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Results for “Signaling pathways”

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PubMed2003

Growth signalling pathways in Arabidopsis and the AGC protein kinases.

Lipid-derived signals are central to regulating a multitude of cellular processes but, in plants, little is known of the downstream signalling pathways. The Arabidopsis 3-phosphoinositide-dependent protein kinase (PDK1) could couple lipid signals to the activation of several protein kinases of the so-called AGC kinase family. The Arabidopsis AGC kinases contain sequence motives required for the docking of PDK1 and phosphorylation of their activation loop in the kinase catalytic domain. It is becoming evident that specific members of the AGC kinases are implicated in key growth signalling pathways. For example, Arabidopsis p70(S6K) might be a nodal point able to integrate hormonal and developmental signals with nutritional inputs, together with the Arabidopsis Target of Rapamycin (TOR) protein.

3-Phosphoinositide-Dependent Protein Kinases↗

PubMed2004

Casein kinase Iepsilon plays a functional role in the transforming growth factor-beta signaling pathway.

The transforming growth factor-beta (TGF-beta) signaling pathway is known to be involved in a wide range of biological events, including development, cellular differentiation, apoptosis, and oncogenesis. The TGF-beta signal is mediated by ligand binding to the type II receptor, leading to the recruitment and activation of the type I receptor, and subsequent activation of a family of intracellular signal transducing proteins called Smads. Here we report a regulatory role for casein kinase Iepsilon (CKIepsilon) in the TGF-beta signaling cascade. We find that CKIepsilon binds to all Smads and the cytoplasmic domains of the type I and type II receptors both in vitro and in vivo. The interaction of CKIepsilon with the type I and type II receptors is independent of TGF-beta stimulation, whereas the CKIepsilon/Smad interaction is transiently disrupted by ligand treatment. Additionally, CKIepsilon is able to phosphorylate the receptor-activated Smads (Smads 1-3 and 5) and the type II receptor in vitro. Transcriptional reporter assays reveal that transient overexpression of wild type CKIepsilon dramatically reduces basal reporter activity but enhances TGF-beta-stimulated transcription. Furthermore, overexpression of a kinase-dead mutant of CKIepsilon inhibits both basal and ligand-induced transcription, whereas inhibition of endogenous CKI catalytic activity with IC261 blocks only TGF-beta-stimulated reporter activity. Finally, knocking down CKIepsilon protein levels results in a significant increase in basal and TGF-beta-induced transcription. These results suggest that CKIepsilon plays a ligand-dependent, differential, and dual regulatory role within the TGF-beta signaling pathway.

Animals↗

PubMed2001

Signaling pathways activated by alpha1-adrenergic receptor subtypes in PC12 cells.

PC12 cells were used to compare signaling pathways activated by alpha1-adrenergic receptor (AR) subtypes. PC12 cells were transfected with human alpha1A, alpha1B, or alpha1D-ARs, and subclones stably expressing receptor densities in physiological ranges isolated and characterized. Norepinephrine (NE) activated a large number of signaling pathways in transfected cells, including inositol phosphate formation, intracellular calcium, all three arms of the mitogen activated protein kinase pathways, and a number of tyrosine kinases. Activation of mitogen activated protein kinase pathways and tyrosine kinases was not blocked by chelation of intracellular calcium with BAPTA or inhibition of protein kinase C. NE also activated luciferase reporter constructs for seven different transcription factors (AP1, SRE, CRE, NFkappaB, NFAT, Stat, GAS) following transfection into alpha1A-AR expressing PC12 cells. However, similar increases in inositol phosphate formation and intracellular Ca2+ caused by purinergic P2Y2 receptor activation did not activate any of these reporters. Comparison of alpha1-AR subtypes showed that the alpha1A activated all seven reporters, the alpha1B showed smaller effects, while the alpha1D was ineffective. NE caused differentiation of alpha1A, but not alpha1B or alpha1D, -AR expressing PC12 cells similar to that caused by NGF. This NE-induced differentiation was reduced or blocked by all inhibitors tested. We conclude that alpha1-ARs activate many signaling pathways and transcriptional responses in PC12 cells, which are not linearly related to second messenger production, and which may differ for different alpha1-AR subtypes.

