Fgf signalling through MAPK cascade is required for development of the subpallial telencephalon in zebrafish embryos

Development ◽  
2001 ◽  
Vol 128 (21) ◽  
pp. 4153-4164 ◽  
Author(s):  
Minori Shinya ◽  
Sumito Koshida ◽  
Atsushi Sawada ◽  
Atsushi Kuroiwa ◽  
Hiroyuki Takeda
Keyword(s):  
Anterior Part ◽  
Immunohistochemical Analysis ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Zebrafish Embryos ◽  
Fgf Receptor ◽  
Erk Activation ◽  
Extracellular Signal ◽  
Mitogen Activated Protein ◽  
Fgf Signalling

The telencephalon is formed in the most anterior part of the central nervous system (CNS) and is organised into ventral subpallial and dorsal pallial domains. In mice, it has been demonstrated that Fgf signalling has an important role in induction and patterning of the telencephalon. However, the precise role of Fgf signalling is still unclear, owing to overlapping functions of Fgf family genes. To address this, we have examined, in zebrafish embryos, the activation of Ras/mitogen-activated protein kinase (MAPK), one of the major downstream targets of Fgf signalling. Immunohistochemical analysis reveals that an extracellular signal-regulated kinase (ERK), a vertebrate MAPK is activated in the anterior neural boundary (ANB) of the developing CNS at early segmentation stages. Experiments with Fgf inhibitors reveal that ERK activation at this stage is totally dependent on Fgf signalling. Interestingly, a substantial amount of ERK activation is observed in ace mutants in which fgf8 gene is mutated. We then examine the function of Fgf signalling in telencephalic development by use of several inhibitors to Fgf signalling cascade, including dominant-negative forms of Ras (RasN17) and the Fgf receptor (Fgfr), and a chemical inhibitor of Fgfr, SU5402. In treated embryos, the induction of telencephalic territory normally proceeded but the development of the subpallial telencephalon was suppressed, indicating that Fgf signalling is required for the regionalisation within the telencephalon. Finally, antisense experiments with morpholino-modified oligonucleotides suggest that zebrafish fgf3, which is also expressed in the ANB, co-operates with fgf8 in subpallial development.

scholarly journals Rac-PAK Signaling Stimulates Extracellular Signal-Regulated Kinase (ERK) Activation by Regulating Formation of MEK1-ERK Complexes

2002 ◽  
Vol 22 (17) ◽  
pp. 6023-6033 ◽  
Author(s):  
Scott T. Eblen ◽  
Jill K. Slack ◽  
Michael J. Weber ◽  
Andrew D. Catling
Keyword(s):  
Protein Kinase ◽  
Signaling Pathway ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Wild Type ◽  
Erk Activation ◽  
Extracellular Signal Regulated Kinase ◽  
Signaling Complex ◽  
Extracellular Signal ◽  
Mitogen Activated Protein

ABSTRACT Utilizing mutants of extracellular signal-regulated kinase 2 (ERK2) that are defective for intrinsic mitogen-activated protein kinase or ERK kinase (MEK) binding, we have identified a convergent signaling pathway that facilitates regulated MEK-ERK association and ERK activation. ERK2-Δ19-25 mutants defective in MEK binding could be phosphorylated in response to mitogens; however, signaling from the Raf-MEK pathway alone was insufficient to stimulate their phosphorylation in COS-1 cells. Phosphorylation of ERK2-Δ19-25 but not of wild-type ERK2 in response to Ras V12 was greatly inhibited by dominant-negative Rac. Activated forms of Rac and Cdc42 could enhance the association of wild-type ERK2 with MEK1 but not with MEK2 in serum-starved adherent cells. This effect was p21-activated kinase (PAK) dependent and required the putative PAK phosphorylation sites T292 and S298 of MEK1. In detached cells placed in suspension, ERK2 was complexed with MEK2 but not with MEK1. However, upon replating of cells onto a fibronectin matrix, there was a substantial induction of MEK1-ERK2 association and ERK activation, both of which could be inhibited by dominant-negative PAK1. These data show that Rac facilitates the assembly of a mitogen-activated protein kinase signaling complex required for ERK activation and that this facilitative signaling pathway is active during adhesion to the extracellular matrix. These findings reveal a novel mechanism by which adhesion and growth factor signals are integrated during ERK activation.

