Expression of osterix Is Regulated by FGF and Wnt/β-Catenin Signalling during Osteoblast Differentiation.
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ABSTRACT:
SUBMITTER: Felber K
PROVIDER: S-EPMC4686927 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BackgroundN-myc downstream regulated gene 1 (NDRG1) plays a role in a variety of biological processes including differentiation of osteoclasts. However, it is not known if and how NDRG1 regulates osteogenic differentiation of marrow stromal progenitor cells.MethodsGene expression profiling analysis was performed to study the expression level of Ndrg1 during osteogenic and adipogenic differentiation. Gain-of-function and/or loss-of function experiments were carried out to study the role of NDRG1 in the proliferation and differentiation of marrow stromal progenitor cells and the mechanism underlying the function was investigated. Finally, in vivo transfection of Ndrg1 siRNA was done and its effect on osteogenic and adipogenic differentiation in mice was explored.ResultsGene expression profiling analysis revealed that NDRG1 level was regulated during osteogenic and adipogenic differentiation of progenitor cells. The functional experiments demonstrated that NDRG1 negatively regulated the cell growth, and reciprocally modulated the osteogenic and adipogenic commitment of marrow stromal progenitor cells, driving the cells to differentiate toward adipocytes at the expense of osteoblast differentiation. Moreover, NDRG1 interacted with low-density lipoprotein receptor-related protein 6 (LRP6) in the stromal progenitor cells and inactivated the canonical Wnt/β-catenin signaling cascade. Furthermore, the impaired differentiation of progenitor cells induced by Ndrg1 siRNA could be attenuated when β-catenin was simultaneously silenced. Finally, in vivo transfection of Ndrg1 siRNA to the marrow of mice prevented the inactivation of canonical Wnt signaling in the BMSCs of ovariectomized mice, and ameliorated the reduction of osteoblasts on the trabeculae and increase of fat accumulation in the marrow observed in the ovariectomized mice.ConclusionThis study has provided evidences that NDRG1 plays a role in reciprocally modulating osteogenic and adipogenic commitment of marrow stromal progenitor cells through inactivating canonical Wnt signaling.
| S-EPMC8817551 | biostudies-literature
Project description:Commitment of Runx2-expressing precursor osteoblasts to functional osteoblasts and then to osteocytes is triggered by Osterix (Osx), which activates its target genes in those cells during bone formation. It is not yet known whether Osx has a role in remodeling the chromatin architecture of its target genes during the transition from preosteoblast to osteoblast. In testing the hypothesis that Osx is indispensable for active chromatin architecture, we first showed that in Osx-null calvarial cells occupancy of the transcriptional activators, including lysine 4 methyl transferase (Wdr5), c-Myc, and H2A.Z, at the Osx target gene Bsp was very markedly decreased. The levels of methylation of lysines 4 and 36 and acetylation of histone H3, markers for active chromatin, were also reduced at the Bsp gene in these cells. In contrast, occupancy of the transcriptional repressors HP1 and the nucleolar protein 66 (NO66), a histone demethylase previously identified as an Osx-interacting protein, was increased at the Bsp gene in Osx-null calvarial cells. Furthermore, the Bsp promoter was hypermethylated in embryonic stem (ES) cells and in embryonic day 9.5 (E9.5) embryos but was markedly hypomethylated in the calvaria of E18.5 embryos, coinciding with robust Bsp expression. In contrast, CpG methylation in the Bsp promoter remained high in Osx-null calvaria compared to Osx-wild-type calvaria. Our data also revealed that NO66 interacted with DNA Methyltransferase 1A (DNMT1A), histone deacetylase 1A (HDAC1A), and HP1, which are known to control histone and DNA methylation. In addition, HP1 stimulated the demethylase activity of NO66 for its substrates "trimethylation of histone H3 at lysine 4" (H3K4me3) and "trimethylation of histone H3 at lysine 36" (H3K36me3). Our findings strongly suggest that in the absence of Osx, the chromatin of Osx target genes is transcriptionally inactive. We propose that Osx is a molecular switch for the formation of an active chromatin state during osteoblast differentiation, whereas NO66 helps gene repression through histone demethylation and/or formation of a repressor complex, resulting in multilayered control of the chromatin architecture of specific osteoblast genes.
