpi that disrupts the interaction between GSTpi and JNK mimics in wild type mice the increased myeloproliferation observed in GSTpi(-/-) animals. Until now, the molecular basis for this effect has not been defined. In an in vitro hematopoiesis assay, interleukin-3, granulocyte colony-stimulating factor, and granulocyte/macrophage colony-stimulating factor were more effective at stimulating proliferation of hematopoietic cells in GSTpi(-/-) mice than in wild type. The JNK inhibitor SP600125 which caused little inhibition of cytokine-induced myeloproliferation in wild type mice, decreased the number of colonies in GSTpi(-/-) animals. A more sustained phosphorylation of the STAT family of proteins was also observed in GSTpi(-/-) bone marrow-derived mast cells exposed to interleukin-3. This was associated with an increased proliferation and a down-regulation of expression of negative regulators of the Janus kinase-STAT pathway SHP, Src homology 2 domain-containing tyrosine phosphatase-1 and -2. The increased activation of JNK and STATs in GSTpi-deficient mice provides a viable mechanism for the increased myeloproliferation in these animals. These data also confirm the important role that GSTpi plays in the regulation of cell signaling pathways in a myeloproliferative setting.
 ...
PMID:Increased myeloproliferation in glutathione S-transferase pi-deficient mice is associated with a deregulation of JNK and Janus kinase/STAT pathways. 1468 49

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that plays a role as an intracellular mediator of the xenobiotic signaling pathway. AhR contains signals for both nuclear localization and nuclear export (NES). The objective of this study was to demonstrate how AhR intracellular distribution was regulated physiologically in cells. We found that cell density, but not the cell cycle, influenced the subcellular distribution of AhR in a keratinocyte cell line, HaCaT: AhR was predominantly nuclear at sparse cell densities, both nuclear and cytoplasmic at subconfluence, and predominantly cytoplasmic at confluence. Stable transfectants of HaCaT carrying a reporter gene fused with xenobiotic responsive element showed an association between xenobiotic responsive element-mediated transcription and AhR relocalization. Leptomycin B promoted nuclear accumulation of AhR irrespective of cell density, suggesting that this alteration may be because of a change of the regulation of the nuclear export of AhR. We found that Ser-68 in the NES of AhR was phosphorylated after nuclear accumulation of activated AhR and the nuclear export of a chimeric GST-AhR-GFP fusion protein was suppressed by substitution of a serine residue (Ser-68) to aspartic acid, which mimics the negative charge of phosphorylation. This novel cell density-dependent AhR relocalization was affected by exposure to SB203580, okadaic acid, and low Ca(2+) concentrations. These findings strongly suggest that cell density regulates the intracellular localization and function of AhR, because of modulation of nuclear export activity. The p38 MAPK-mediated phosphorylation of the NES and its dephosphorylation, regulated by cell-cell contact signals, may have pivotal roles in the novel AhR relocalization.
 ...
PMID:Cell density regulates intracellular localization of aryl hydrocarbon receptor. 1498 36

We have previously found that the pi-isozyme of glutathione-S-transferase (GST-pi) is a strong and selective inhibitor of the phosphorylation of the transcriptional activating protein jun by its activating kinase, jun-N-terminal kinase (JNK). We further performed molecular dynamics calculations on the 3-dimensional structure of GST-pi free and bound to an inhibitor that blocks its ability to inhibit the JNK-jun activation. We thus identified 4 putative domains that may be involved in the interaction between GST-pi and the JNK-jun complex: residues 34-50, 99-121, 165-182 (with 2 overlapping sub-domains 165-175 and 169-182), and 194-201. We have synthesized each of these domains and tested them for their abilities to affect the GST-JNK-jun system, first in a cell-free system. We find that peptides corresponding to residues 99-121 and 194-201 strongly inhibit the binding of GST to the JNK-jun complex but do not inhibit JNK-induced phosphorylation of jun, while peptides corresponding to residues 34-50 and 165-182 do not inhibit GST binding but, except for the 165-175 subdomain peptide, strongly inhibit jun phosphorylation. A control peptide, X13, had no effect on either process. Peptide effects on jun phosphorylation appear to be selective for the JNK-jun system since the 34-50 peptide has no effect on other kinase systems (eg, casein kinase, MAP kinase). Three of the domain peptides, 34-50, 165-175, and 194-201 have been attached on their carboxyl-terminal ends to a penetratin sequence, enabling transmembrane transport into cells, and have been introduced into human astrocytes in which JNK was activated with anisomycin. We find that the 34-50-penetratin peptide strongly inhibits intracellular jun phosphorylation while the 194-201-penetratin peptide has no effect; the 165-175-penetratin peptide has a weak effect on this process. Thus, the effects in cells parallel those in the cell-free system. We conclude that all putative domains, identified in our prior structural studies, appear to interact with the JNK-jun complex. The 34-50 peptide may be useful in selectively blocking uncontrolled mitogenic signaling involving the JNK-jun pathway and may be a potential agent for blocking oncogenic ras-p21-induced cell transformation.
 ...
PMID:Effector peptides from glutathione-S-transferase-pi affect the activation of jun by jun-N-terminal kinase. 1503 66