Synergistic Activation of Mitogen-Activated Protein Kinase by Cyclic AMP and Myeloid Growth Factors Opposes Cyclic AMP’s Growth-Inhibitory Effects

Blood ◽  
1999 ◽  
Vol 93 (2) ◽  
pp. 537-553 ◽  
Author(s):  
Angel Wai-mun Lee
Keyword(s):  
Protein Kinase ◽  
Cyclic Amp ◽  
Cell Line ◽  
Mapk Pathway ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Erk Activation ◽  
Mitogen Activated Protein ◽  
Erk Activity ◽  
Myeloid Progenitors

Abstract Colony-stimulating factors (CSFs) promote the proliferation, differentiation, commitment, and survival of myeloid progenitors, whereas cyclic AMP (cAMP)-mediated signals frequently induce their growth arrest and apoptosis. The ERK/mitogen-activated protein kinase (MAPK) pathway is a target for both CSFs and cAMP. We investigated how costimulation by cAMP and colony-stimulating factor-1 (CSF-1) or interleukin-3 (IL-3) modulates MAPK in the myeloid progenitor cell line, 32D. cAMP dramatically increased ERK activity in the presence of CSF-1 or IL-3. IL-3 also synergized with cAMP to activate ERK in another myeloid cell line, FDC-P1. The increase in ERK activity was transmitted to a downstream target, p90rsk. cAMP treatment of 32D cells transfected with oncogenic Ras was found to recapitulate the superactivation of ERK seen with cAMP and CSF-1 or IL-3. ERK activation in the presence of cAMP did not appear to involve any of the Raf isoforms and was blocked by expression of dominant-negative MEK1 or treatment with a MEK inhibitor, PD98059. Although cAMP had an overall inhibitory effect on CSF-1–mediated proliferation and survival, the inhibition was markedly increased if ERK activation was blocked by PD98059. These findings suggest that upregulation of the ERK pathway is one mechanism induced by CSF-1 and IL-3 to protect myeloid progenitors from the growth-suppressive and apoptosis-inducing effects of cAMP elevations.

Phorbol ester-induced activation of mitogen-activated protein kinase/extracellular-signal-regulated kinase kinase and extracellular-signal-regulated protein kinase decreases glucose-6-phosphatase gene expression

2001 ◽  
Vol 357 (3) ◽  
pp. 867-873 ◽  
Author(s):  
Dieter SCHMOLL ◽  
Rolf GREMPLER ◽  
Andreas BARTHEL ◽  
Hans-Georg JOOST ◽  
Reinhard WALTHER
Keyword(s):  
Gene Expression ◽  
Protein Kinase ◽  
Phorbol Ester ◽  
Regulation Of Gene Expression ◽  
Insulin Response ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Extracellular Signal Regulated Kinase ◽  
Extracellular Signal ◽  
Mitogen Activated Protein

Glucose-6-phosphatase (G6Pase) plays a central role in blood glucose homoeostasis, and insulin suppresses G6Pase gene expression by the activation of phosphoinositide 3-kinase (PI 3-kinase). Here, we show that the phorbol ester PMA decreases both basal and dexamethasone/cAMP-induced expression of a luciferase gene under the control of the G6Pase promoter in transiently transfected H4IIE hepatoma cells. This regulation was suppressed by the inhibitors of the mitogen-activated protein kinase/extracellular-signal-regulated kinase kinase (MEK), PD98059 and U0126, but not by the inhibitor of PI 3-kinase, LY294002. The co-expression of a constitutively active mutant of MEK mimicked the regulation of G6Pase promoter activity by PMA. The effect of PMA on both basal and induced G6Pase gene transcription was impaired by the overexpression of a dominant negative MEK construct, as well as by the expression of mitogen-activated protein kinase phosphatase-1. The mutation of the forkhead-binding sites within the insulin-response unit of the G6Pase promoter, which decreases the effect of insulin on G6Pase gene expression, did not alter the regulation of gene expression by PMA. The data show that PMA decreases G6Pase gene expression by the activation of MEK and extracellular-signal regulated protein kinase. With that, PMA mimics the effect of insulin on G6Pase gene expression by a different signalling pathway.