Animals↗

PubMed2004

Rapid signalling pathway activation by androgens in epithelial and stromal cells.

Estradiol rapidly activates Src as well as the Src-dependent pathway in human mammary cancer-derived MCF-7 cells, in human prostate cancer-derived LNCaP cells and in Cos cells transiently expressing hERs [EMBO J. 15 (1996) 1292; EMBO J. 17 (1998) 2008]. In addition, estradiol immediately stimulates, yes, an ubiquitous member of the Src kinase family, in human colon carcinoma-derived Caco-2 cells [Cancer Res. 56 (1996) 4516]. Progestins and androgens activate the same pathway in human mammary and prostate cancer-derived cells [EMBO J. 17 (1998) 2008; EMBO J. 19 (2000) 5406]. We observed that estradiol also stimulates the phosphatidylinositol-3-kinase (PI3K)/AKT pathway in MCF-7 cells [EMBO J. 20 (2001) 6050]. In these cells, activation of the Src- and the PI3 K-dependent pathways is simultaneous and mediated by direct interactions of the two kinases with ERalpha. The signalling pathway activation by sex-steroid hormones leads to DNA synthesis and cell growth in human mammary and prostate cancer-derived cells [EMBO J. 19 (2000) 5406; EMBO J. 20 (2001) 6050; EMBO J. 18 (1999) 2500]. Furthermore, androgen stimulation of NIH3T3 fibroblasts activates the same pathways triggered by this hormone in LNCaP cells and promotes the S-phase entry or cytoskeleton changes in these cells [J. Cell Biol. 161 (2003) 547]. All the described effects are rapid and require classic steroid receptors, but, surprisingly, not their transcriptional activity. Indeed, a transcriptionally inactive mutant of hER mediates the estrogen-stimulated DNA synthesis of NIH3T3 fibroblasts [EMBO J. 18 (1999) 2500]. Furthermore, AR in NIH3T3 cells does not enter nuclei and is unable to respond to the hormone with transcription stimulation, whereas it activates signaling pathways and triggers important biological responses. Signaling pathway activation by steroids has also been described by other groups under different experimental conditions and/or in different cell types. In these cells, steroid stimulation triggers various effects, such as neuroprotection, vasorelaxation or bone protection [J. Neurosci. Res. 60 (2000) 321; Nature 407 (2000) 538; J. Cell Biochem. 76 (1999) 206]. Analysis of the mechanisms responsible for the hormone-dependent and steroid receptor-mediated pathway activation in epithelial as well as stromal cells reveals immediate association of steroid receptors with extranuclear signaling effectors [EMBO J. 17 (1998) 2008; Cancer Res. 56 (1996) 4516; EMBO J. 19 (2000) 5406; EMBO J. 20 (2001) 6050; J. Cell Biol. 161 (2003) 547]. These results further highlight the central role of the hormone-regulated protein-protein interactions in the steroid action. They also offer the possibility of interfering with important activities of hormones, such as proliferation or survival, cytoskeleton changes as well as invasiveness and vasorelaxation, without affecting the steroid effects that depend on receptor transcriptional activity.

Androgens↗

PubMed2003

Calcium signaling pathway involving calcineurin regulates interleukin-8 gene expression through activation of NF-kappaB in human osteoblast-like cells.