Alien was previously identified as a corepressor for the thyroid hormone receptor (TR) and DAX-1 which belong both to the superfamily of nuclear receptors. Here, we isolated the mixed lineage kinase 2 (MLK2) as an interacting partner for the corepressor Alien using a yeast two hybrid screen. MLK2 is an upstream activator of JNKs and activation of MLK2-mediated signaling cascades play roles in neurodegenerative and apoptotic mechanisms in the central nervous system. MLK2 has been shown to be localized both in the cytoplasm and cell nucleus. We confirmed the Alien-MLK2 interaction using GST pull-down experiments and also show that MLK2 is able to phosphorylate Alien in immune-kinase assays. Functional analyses revealed that Alien, DAX-1 and thyroid hormone receptor mediated transcriptional silencing is strongly enhanced in the presence of active MLK2. Since MAP kinase signaling pathways are important mediators of cellular responses to a wide variety of stimuli, our data suggest that signaling pathways not only regulate transactivation but also enhancement of transcriptional silencing. This novel cross-talk may represent a link between MLK2-mediated signaling and transcriptional repression of target genes during neuronal differentiation processes.
 ...
PMID:Mixed lineage kinase 2 enhances trans-repression of Alien and nuclear receptors. 1506 75

Kinase Suppressor of Ras1 (KSR1) functions as a positive modulator of Ras-dependent signaling either upstream of or parallel to Raf-1, and pharmacologic inactivation of KSR1 may serve as a treatment for Rasdriven malignancies such as pancreatic cancer (Xing, H. R., Cordon-Cardo, C., Deng, X., Tong, W., Campodonico, L., Fuks, Z., and Kolesnick, R. (2003) Nat. Med. 9, 1266-1268). Although some studies demonstrated a requirement for KSR1 kinase activity for its action, others suggested KSR1 acts primarily as a scaffold facilitating assembly of the c-Raf-1/MEK module. We recently established a two-stage in vitro reconstitution assay to measure KSR1 kinase activity (Xing, H. R., Lozano, J., and Kolesnick, R. (2000) J. Biol. Chem. 275, 17276-17280). In this assay, KSR1, immunopurified to apparent homogeneity, never comes in contact with recombinant kinases other than c-Raf-1. In the first assay stage, activated KSR1 is incubated with recombinant c-Raf-1 and ATP. In the second stage, activated c-Raf-1 is separated from KSR1, and incubated with unactivated MEK1, unactivated MAPK, Elk-1, and ATP. Elk-1 phosphorylation serves as a specific readout for MAPK activation. However, because KSR1 constitutively associates with MEK1 and this interaction appears critical for KSR1 scaffolding function, it has been argued that the kinase activity detected is an artifact of KSR1-bound MEK1. To address these concerns, we depleted as much as 90% of KSR1-bound MEK1 by high salt washing without altering KSR1 kinase activity. Further, a complete inactivation of KSR1-bound MEK1 by pretreating with the MEK inhibitor PD 98059 prior to the first assay stage did not alter KSR1 kinase activity. In addition, the omission of exogenous recombinant GST-MEK1 from the reaction mixture during the second assay stage abolished Elk-1 phosphorylation confirming KSR1-bound MEK1 does not support MAPK activation in our in vitro assay. Moreover, a kinase-inactive mutant, FLAG-Ki-KSR1(D683A/D700A), which efficiently interacts with endogenous MEK1, lacks kinase activity. These results collectively support our contention that the kinase activity of KSR1 is an intrinsic property of this protein independent of KSR1-bound endogenous MEK.
 ...
PMID:The kinase activity of kinase suppressor of Ras1 (KSR1) is independent of bound MEK. 2427 36


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>