scholarly journals Differential inhibition of signaling pathways by dominant-negative SH2/SH3 adapter proteins.

1995 ◽  
Vol 15 (12) ◽  
pp. 6829-6837 ◽  
Author(s):  
M Tanaka ◽  
R Gupta ◽  
B J Mayer
Keyword(s):  
Protein Kinase ◽  
Signaling Pathways ◽  
Tyrosine Kinases ◽  
Egf Receptor ◽  
Sh3 Domain ◽  
Sh2 Domain ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Erk Activation ◽  
Mitogen Activated Protein

SH2/SH3 adapters are thought to function in signal transduction pathways by coupling inputs from tyrosine kinases to downstream effectors such as Ras. Members of the mitogen-activated protein kinase family are known to be activated by a variety of mitogenic stimuli, including tyrosine kinases such as Abl and the epidermal growth factor (EGF) receptor. We have used activation of the mitogen-activated protein kinase Erk-1 as a model system with which to examine whether various dominant-negative SH2/SH3 adapters (Grb2, Crk, and Nck) could block signaling pathways leading to Erk activation. Activation of Erk-1 by oncogenic Abl was effectively inhibited by Grb2 with mutations in either its SH2 or SH3 domain or by Crk-1 with an SH3 domain mutation. The Crk-1 SH2 mutant was less effective, while Nck SH2 and SH3 mutants had little or no effect on Erk activation. These results suggest that both Crk and Grb2 may contribute to the activation of Erk by oncogenic Abl, whereas Nck is unlikely to participate in this pathway. Next we examined whether combinations of these dominant-negative adapters could inhibit Erk activation more effectively than each mutant alone. When combinations of Crk-1 and Grb2 mutants were analyzed, the combination of the Crk-1 SH3 mutant plus the Grb2 SH3 mutant gave a striking synergistic effect. This finding suggests that in Abl-transformed cells, more than one class of tyrosine-phosphorylated sites (those that bind the Grb2 SH2 domain and those that bind the Crk SH2 domain) can lead to Ras activation. In contrast to results with Abl, Erk activation by EGF was strongly inhibited only by Grb2 mutants; Crk and Nck mutants had little or no effect. This finding suggests that Grb2 is the only adapter involved in the activation of Erk by EGF. Dominant-negative adaptors provide a novel means to identify binding interactions important in vivo for signaling in response to a variety of stimuli.

5-HT2A receptors stimulate mitogen-activated protein kinase via H2O2 generation in rat renal mesangial cells

2000 ◽  
Vol 278 (4) ◽  
pp. F650-F658 ◽  
Author(s):  
Eddie L. Greene ◽  
Odette Houghton ◽  
Georgiann Collinsworth ◽  
Maria N. Garnovskaya ◽  
Toshio Nagai ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Phospholipase C ◽  
Mesangial Cells ◽  
Signal Transduction Pathway ◽  
Peak Effect ◽  
Mitogen Activated Protein Kinase ◽  
Similar Degree ◽  
Erk Activation ◽  
Extracellular Signal ◽  
Mitogen Activated Protein