UNLABELLED: Involvement of aberrant IL-8 production by osteoblasts was demonstrated in pathogenesis of inflammatory joint diseases. We thus investigated intracellular signaling pathways leading to IL-8 expression in human osteoblast-like HOS-TE85 cells. It was demonstrated that Ca2+ signaling pathway involving calcineurin regulates IL-8 gene expression through activation of a transcription factor, NF-kappaB. INTRODUCTION: Involvement of aberrant interleukin (IL)-8 production by osteoblasts was demonstrated in pathogenesis of inflammatory joint diseases. However, intracellular signaling pathways leading to IL-8 expression in osteoblasts have been poorly explored. Because a variety of external stimuli was shown to increase intracellular Ca2+ in osteoblasts, we investigated effects of Ca(2+)-ionophore and phorbol-myristate-acetate (Ion/PMA) on IL-8 expression in human osteoblast-like HOS-TE85 cells and compared the effects with those elicited by TNF-alpha. MATERIALS AND METHODS: HOS-TE85 cells were treated with Ion/PMA or TNF-alpha in the presence and absence of calcineurin inhibitors (CnI), cyclosporin A, and FK506. IL-8 mRNA levels and its promoter activities were examined by Northern blot and luciferase reporter analyses, respectively. Electrophoretic mobility shift assay (EMSA) was used to evaluate DNA binding activities of transcription factors such as NF-kappaB. Degradation of IkappaB, a cytoplasmic NF-kappaB-inhibitory protein, was examined by Western blot analysis. RESULTS: Ion/PMA and TNF-alpha induced IL-8 mRNA expression. Interestingly, CnI attenuated the induction by Ion/PMA, but not that by TNF-alpha. Promoter activity was also increased by both stimuli, and only the Ion/PMA-dependent increase was suppressed by CnI. Introduction of mutations in the promoter demonstrated that one NF-kappaB site was responsible for the suppression by CnI. EMSA revealed that this site binds with NF-kappaB containing p65 that was activated by Ion/PMA and TNF-alpha and that CnI inhibited only Ion/PMA-dependent NF-kappaB activation. Accordingly, CnI blocked only Ion/PMA-dependent degradation of IkappaB-alpha. In addition, the basal and Ion/PMA-dependent IL-8 promoter activities were enhanced by co-transfection of constitutively active calcineurin. CONCLUSION: These results show that the Ca2+ signaling pathway involving calcineurin regulates IL-8 gene expression through activation of NF-kappaB in human osteoblast-like cells.

Antineoplastic Agents↗

PubMed1997

Involvement of hippocampal cAMP/cAMP-dependent protein kinase signaling pathways in a late memory consolidation phase of aversively motivated learning in rats.

cAMP/cAMP-dependent protein kinase (PKA) signaling pathway has been recently proposed to participate in both the late phase of long term potentiation in the hippocampus and in the late, protein synthesis-dependent phase of memory formation. Here we report that a late memory consolidation phase of an inhibitory avoidance learning is regulated by an hippocampal cAMP signaling pathway that is activated, at least in part, by D1/D5 receptors. Bilateral infusion of SKF 38393 (7.5 microg/side), a D1/D5 receptor agonist, into the CA1 region of the dorsal hippocampus, enhanced retention of a step-down inhibitory avoidance when given 3 or 6 h, but not immediately (0 h) or 9 h, after training. In contrast, full retrograde amnesia was obtained when SCH 23390 (0.5 microg/side), a D1/D5 receptor antagonist, was infused into the hippocampus 3 or 6 h after training. Intrahippocampal infusion of 8Br-cAMP (1.25 microg/side), or forskolin (0.5 microg/side), an activator of adenylyl cyclase, enhanced memory when given 3 or 6 h after training. KT5720 (0.5 microg/side), a specific inhibitor of PKA, hindered memory consolidation when given immediately or 3 or 6 h posttraining. Rats submitted to the avoidance task showed learning-specific increases in hippocampal 3H-SCH 23390 binding and in the endogenous levels of cAMP 3 and 6 h after training. In addition, PKA activity and P-CREB (phosphorylated form of cAMP responsive element binding protein) immunoreactivity increased in the hippocampus immediately and 3 and 6 h after training. Together, these findings suggest that the late phase of memory consolidation of an inhibitory avoidance is modulated cAMP/PKA signaling pathways in the hippocampus.

Animals↗

PubMed2003

Estrogen and growth factor signaling pathway: basic approaches for clinical application.

