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)

The P-450 side chain cleavage (CYP11A1) gene encodes the enzyme that catalyzes the initial step in steroid biosynthesis, resulting in the conversion of cholesterol to pregnenolone. Expression of the CYP11A1 gene is increased by hormones, such as adrenocorticotropin and luteinizing hormone, as well as by a number of growth factors, suggesting that its promoter may contain regulatory elements that respond to multiple signal transduction pathways. Using transient expression assays of the ovine CYP11A1 promoter in JEG-3 placental cells, distinct regulatory elements were found to mediate transcriptional stimulation by cAMP and epidermal growth factor (EGF). The cAMP response was mediated through a GC-rich sequence localized between -117 and -92. In contrast, EGF induced CYP11A1 transcription through an adjacent but distinct sequence (-92 to -77 base pairs) that was shown previously to bind nuclear proteins in DNase I footprinting reactions. This EGF-responsive element (EGF-RE) resembles an activator protein-1 (AP-1) site and was also required for transactivation by co-transfected c-Jun. A point mutation within the EGF-RE impaired stimulation by both EGF and c-Jun, suggesting that these pathways converge on a common regulatory element. Transfer of single or multiple copies of the EGF-RE upstream of an heterologous promotor conferrd EGF and c-Jun responses, providing further evidence that this element is sufficient for both responses. Transfection studies employing mutant c-Jun proteins confirmed a requirement for its DNA binding, leucine zipper and amino-terminal domains, each of which are required for activation of a classical AP-1 reporter. Gel shift studies demonstrated that protein binding to the CYP11A1 EGF-RE was competed specifically by a canonical AP-1 site, and the addition of an anti-JUN antibody confirmed the presence of AP-1 proteins. Consistent with the possibility that EGF may act in part via c-Jun, EGF stimulated the activity of a chimeric GAL4 c-Jun protein, indicating that JUN can serve as a potential target of EGF in JEG-3 cells. EGF also induced mitogen-activated protein kinase activity, and a dominant negative mutant of mitogen-activated protein kinase partially blocked EGF stimulation of GAL4 c-Jun activity. We conclude that EGF stimulates the CYP11A1 promoter through an AP-1 like element and that c-Jun is one of the targets of EGF action.
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PMID:Epidermal growth factor and c-Jun act via a common DNA regulatory element to stimulate transcription of the ovine P-450 cholesterol side chain cleavage (CYP11A1) promoter. 762 50

The 92 kDa type IV collagenase (MMP-9), which degrades type IV collagen, has been implicated in tissue remodeling. The purpose of the current study was to determine the role of Jun amino-terminal kinase (JNK)- and extracellular signal-regulated kinase- (ERK)-dependent signaling cascades in the regulation of MMP-9 expression. Towards this end, we first determined the transcriptional requirements for MMP-9 promoter activity in a cell line (UM-SCC-1) which is an avid secretor of this collagenase. Transfection of these cells with a CAT reporter driven by progressive 5' deleted fragments of the MMP-9 promoter indicated the requirement of a region spanning -144 to -73 for optimal promoter activity. DNase I footprinting revealed a protected region of the promoter spanning nucleotides -91 to -68 and containing a consensus AP-1 motif at -79. Mutation of this AP-1 motif practically abolished the activity of the MMP-9 promoter-driven CAT reporter. Mobility shift assays indicated c-Fos and Jun-D bound to this motif and transfection of the cells with a mutated c-Jun, which quenches the function of endogenous Jun and Fos proteins, decreased MMP-9 promoter activity by 80%. UM-SCC-1 cells contained a constitutively activated JNK and the expression of a kinase-deficient JNK1 reduced the activity of a CAT reporter driven either by the MMP-9 promoter or by three tandem AP-1 repeats upstream of a thymidine kinase minimal promoter. Conditioned medium collected from UM-SCC-1 cells transfected with the dominant negative JNK1 expression vector diminished 92 kDa gelatinolysis. Similarly, interfering with MEKK, which lies upstream of JNK1, using a dominant negative expression vector reduced MMP-9 promoter activity over the same concentration range which repressed the AP-1-thymidine kinase CAT reporter construct. UM-SCC-1 cells also contained a constitutively activated ERK1. MMP-9 expression, as determined by CAT assays and by zymography, was reduced by the co-expression of a kinase-deficient ERK1. Interfering with MEK1, which is an upstream activator of ERK1, either with PD 098059, which prevents the activation of MEK1, or with a dominant negative expression construct, reduced 92 kDa gelatinolysis and MMP-9 promoter activity respectively. c-Raf-1 is an upstream activator of MEK1 and a kinase-deficient c-Raf-1 expression construct decreased the activity of a promoter driven by either the MMP-9 promoter or three tandem AP-1 repeats. Conversely, treatment of UM-SCC-1 cells with PMA, which activates c-Raf-1, increased 92 kDa gelatinolysis. These data suggest that MMP-9 expression in UM-SCC-1 cells, is regulated by JNK- and ERK-dependent signaling pathways.
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PMID:Regulation of 92 kDa type IV collagenase expression by the jun aminoterminal kinase- and the extracellular signal-regulated kinase-dependent signaling cascades. 913 92