Serotonin (5-HT) stimulates mitogenesis in rat renal mesangial cells through a G protein-coupled 5-HT2A receptor. We tested the hypothesis that oxidants might be involved in the signal transduction pathway linking the receptor to extracellular signal-regulated protein kinase (ERK). 5-HT rapidly increased the activity and phosphorylation of ERK. These effects were blocked by the 5-HT2A receptor antagonist ketanserin. The peak effect was noted at 5–10 min, and half-maximal stimulation was achieved at 10–30 nM 5-HT. Chemical inhibitor and activator studies supported the involvement of phospholipase C, protein kinase C (PKC), and reactive oxygen species (ROS, i.e., H2O2 and superoxide) generated by an NAD(P)H oxidase-like enzyme in the ERK activation cascade. Mapping studies supported a location for the NAD(P)H oxidase enzyme and the ROS downstream from PKC. Our studies are most consistent with an ERK activation pathway as follows: 5-HT2A receptor → Gq protein → phospholipase C → diacylglycerol → classical PKC → NAD(P)H oxidase → superoxide → superoxide dismutase → H2O2 → mitogen-activated extracellular signal-regulated kinase → ERK. These studies demonstrate a role for the 5-HT2A receptor in rapid, potent, and efficacious activation of ERK in rat renal mesangial cells. They support a role for oxidants in conveying the stimulatory signal from 5-HT, because 1) chemical antioxidants attenuate the 5-HT signal, 2) oxidants and 5-HT selectively activate ERK to a similar degree, 3) 5-HT produces superoxide and H2O2 in these cells, and 4) a specific enzyme [NAD(P)H oxidase] has been implicated as the source of the ROS, which react selectively downstream of classical PKC.

MAPK-induced Ser727 phosphorylation promotes SUMOylation of STAT1

2007 ◽  
Vol 409 (1) ◽  
pp. 179-185 ◽  
Author(s):  
Sari Vanhatupa ◽  
Daniela Ungureanu ◽  
Maija Paakkunainen ◽  
Olli Silvennoinen
Keyword(s):  
Interferon Γ ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Extracellular Signal Regulated Kinase ◽  
Extracellular Signal ◽  
Microbial Infections ◽  
Mitogen Activated Protein ◽  
P38mapk Pathway ◽  
Ifn Γ

STAT1 (signal transducer and activator of transcription 1) is a critical mediator of IFN-γ (interferon-γ)-induced gene responses, and its function is regulated through phosphorylation of Tyr701 and Ser727. MAPK (mitogen-activated protein kinase) pathways mediate phosphorylation of Ser727 in response to microbial infections, stress stimuli and growth factors. Recently, STAT1 was found to become modified by PIAS (protein inhibitor of activated STAT)-mediated SUMO-1 (small ubiquitin-related modifier-1) conjugation at Lys703, but the regulation of this modification is largely unknown. Here, we have investigated the role of MAPK-induced Ser727 phosphorylation in regulation of STAT1 SUMOylation. Activation of the p38MAPK pathway by upstream activating kinase, MKK6 (MAPK kinase-6) or osmotic stress enhanced the SUMOylation of STAT1, which was counteracted by the p38MAPK inhibitor SB202190 or by dominant-negative p38MAPK. Activation of the ERK1/2 (extracellular-signal-regulated kinase 1/2) pathway by Raf-1 also enhanced Ser727 phosphorylation and SUMOylation of STAT1, and this induction was counteracted by PD98059 inhibitor. Mutation of Ser727 to alanine abolished the p38MAPK-induced SUMOylation. Furthermore, S727D and S727E mutations, which mimic the phosphorylation of Ser727, enhanced the basal SUMOylation of STAT1 and interaction between PIAS1 and STAT1. Taken together, these results identify Ser727 phosphorylation as a regulator of STAT1 SUMOylation and highlight the central role of Ser727 in co-ordination of STAT1 functions in cellular responses.