Estrogen and its receptor play important roles in genesis and malignant progression of estrogen-dependent cancers, together with various growth factors. Functional cross-talk between estrogen-signaling and growth factor-mediated signaling pathways has been reported. Firstly, we show an example of the cross-talk that may alter the effect of antagonist on the breast and endometrial cancer cell growth. Our observations suggest that the constitutively activated MAP kinase-signaling pathway in endometrial cancer cells might enhance the transcriptional activity of ERalpha via phosphorylation of AF-1 domain. This mechanism may cause the growth stimulative effect of tamoxifen on the endometrium. Secondly, we show our recent study for comprehensive understanding of estrogen-signaling pathway using cDNA microarray. According to the results of the expression profiling of estrogen-responsive genes in ER-positive breast cancer cells using large-scale cDNA microarray, the custom-made cDNA microarray, on which only estrogen-responsive genes were loaded, was produced. Using this microarray consisting of the narrowed gene subset, we analyzed estrogen responsiveness of various cell lines and effect of estrogen antagonists. Aim of this study is not only to address the molecular mechanisms of estrogen-dependent growth of breast cancer, but also to develop the new diagnostic tools for responsiveness to hormone therapy of primary breast cancer patients. Finally, in order to understand the local tumor biology including stroma-cancer interaction, we recently developed the new analytical system using ERE-GFP introduced into breast cancer cells. Several observations indicated that these reporter cells were useful for assessment of stimulative effects of stroma cells adjacent to breast cancer on the estrogen-signaling pathway. These studies may provide not only new clues for elucidation of the molecular mechanisms of estrogen-dependent growth of breast cancer, but also assessment of anti-estrogen responses of individual breast cancer for patient-tailored hormone therapy.

Breast Neoplasms↗

PubMed2001

Synergistic activation of the Wnt signaling pathway by Dvl and casein kinase Iepsilon.

Although casein kinase Iepsilon (CKIepsilon) has been shown to regulate the Wnt signaling pathway positively, its mode of action is not clear. In this study we show that CKIepsilon activates the Wnt signaling pathway in co-operation with Dvl. CKIepsilon and Axin associated with different sites of Dvl, and CKIepsilon and Dvl interacted with distinct regions on Axin. Therefore, these three proteins formed a ternary complex. Either low expression of Dvl or CKIepsilon alone did not accumulate beta-catenin, but their co-expression accumulated greatly. Dvl and CKIepsilon activated the transcriptional activity of T cell factor (Tcf) synergistically. Although the Dvl mutant that binds to Axin but not to CKIepsilon activated Tcf, it did not synergize with CKIepsilon. Another Dvl mutant that does not bind to Axin did not activate Tcf irrespective of the presence of CKIepsilon. Furthermore, Dvl and CKIepsilon co-operatively induced axis duplication of Xenopus embryos. These results indicate that Dvl and CKIepsilon synergistically activated the Wnt signaling pathway and that the binding of the complex of Dvl and CKIepsilon to Axin is necessary for their synergistic action.

Adaptor Proteins, Signal Transducing↗

PubMed2005

The regulation of the expression of inducible nitric oxide synthase by Src-family tyrosine kinases mediated through MyD88-independent signaling pathways of Toll-like receptor 4.

Bacterial lipopolysaccharide (LPS) activates Toll-like receptor 4 (TLR4) leading to the expression of inflammatory gene products. Src-family tyrosine kinases (STKs) are known to be activated by LPS in monocytes/macrophages. Therefore, we determined the role of STKs in TLR4 signaling pathways and target gene expression in macrophages. The activation of NFkappaB, and p38 MAPK, and the expression of inducible nitric oxide synthase (iNOS) induced by LPS were not affected in macrophages deficient in three STKs (Lyn, Hck, and Fgr). These results suggest that the deletion of the three STKs among possibly nine STKs is not sufficient to abolish total activity of STKs possibly due to the functional redundancy of other STKs present in macrophages. However, two structurally unrelated pan-inhibitors of STKs, PP1 and SU6656, suppressed LPS-induced iNOS expression in MyD88-knockout as well as wild-type macrophages. The suppression of iNOS expression by the inhibitors was correlated with the downregulation of IFNbeta (a MyD88-independent gene) expression and subsequent decrease in STAT1 phosphorylation. Moreover, PP1 suppressed the expression of IFNbeta and iNOS induced by TRIF, a MyD88-independent adaptor of TLR4. PP1 suppressed STAT1 phosphorylation induced by LPS, but not by IFNbeta suggesting that STKs are involved in the primary downstream signaling pathways of TLR4, but not the secondary signaling pathways downstream of IFNbeta receptor. Together, these results demonstrate that STKs play a positive regulatory role in TLR4-mediated iNOS expression in a MyD88-independent (TRIF-dependent) manner. These results provide new insight in understanding the role of STKs in TLR4 signaling pathways and inflammatory target gene expression.