To determine whether M2 muscarinic receptors are linked to the monomeric G protein Rho, we studied the effect of carbachol on actin reorganization (stress fiber formation) in cultured human airway smooth muscle cells that expressed mainly M2 muscarinic receptors by dual-fluorescence labeling of filamentous (F) and monomeric (G) actin. F-actin was labeled with FITC-labeled phalloidin, and G-actin was labeled with Texas Red-labeled DNase I. Carbachol stimulation induced stress fiber formation (increased F-actin staining) in the cells and increased the F- to G-actin ratio 3.6 +/- 0.4-fold (mean +/- SE; n = 5 experiments). Preincubation with pertussis toxin, Clostridium C3 exoenzyme, or tyrosine kinase inhibitors reduced the carbachol-induced increase in stress fiber formation and significantly decreased the F- to G-actin ratio, whereas a mitogen-activated protein kinase inhibitor, a phosphatidylinositol 3-kinase inhibitor, and a protein kinase C inhibitor were without effect. This study demonstrates that in cultured human airway smooth muscle cells, muscarinic-receptor activation induces stress fiber formation via a pathway involving a pertussis-sensitive G protein, Rho proteins, and tyrosine phosphorylation.
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PMID:Carbachol-induced actin reorganization involves Gi activation of Rho in human airway smooth muscle cells. 961 96

We have previously shown that the maximal promoter activity of the gene for extracellular signal-regulated protein kinase 2 (ERK2; also known as p42 mitogen-activated protein kinase) resides in the 371 bp 5'-flanking sequence. In the present study we defined roles for a CCAAT box and two adjacent GC boxes in the activity of this promoter. Deletion analysis and DNase I footprinting of this 371 bp region indicated that the CCAAT box at -64 and GC boxes at -86 and -39 are crucial for promoter activity. Electrophoretic mobility-shift assays showed that transcription factor NF-Y/CBF binds to the CCAAT box. Sp1 and Sp3, members of the Sp family of transcription factors, bind to the GC boxes of the ERK2 promoter. The binding of Sp3 was predominant over that of Sp1. Disruption by mutation of any of the CCAAT box and GC boxes similarly decreased promoter activity. These three cis elements exhibited a moderate synergy in promoter function. The transactivating role of NF-Y was corroborated by the finding that a dominant-negative form of NF-YA diminished the promoter activity. These results provide clues for refining our understanding of not only the regulation of expression of the gene for ERK2 but also mechanisms by which NF-Y and Sp1/Sp3 regulate transcription.
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PMID:Regulation of the gene promoter for extracellular signal-regulated protein kinase 2 by transcription factors NF-Y and Sp3. 1072 14

Transformation of normal cloned rat embryo fibroblast (CREF) cells with cellular oncogenes results in acquisition of anchorage-independent growth and oncogenic potential in nude mice. These cellular changes correlate with an induction in the expression of a cancer progression-promoting gene, progression elevated gene-3 (PEG-3). To define the mechanism of activation of PEG-3 as a function of transformation by the Ha-ras and v-raf oncogenes, evaluations of the signaling and transcriptional regulation of the approximately 2.0 kb promoter region of the PEG-3 gene, PEG-Prom, was undertaken. The full-length and various mutated regions of the PEG-Prom were linked to a luciferase reporter construct and tested for promoter activity in CREF and oncogene-transformed CREF cells. An analysis was also performed using CREF cells doubly transformed with Ha-ras and the Ha-ras specific suppressor gene Krev-1, which inhibits the transformed phenotype in vitro. These assays document an association between expression of the transcription regulator PEA3 and PEG-3. The levels of PEA3 and PEG-3 RNA and proteins are elevated in the oncogenically transformed CREF cells, and reduced in transformation and tumorigenic suppressed Ha-ras/Krev-1 doubly transformed CREF cells. Enhanced tumorigenic behavior, PEG-3 promoter function and PEG-3 expression in Ha-ras transformed cells were all dependent upon increased activity within the mitogen-activated protein kinase (MAPK) pathway. Electrophoretic mobility shift assays and DNase I footprinting experiments indicate that PEA3 binds to sites within the PEG-Prom in transformed rodent cells in an area adjacent to the TATA box in a MAPK-dependent fashion. These findings demonstrate an association between Ha-ras and v-raf transformation of CREF cells with elevated PEA3 and PEG-3 expression, and they implicate MAPK signaling via PEA3 as a signaling cascade involved in activation of the PEG-Prom.
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PMID:PEA3 sites within the progression elevated gene-3 (PEG-3) promoter and mitogen-activated protein kinase contribute to differential PEG-3 expression in Ha-ras and v-raf oncogene transformed rat embryo cells. 1129 38