Involvement of p38 mitogen–activated protein kinase in different stages of thymocyte development

Blood ◽  
2003 ◽  
Vol 101 (3) ◽  
pp. 970-976 ◽  
Author(s):  
Shu-Ching Hsu ◽  
Chia-Cheng Wu ◽  
Jiahuai Han ◽  
Ming-Zong Lai
Keyword(s):  
T Cell ◽  
Positive Selection ◽  
P38 Mapk ◽  
Cell Receptor ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Thymocyte Development ◽  
Dominant Negative Mutation ◽  
Extracellular Signal ◽  
Mitogen Activated Protein

Abstract Positive selection of thymocytes during T-cell development is mediated by T-cell receptor (TCR)–activated signals. For different mitogen-activated protein kinases (MAPKs) activated by TCR complex, a selective involvement of extracellular signal–regulated kinase, but not p38 MAPK, in positive selection has been suggested. Using transgenic mice with dominant-negative mutation of both MAP kinase kinase 3 (MMK3) and MKK6, we obtained mice with different extents of inhibition of p38 MAPK activation. Partial inhibition of p38 MAPK impaired CD4−CD8− thymocyte development and T-cell proliferation, but not positive selection. Interference with thymocyte positive selection was observed in mice with effective suppression of p38 MAPK. Our results suggest that, in addition to early thymocyte development, p38 is involved in positive selection.

PKC-ε regulation of extracellular signal-regulated kinase: a potential role in phenylephrine-induced cardiocyte growth

2004 ◽  
Vol 286 (6) ◽  
pp. H2195-H2203 ◽  
Author(s):  
Vijay U. Rao ◽  
Hirokazu Shiraishi ◽  
Paul J. McDermott
Keyword(s):  
Growth Regulation ◽  
Dominant Negative ◽  
Mitogen Activated Protein Kinase ◽  
Neonatal Rat ◽  
Inhibitory Effects ◽  
Hypertrophic Growth ◽  
Extracellular Signal Regulated Kinase ◽  
Extracellular Signal ◽  
Mitogen Activated Protein ◽  
Acute Increase

Hypertrophic growth of cardiac muscle is dependent on activation of the PKC-ε isoform. To define the effectors of PKC-ε involved in growth regulation, recombinant adenoviruses were used to overexpress either wild-type PKC-ε (PKC-ε/WT) or dominant negative PKC-ε (PKC-ε/DN) in neonatal rat cardiocytes. PKC-ε/DN inhibited acute activation of PKC-ε produced in response to phorbol ester and reduced ERK1/2 activity as measured by the phosphorylation of p42 and p44 isoforms. The inhibitory effects were specific to PKC-ε because PKC-ε/DN did not prevent translocation of either PKC-α or PKC-δ. Overexpression of PKC-ε/DN blunted the acute increase in ERK1/2 phorphorylation induced by the α1-adrenergic agonist phenylephrine (PE ). Inhibition of PKC-δ with rottlerin potentiated the effects of PE on ERK1/2 phosphorylation. PKC-ε/DN adenovirus also blocked cardiocyte growth as measured after 48 h of PE treatment, although the multiplicity of infection was lower than that required to block acute ERK1/2 activation. PE activated p38 mitogen-activated protein kinase as measured by its phosphorylation, but the response was not blocked by PKC inhibitors or by overexpression of PKC-ε/DN. Taken together, these studies show that the hypertrophic agonist PE regulates ERK1/2 activity in cardiocytes by a pathway dependent on PKC-ε and that PE-induced growth is mediated by PKC-ε.

The quantity and duration of FcRγ signals determine mast cell degranulation and survival

Blood ◽  
2004 ◽  
Vol 103 (8) ◽  
pp. 3093-3101 ◽  
Author(s):  
Sho Yamasaki ◽  
Eri Ishikawa ◽  
Masayuki Kohno ◽  
Takashi Saito
Keyword(s):  
Mast Cell ◽  
Cell Survival ◽  
Immunoglobulin E ◽  
Mast Cell Degranulation ◽  
Mitogen Activated Protein Kinase ◽  
Cross Linking ◽  
Erk Activation ◽  
Extracellular Signal Regulated Kinase ◽  
Extracellular Signal ◽  
Mitogen Activated Protein