| S-EPMC3961497 | biostudies-literature
Project description:Bone formation is a dynamic process directed by osteoblast activity. The transition from the proliferation to differentiation stage during osteoblast maturation involves the downregulation of the Wnt/β-catenin signaling pathway, and extracellular antagonists are important for the regulation of Wnt signaling. However, the expression levels of Wnt antagonists in these stages of human osteoblast maturation have not been fully elucidated. Therefore, the aim of the present study was to investigate the expression levels of extracellular Wnt antagonists during proliferation and differentiation in osteoblast-like cell lines. The results demonstrated an overlap between the differential expression of secreted Frizzled-related protein (SFPR)2, SFRP3, SFRP4 and Dickkopf (DKK) 2 genes during the differentiation stage in the MG-63 and Saos-2 cells. Furthermore, high expression levels of DKK3 in MG-63 cells, Wnt inhibitory factor 1 (WIF1) in Saos-2 cells and DKK4 in hFOB 1.19 cells during the same stage (differentiation), were observed. The upregulated expression levels of Wnt antagonists were also correlated with the high expression of anxin 2 during the differentiation stage. These findings suggested that Wnt-related antagonists could modulate the Wnt/β-catenin signaling pathway. By contrast, DKK1 was the only gene that was found to be upregulated during the proliferation stage in hFOB 1.19 and Saos-2 cells. To the best of our knowledge, the present study provides, for the first time, the expression profile of Wnt antagonists during the proliferation stage and the initial phases of differentiation in osteoblast-like cell lines. The current results offer a basis to investigate potential targets for bone-related Wnt-signaling modulation in bone metabolism research.
| S-EPMC7654218 | biostudies-literature
Project description:Patients with Hutchinson-Gilford progeria syndrome (HGPS) have low bone mass and an atypical skeletal geometry that manifests in a high risk of fractures. Using both in vitro and in vivo models of HGPS, we demonstrate that defects in the canonical WNT/β-catenin pathway, seemingly at the level of the efficiency of nuclear import of β-catenin, impair osteoblast differentiation and that restoring β-catenin activity rescues osteoblast differentiation and significantly improves bone mass. Specifically, we show that HGPS patient-derived iPSCs display defects in osteoblast differentiation, characterized by a decreased alkaline phosphatase activity and mineralizing capacity. We demonstrate that the canonical WNT/β-catenin pathway, a major signaling cascade involved in skeletal homeostasis, is impaired by progerin, causing a reduction in the active β-catenin in the nucleus and thus decreased transcriptional activity, and its reciprocal cytoplasmic accumulation. Blocking farnesylation of progerin restores active β-catenin accumulation in the nucleus, increasing signaling, and ameliorates the defective osteogenesis. Moreover, in vivo analysis of the Zmpste24-/- HGPS mouse model demonstrates that treatment with a sclerostin-neutralizing antibody (SclAb), which targets an antagonist of canonical WNT/β-catenin signaling pathway, fully rescues the low bone mass phenotype to wild-type levels. Together, this study reveals that the β-catenin signaling cascade is a therapeutic target for restoring defective skeletal microarchitecture in HGPS. © 2018 American Society for Bone and Mineral Research.
| S-EPMC7739562 | biostudies-literature
Project description:Wnt signalling has pivotal roles in tumour progression and metastasis; however, the exact molecular mechanism of Wnt signalling in the metastatic process is as yet poorly defined. Here we demonstrate that the tumour metastasis suppressor gene, NDRG1, interacts with the Wnt receptor, LRP6, followed by blocking of the Wnt signalling, and therefore, orchestrates a cellular network that impairs the metastatic progression of tumour cells. Importantly, restoring NDRG1 expression by a small molecule compound significantly suppressed the capability of otherwise highly metastatic tumour cells to thrive in circulation and distant organs in animal models. In addition, our analysis of clinical cohorts data indicate that Wnt+/NDRG-/LRP+ signature has a strong predictable value for recurrence-free survival of cancer patients. Collectively, we have identified NDRG1 as a novel negative master regulator of Wnt signalling during the metastatic progression, which opens an opportunity to define a potential therapeutic target for metastatic disease.