The actin cytoskeleton is important for stress fiber formation and contributes to the initiation and maintenance of smooth muscle contraction. To determine if oxytocin and lysophosphatidic acid (LPA) induce stress fiber formation, cultured human myometrial cells were exposed to oxytocin (10(-5) M) or LPA (10(-6) M), and filamentous (F) and globular (G) actin pools were stained with fluorescein isothiocyanate-phalloidin and Texas red DNase I, respectively. The F- to G-actin fluorescent-staining ratio was measured by fluorescence microscopy. Oxytocin and LPA increased stress fiber formation, as indicated by an increase in the F- to G-actin fluorescent-staining ratio. The Rho-kinase inhibitor Y-27632 markedly attenuated this increase. Oxytocin-induced stress fiber formation was completely inhibited in the presence of the oxytocin antagonist compound VI. Tyrosine kinase inhibition with tyrphostin A23 partially blocked the increase induced by oxytocin but had no effect on LPA-induced stress fiber formation. Stress fiber formation was not blocked by pertussis toxin, mitogen-activated protein kinase, or protein kinase C inhibitors. Our results show that human myometrial cells respond to oxytocin and LPA with the formation of stress fibers that may be involved in the maintenance of uterine contractions. Rho-kinase appears to be a key signaling factor in this pathway.
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PMID:Oxytocin and lysophosphatidic acid induce stress fiber formation in human myometrial cells via a pathway involving Rho-kinase. 1146 6

MKP-M is a dual specificity phosphatase that preferentially inactivates JNK. mkp-M gene expression is rapidly induced by lipopolysaccharide (LPS) stimulation in macrophages and is involved in the negative regulation of LPS-mediated JNK activation and tumor necrosis factor-alpha secretion. To reveal the transcriptional regulation of the mkp-M gene, we isolated the mouse mkp-M gene and mapped its transcriptional start site. Luciferase reporter plasmids containing 5'-upstream regions of the mkp-M gene were stably transfected into RAW264.7 cells. The assays using these cells revealed that the promoter region between -252 and -135 is required for mkp-M promoter activation. Sequencing analysis revealed E box and CREB-responsive elements in this region, and electromobility shift assays and mutagenesis confirmed that both of these elements are essential for LPS responsiveness of the mkp-M gene. We also utilized chromatin immunoprecipitation assay and found that LPS stimulation caused acetylation of histone H3 and H4 at mkp-M promoter in RAW264.7 cells. Consistent with this, a histone deacetylase inhibitor, trichostatin A, increased endogenous mkp-M gene transcription. Finally, DNase I hypersensitivity site mapping revealed the inducible hypersensitivity site after LPS stimulation around the location of the E box and CREB-responsive elements. Altogether, our data indicated that the activation of mkp-M gene transcription in macrophages by LPS is associated with histone acetylation and chromatin remodeling.
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PMID:Histone acetylation and activation of cAMP-response element-binding protein regulate transcriptional activation of MKP-M in lipopolysaccharide-stimulated macrophages. 1251 74

In a previous study, we showed that isoproterenol induced actin depolymerization in human airway smooth muscle cells by both protein kinase A (PKA)-dependent and -independent signaling pathways. We now investigate the signaling pathway of PKA-independent actin depolymerization induced by isoproterenol in these cells. Cells were briefly exposed to isoproterenol or PGE(1) in the presence and absence of specific inhibitors of Src-family tyrosine kinases, phosphatidylinositol-3-kinase (PI3 kinase), or MAP kinase, and actin depolymerization was measured by concomitant staining of filamentous actin with FITC-phalloidin and globular actin with Texas red DNase I. Isoproterenol, cholera toxin, and PGE(1) induced actin depolymerization, indicated by a decrease in the intensity of filamentous/globular fluorescent staining. Pretreatment with the Src kinase inhibitors 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyriimidine (PP2) or geldanamycin or the PKA inhibitor Rp-cAMPS only partly inhibited isoproterenol- or PGE(1)-induced actin depolymerization. In contrast, PP2 and geldanamycin did not inhibit forskolin-induced actin depolymerization, and AG-213 (an EGF receptor tyrosine kinase inhibitor) did not inhibit isoproterenol- or PGE(1)-induced actin depolymerization. PI3 kinase or MAP kinase inhibition did not inhibit isoproterenol-induced actin depolymerization. Moreover, isoproterenol but not forskolin induced tyrosine phosphorylation of an Src family member at position 416. These results further confirm that both PKA-dependent and PKA-independent pathways mediate actin depolymerization in human airway smooth muscle cells and that the PKA-independent pathway by which isoproterenol induces actin depolymerization in human airway smooth muscle cells involves Src protein tyrosine kinases and the G(s) protein.
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PMID:Isoproterenol induces actin depolymerization in human airway smooth muscle cells via activation of an Src kinase and GS. 1582 Oct 21