Adaptor Proteins, Signal Transducing↗

PubMed2003

Multiple signalling pathways mediate fungal elicitor-induced beta-thujaplicin biosynthesis in Cupressus lusitanica cell cultures.

The biosynthesis of a phytoalexin, beta-thujaplicin, in Cupressus lusitanica cell cultures can be stimulated by a yeast elicitor, H(2)O(2), or methyl jasmonate. Lipoxygenase activity was also stimulated by these treatments, suggesting that the oxidative burst and jasmonate pathway may mediate the elicitor-induced accumulation of beta-thujaplicin. The elicitor signalling pathway involved in beta-thujaplicin induction was further investigated using pharmacological and biochemical approaches. Treatment of the cells with calcium ionophore A23187 alone stimulated the production of beta-thujaplicin. A23187 also enhanced the elicitor-induced production of beta-thujaplicin. EGTA, LaCl(3), and verapamil pretreatments partially blocked A23187- or yeast elicitor-induced accumulation of beta-thujaplicin. These results suggest that Ca(2+) influx is required for elicitor-induced production of beta-thujaplicin. Treatment of cell cultures with mastoparan, melittin or cholera toxin alone or in combination with the elicitor stimulated the production of beta-thujaplicin or enhanced the elicitor-induced production of beta-thujaplicin. The G-protein inhibitor suramin inhibited the elicitor-induced production of beta-thujaplicin, suggesting that receptor-coupled G-proteins are likely to be involved in the elicitor-induced biosynthesis of beta-thujaplicin. Indeed, both GTP-binding activity and GTPase activity of the plasma membrane were stimulated by elicitor, and suramin and cholera toxin affected G-protein activities. In addition, all inhibitors of G-proteins and Ca(2+) flux suppressed elicitor-induced increases in lipoxygenase activity whereas activators of G-proteins and the Ca(2+) signalling pathway increased lipoxygenase activity. These observations suggest that Ca(2+) and G-proteins may mediate elicitor signals to the jasmonate pathway, and the jasmonate signalling pathway may then lead to the production of beta-thujaplicin.

Acetates↗

PubMed1991

Growth factor and oncogene signalling pathways as targets for rational anticancer drug development.

There is a critical need for new targets, in addition to DNA, for anticancer drug development. A recently discovered target is the intracellular signalling pathways that mediate the actions of growth factors and oncogenes on cell proliferation. Two important pathways, the myo-inositol and protein tyrosine kinase signalling pathways are reviewed. Three classes of compounds that modulate myo-inositol signalling are discussed. These are: 1) the D-3-substituted-3-deoxy-myo-inositol analogues that act as antimetabolites of myo-inositol and show selective growth inhibition of some transformed cells; 2) the alkaloid staurosporine that acts as a potent inhibitor of protein kinase C and of platelet-derived growth factor (PDGF) receptor protein tyrosine kinase activity; 3) the ether lipid analogues that block growth factor signalling at several points by acting as inhibitors of protein kinase C, phosphoinositide specific phospholipase C and inositol(1,4,5)trisphosphate-induced Ca2+ release. It is suggested that inhibition of signalling pathways may explain the growth inhibitory effects of these compounds. Other potential signalling target sites for anticancer drug development are discussed.

Alkaloids↗

PubMed2005

[Effects of angiotensin II on extracellular signal-regulated protein kinases signaling pathway in cultured vascular smooth muscle cells from Wistar-Kyoto rats and spontaneously hypertensive rats].