Abstract Immunoglobulin E (IgE) bound to multivalent antigen (Ag) elicits mast cell degranulation but not survival; on the contrary, IgE in the absence of Ag (IgE(-Ag)) induces survival only but not degranulation. Although these distinct responses are mediated through the same receptor, FcϵRI, the molecular mechanism generating the divergence is largely unknown. We recently showed that the signals through FcRγ chain are essential for IgE(-Ag)–induced mast cell survival as well as IgE(+Ag)–induced degranulation. To determine whether the cellular output is regulated by the quantity of FcRγ signal, we expressed CD8/FcRγ chimeras (CD8/γ) in bone marrow–derived mast cells (BMMCs) from FcRγ-/- mice to manipulate the strength of FcRγ signals by anti-CD8 cross-linking. Cross-linking of CD8/γ induced mast cell survival and degranulation. Survival was induced by weaker stimulation than needed for degranulation in terms of anti-CD8 concentration and the valency of chimera. However, sustained extracellular signal-regulated kinase (Erk) activation seems to regulate survival even when the activation signal was strong enough to elicit degranulation. Generation of sustained Erk activation by active mitogen-activated protein kinase kinase (MEK) induced BMMC survival. These results suggest that the duration and the magnitude of FcRγ signals may determine mast cell survival and degranulation, respectively. (Blood. 2004;103:3093-3101)

Regulation of polymorphonuclear leukocyte phagocytosis by myosin light chain kinase after activation of mitogen-activated protein kinase

Blood ◽  
2000 ◽  
Vol 95 (7) ◽  
pp. 2407-2412 ◽  
Author(s):  
Pamela J. Mansfield ◽  
James A. Shayman ◽  
Laurence A. Boxer
Keyword(s):  
Map Kinase ◽  
Light Chain ◽  
Polymorphonuclear Leukocyte ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Mek Inhibitor ◽  
Mitogen Activated Protein Kinase ◽  
Erk Activation ◽  
Extracellular Signal ◽  
Mitogen Activated Protein

Abstract Polymorphonuclear leukocyte (PMNL) phagocytosis mediated by FcγRII proceeds in concert with activation of the mitogen-activated protein (MAP) kinase, extracellular signal-regulated kinase ERK2. We hypothesized that myosin light chain kinase (MLCK) could be phosphorylated and activated by ERK, thereby linking the MAP kinase pathway to the activation of cytoskeletal components required for pseudopod formation. To explore this potential linkage, PMNLs were challenged with antibody-coated erythrocytes (EIgG). Peak MLCK activity, 3-fold increased over controls, occurred at 4 to 6 minutes, corresponding with the peak rate of target ingestion and ERK2 activity. The MLCK inhibitor ML-7 (10 μmol/L) inhibited both phagocytosis and MLCK activity to basal values, thereby providing further support for the linkage between the functional response and the requirement for MLCK activation. The MAPK kinase (MEK) inhibitor PD098059 inhibited phagocytosis, MLCK activity, and ERK2 activity by 80% to 90%. To directly link ERK activation to MLCK activation, ERK2 was immunoprecipitated from PMNLs after EIgG ingestion. The isolated ERK2 was incubated with PMNL cytosol as a source of unactivated MLCK and with MLCK substrate; under these conditions ERK2 activated MLCK, resulting in phosphorylation of the MLCK substrate or of the myosin light chain itself. Because MLCK activates myosin, we evaluated the effect of directly inhibiting myosin adenosine triphosphatase using 2,3-butanedione monoxime (BDM) and found that phagocytosis was inhibited by more than 90% but MLCK activity remained unaffected. These results are consistent with the interpretation that MEK activates ERK, ERK2 then activates MLCK, and MLCK activates myosin. MLCK activation is a critical step in the cytoskeletal changes resulting in pseudopod formation.

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