Increased expression of the IGF-I receptor (IGF-IR) is associated with proliferation and survival of vascular smooth muscle cells (VSMCs). In cultured VSMCs, we reported that angiotensin II (Ang II) increases transcription and expression of IGF-IR. Now, we show that mesenteric arteries of rats infused with Ang II develop thickening and increased IGF-IR expression. To determine how Ang II transcriptionally regulates IGF-IR expression in VSMCs, we generated 5'-end deletions of the IGF-IR promoter and measured Ang II-induced promoter-luciferase activity in VSMCs. Activities from these promoter sequences suggested that the Ang II-responsive region is located between -270 and -135 of the IGF-IR promoter. Using a DNase I foot printing analysis, we identified two putative nuclear factor-kappaB (NF-kappaB)-like sequences located in the same region of the IGF-IR promoter. When we mutated either of these NF-kappaB-like sites, Ang II-induced IGF-IR promoter activity decreased sharply. Electrophoretic mobility gel shift, anti-p50 of NF-kappaB supershift and chromatin immunoprecipitation assays demonstrated that both the p65 and p50 subunits of NF-kappaB will bind to this Ang II response element in the IGF-IR promoter. When we blocked the Ras/MAPK kinase 1 pathway or the inhibitory-kappaB kinase pathway, both Ang II-induced IGF-IR promoter activity and expression of IGF-IR protein significantly declined. Our results indicate that the mechanism by which Ang II stimulates IGF-IR expression in VSMCs involves NF-kappaB binding to NF-kappaB sites in the IGF-IR promoter, leading to expression of IGF-IR through both Ras/MAPK kinase 1-and inhibitory-kappaB kinase-dependent pathways. Because IGF-IR is a major factor associated with thickening of mesenteric vessels, our results provide potential therapeutic targets.
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PMID:Angiotensin II stimulates transcription of insulin-like growth factor I receptor in vascular smooth muscle cells: role of nuclear factor-kappaB. 1632 63

Caffeine treatment of ovine oocytes increases the activity of maturation-promoting factor (MPF) and mitogen-activated protein kinases (MAPKs) and, in somatic cell nuclear transfer (SCNT) embryos, increases the frequency of nuclear envelope breakdown (NEBD) and premature chromosome condensation (PCC). At the blastocyst stage, caffeine-treated SCNT embryos have increased cell numbers. One explanation for this is that NEBD and PCC release chromatin-bound somatic factors, allowing greater access of oocyte factors involved in DNA synthesis and nuclear reprogramming to donor chromatin. This could advance DNA replication and cleavage in the first cell cycle, resulting in increased cell numbers. Alternatively, increased MAPK activity may affect localisation of heat shock proteins (HSPs) and reduce apoptosis. To investigate these possibilities, we investigated chromatin accessibility, the timing of DNA synthesis and first cleavage, the localisation of HSP27 during early development and the frequency of apoptotic nuclei at the blastocyst stage. Compared with control SCNT (non-caffeine treatment), caffeine treatment (10 mM caffeine for 6 h prior to activation) increased the accessibility of DNase I to donor chromatin (P < 0.05 at 1.5 h post activation (h.p.a.)), advanced DNA synthesis (43.5% v. 67.6%, respectively; P < 0.01 at 6 h.p.a.) and first cleavage (27.3% v. 40.5% at 20 h.p.a., respectively) and increased nuclear localisation of HSP27. Although development to the blastocyst stage was not affected, caffeine increased total cell numbers (98.5 v. 76.6; P < 0.05) and reduced the frequency of apoptotic nuclei (11.27% v. 20.3%; P < 0.05) compared with control SCNT group.
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PMID:Treatment of ovine oocytes with caffeine increases the accessibility of DNase I to the donor chromatin and reduces apoptosis in somatic cell nuclear transfer embryos. 2059 34


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