The aim of this study was to investigate the effects of angiotensin II (Ang II) on extracellular signal-regulated protein kinase (ERK) signaling pathway in cultured vascular smooth muscle cells (VSMCs) from spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats. VSMCs from SHR and WKY rats were treated with 1x10(-7) mmol/L Ang II for 24 h in the absence or presence of 30 min of pre-treatment of valsartan (1x10(-5) mmol/L) or PD98059 (1x10(-5)mmol/L), selective inhibitor of ERKs- dependent pathways, when they were cultured in 20% calf serum medium. VSMCs of SHR and WKY cultured in serum-free medium were used as control groups. Among the different treatments, VSMCs from the SHR and WKY were devided into four groups: (1) control, (2) Ang II, (3) Ang II + valsartan, (4) Ang II + PD98059. ERK activity in VSMCs was measured by immuno-precipitation. Proteins of total ERK (t-ERK), phosphorylated-ERK (p-ERK) and mitogen-activated protein kinases phosphatase-1 (MKP-1) in VSMCs were detected by Western blot. MKP-1 mRNA in VSMCs was measured by RT-PCR. In VSMCs from WKY or SHR rats, ERK activity, p-ERK, MKP-1 and MKP-1 mRNA in Ang II group were higher than those in control group (P<0.05). In both SHRs and WKYs, there were no significant differences in ERK activity, p-ERK, MKP-1 and MKP-1 mRNA among the control group, Ang II + valsartan group and Ang II + PD98059 group. ERK activity, p-ERK, MKP-1 and MKP-1 mRNA in SHRs were significantly higher than those in WKYs with same treatments (P<0.01). There was no significant difference in t-ERK among different groups and no difference in t-ERK between SHRs and WKYs (P>0.05). Our results show that Ang II activates VSMCs ERK signaling pathways via Ang II type 1 (AT(1)) receptors. Ang II increased ERK activity and p-ERK, but not t-ERK, accompanied by an increase in MKP-1 mRNA expression and protein. Among the different treatments, ERK activity and p-ERK were higher in SHR than in WKY. Valsartan and PD98059 blocked Ang II-stimulated ERK activation. These results suggest that ERK signaling pathway plays an important role in the pathogenesis of hypertension. The effect of Ang II on SHR and WKY VSMCs' ERK signaling pathway may be mediated by AT(1) receptors, enhancing ERK activity and the amount of p-ERK, and then increasing MKP-1 mRNA and its expression.

Angiotensin II↗

PubMed2001

CD40-mediated signaling in monocytic cells: up-regulation of tumor necrosis factor receptor-associated factor mRNAs and activation of mitogen-activated protein kinase signaling pathways.

The biochemical pathways involved in CD40 signaling have been extensively studied in B cells and B cell lines, and appear to be primarily initiated by recruitment of the tumor necrosis factor (TNF) receptor-associated factor (TRAF) signaling proteins to the CD40 cytoplasmic domain. Signaling pathways activated through CD40 in monocytes/macrophages have not been characterized as well as in B cells. Using human monocytes and the human monocytic cell line THP1, we examined signal transduction events induced by CD40 engagement with its ligand, CD154. In human monocytes, all TRAF mRNAs were expressed constitutively and CD40 ligation resulted in a strong up-regulation of TRAF1 mRNA. In THP1 cells, CD40 ligation induced expression of TRAF1 and TRAF5 mRNAs. Engagement of CD40 in both monocytes and THP1 cells led to the rapid and transient activation of the extracellular signal-regulated kinases (ERK) 1 and 2, and to low levels of JNK activation. No CD40-dependent activation of p38 mitogen-activated protein kinase (MAPK) was found. In CD154-stimulated monocytes and THP1 cells the upstream ERK1/2 activator, MAPK kinase (MEK) 1/2, and downstream substrate, c-Myc, were activated. By blocking activation of ERK1/2 with a MEK-specific inhibitor, PD98059, CD40-dependent secretion of the pro-inflammatory cytokines, TNF-alpha, IL-6 and IL-8, was demonstrated to be linked to the ERK1/2 pathway. The ERK1/2 pathway did not appear to be involved in up-regulating TRAF1 and TRAF5 mRNAs in THP1 cells. Collectively, these results suggest distinct differences between B cells and monocytic cells in CD40-dependent activation of MAPK pathways.

B-Lymphocytes↗

PubMed2006

Structural modeling and analysis of signaling pathways based on Petri nets.

The purpose of this paper is to discuss how to model and analyze signaling pathways by using Petri net. Firstly, we propose a modeling method based on Petri net by paying attention to the molecular interactions and mechanisms. Then, we introduce a new notion "activation transduction component" in order to describe an enzymic activation process of reactions in signaling pathways and shows its correspondence to a so-called elementary T-invariant in the Petri net models. Further, we design an algorithm to effectively find basic enzymic activation processes by obtaining a series of elementary T-invariants in the Petri net models. Finally, we demonstrate how our method is practically used in modeling and analyzing signaling pathway mediated by thrombopoietin as an example.

Algorithms↗

PubMed1996

Retinoic acid receptor- and retinoid X receptor-selective retinoids activate signaling pathways that converge on AP-1 and inhibit squamous differentiation in human bronchial epithelial cells.

Human bronchial epithelial (HBE) cells undergo squamous differentiation in response to a variety of conditions in tissue culture, and retinoid treatment has been shown to reverse this process. Retinoids mediate their effects through the retinoic acid and retinoid X nuclear receptors (RAR and RXR, respectively), which form RAR-RXR heterodimers, RXR homodimers, and heterodimers of RXR and certain orphan receptors. These receptor dimers bind to distinct response elements, activating separate pathways. In this study, we investigated the roles of RAR and RXR signaling pathways in the inhibition of HBE squamous differentiation. After induction of squamous differentiation by confluent growth, HBE cells were treated with retinoids that selectively activate RARs (E-4-[2-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-2-naphthaienyl)-1- propenyl] benzoic acid), RXRs (4-[1-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-2-naphthyl)ethenyl]benzoic acid), or both RARs and RXRs (9-c/s retinoic acid). These retinoids inhibited the mRNA expression of the squamous differentiation markers transglutaminase type I, involucrin, keratin 5, and keratin 13, suggesting that inhibition of HBE squamous differentiation could be mediated by activation of either RAR or RXR signaling pathways. We examined the role of AP-1 as a potential effector of these retinoid pathways. AP-1 transcriptional activity was reduced markedly by these retinoids, and a concomitant, but proportionally smaller, reduction in AP-1 DNA binding was observed. Furthermore, treatment of squamous HBE cells with the retinoid SR11238, which inhibited AP-1 without activating retinoid receptor transcriptional properties, reduced the expression of transglutaminase type I and involucrin. These findings support the hypothesis that, in HBE cells, RAR and RXR signaling pathways inhibit AP-1 transcriptional activity, and this contributes to retinoid-induced reversal of HBE squamous differentiation.

Alitretinoin↗

PubMed1998

beta-TrCP is a negative regulator of Wnt/beta-catenin signaling pathway and dorsal axis formation in Xenopus embryos.

The Wnt/beta-catenin signaling pathway is responsible for the establishment of dorsoventral axis of Xenopus embryos. The recent finding of the F-box/WD40-repeat protein slimb in Drosophila, whose loss-of-function mutation causes ectopic activation of wingless signaling (Jiang, J., Struhl, G., 1998. Nature 391, 493-496), led us to examine the role of its vertebrate homolog betaTrCp in the Wnt/beta-catenin signaling and dorsal axis formation in Xenopus embryos. Co-injection of betaTrCp mRNA diminished Xwnt8 mRNA-induced axis formation and expression of Siamois and Xnr3, suggesting that betaTrCP is a negative regulator of the Wnt/beta-catenin signaling pathway. An mRNA for a betaTrCp mutant construct (DeltaF), which lacked the F-box domain, induced an ectopic axis and expression of Siamois and Xnr3. Because this activity of DeltaF was suppressed by co-injection of DeltaF TrCP mRNA, DeltaF likely acts in a dominant negative fashion. The activity of DeltaF was diminished by C-cadherin, glycogen synthase kinase 3 and Axin, but not by a dominant negative dishevelled. These results suggest that betaTrCp can act as a negative regulator of dorsal axis formation in Xenopus embryos.

Animals↗

PubMed2005

Tobacco ZFT1, a transcriptional repressor with a Cys2/His2 type zinc finger motif that functions in spermine-signaling pathway.

We previously proposed that a spermine (Spm)-mediated signal transduction pathway is involved in the hypersensitive response induced by Tobacco mosaic virus (TMV) in tobacco plants. To identify regulatory component(s) of this pathway, we surveyed a tobacco cDNA library and found that the ZFT1 gene, which encodes a Cys2/His2 type zinc-finger protein, is Spm-responsive. ZFT1 was not induced by two other polyamines, putrescine and spermidine, or by salicylic acid (SA), jasmonic acid or ethylene. Furthermore, ZFT1 was upregulated in TMV- inoculated tobacco plants in an N gene-dependent manner. Notably, induction of ZFT1 by Spm and by TMV infection was unimpaired in NahG-transgenic tobacco plants, indicating that cross-talk with an SA signaling pathway is not involved in this response. Within the Spm-signaling pathway, we found that ZFT1 functioned downstream of both mitochondrial dysfunction and mitogen-activated protein kinase activation. The ZFT1 protein has two zinc finger motifs and shows a high degree of similarity to ZPT2-3 in petunia and SCOF1 in soybean. However, unlike the latter two proteins, ZFT1 binds to the EP1S sequence and functions as a transcription repressor. Moreover, interestingly, ZFT1 overexpression rendered tobacco plants more tolerant to TMV. Based on the results presented here, we propose that ZFT1 functions as a transcription repressor in a Spm signaling pathway, thereby accelerating necrotic local region formation in tobacco leaves.

Amino Acid Sequence↗

PubMed2006

Regulation of phosphatidylinositol 3-kinase (PI3K)/Akt and nuclear factor-kappa B signaling pathways in dystrophin-deficient skeletal muscle in response to mechanical stretch.

Phosphatidylinositol 3-kinase (PI3K)/Akt and nuclear factor-kappa B (NF-kappaB) signaling pathways play a critical role in mediating survival signals. In this study we have investigated how loss of dystrophin (the primary cause of Duchenne muscular dystrophy) modulates the activation of PI3K/Akt and NF-kappaB signaling pathways in skeletal muscle in response to mechanical stimulation. Activation of Akt was significantly higher in diaphragm muscle from dystrophin-deficient mdx mice compared to normal mice at both prenecrotic and necrotic states. Higher activation of Akt was also observed in cultured dystrophin-deficient primary myotubes differentiated in vitro. Application of passive mechanical stretch ex vivo synergistically increased the activation of Akt in diaphragm of mdx mice. Stretch-induced activation of PDK-1 and PI3K were also higher in diaphragm of mdx mice compared to normal mice. Pretreatment of diaphragm with PI3K inhibitor LY294002 blocked the activation of Akt in normal and mdx mice. Higher activation of Akt was associated with increased phosphorylation of its downstream targets glycogen synthase kinase 3beta (GSK3beta), FKHR, and mammalian target of rapamycin (mTOR). Treatment of diaphragm muscle with LY294002 inhibited the stretch-induced activation of IkappaB (IkappaB) kinase (IKK) and NF-kappaB transcription factor in normal and mdx mice. Mechanical stretch also reduced the interaction of HDAC1 with RelA subunit of NF-kappaB in diaphragm muscle. Finally, cellular levels of Bcl-2, cIAP1, and integrin beta1 and activation of integrin linked kinase were higher in diaphragm muscle of mdx mice compared to normal mice. Taken together, our data suggest that loss of dystrophin and/or mechanical stretch results in the up-regulation of P13K/Akt and NF-kappaB signaling pathways in skeletal muscle.

Animals↗