| S-EPMC3306556 | biostudies-literature
Project description:The Wnt/beta-catenin signalling pathway appears to operate to maintain the undifferentiated state of preadipocytes by inhibiting adipogenic gene expression. To define the mechanisms regulating suppression of Wnt/beta-catenin signalling, we analysed the beta-catenin expression in response to activation of transcription factors that regulate adipogenesis. The results show an extensive down-regulation of nuclear beta-catenin that occurs during the first few days of differentiation of 3T3-L1 preadipocytes and coincides with the induction of the adipogenic transcription factors, C/EBPbeta (CCAAT-enhancer-binding protein) and PPARgamma (peroxisome-proliferator-activated receptor). To assess the role of each of these factors in this process, we conditionally overexpressed C/EBPbeta in Swiss mouse fibroblasts using the TET-off system. Abundant expression of C/EBPbeta alone had minimal effect on beta-catenin expression, whereas expression of C/EBPbeta, in the presence of dexamethasone, induced PPARgamma expression and caused a measurable decrease in beta-catenin. In addition, exposure of cells expressing both C/EBPbeta and PPARgamma to a potent PPARgamma ligand resulted in an even greater decrease in beta-catenin by mechanisms that involve the proteasome. Our studies also suggest a reciprocal relationship between PPARgamma activity and beta-catenin expression, since ectopic production of Wnt-1 in preadipocytes blocked the induction of PPARgamma gene expression. Moreover, by suppressing beta-catenin expression, ectopic expression of PPARgamma in Wnt-1-expressing preadipocytes rescued the block in adipogenesis after their exposure to the PPARgamma ligand, troglitazone.
| S-EPMC1223802 | biostudies-other
Project description:Wnt proteins modulate cell proliferation and differentiation and the self-renewal of stem cells by inducing β-catenin-dependent signalling through the Wnt receptor frizzled (FZD) and the co-receptors LRP5 and LRP6 to regulate cell fate decisions and the growth and repair of several tissues. The 19 mammalian Wnt proteins are cross-reactive with the 10 FZD receptors, and this has complicated the attribution of distinct biological functions to specific FZD and Wnt subtype interactions. Furthermore, Wnt proteins are modified post-translationally by palmitoylation, which is essential for their secretion, function and interaction with FZD receptors. As a result of their acylation, Wnt proteins are very hydrophobic and require detergents for purification, which presents major obstacles to the preparation and application of recombinant Wnt proteins. This hydrophobicity has hindered the determination of the molecular mechanisms of Wnt signalling activation and the functional importance of FZD subtypes, and the use of Wnt proteins as therapeutic agents. Here we develop surrogate Wnt agonists, water-soluble FZD-LRP5/LRP6 heterodimerizers, with FZD5/FZD8-specific and broadly FZD-reactive binding domains. Similar to WNT3A, these Wnt agonists elicit a characteristic β-catenin signalling response in a FZD-selective fashion, enhance the osteogenic lineage commitment of primary mouse and human mesenchymal stem cells, and support the growth of a broad range of primary human organoid cultures. In addition, the surrogates can be systemically expressed and exhibit Wnt activity in vivo in the mouse liver, regulating metabolic liver zonation and promoting hepatocyte proliferation, resulting in hepatomegaly. These surrogates demonstrate that canonical Wnt signalling can be activated by bi-specific ligands that induce receptor heterodimerization. Furthermore, these easily produced, non-lipidated Wnt surrogate agonists facilitate functional studies of Wnt signalling and the exploration of Wnt agonists for translational applications in regenerative medicine.
| S-EPMC5815871 | biostudies-literature
Project description:Fibrosis in animal models and human diseases is associated with aberrant activation of the Wnt/β-catenin pathway. Despite extensive research efforts, effective therapies are still not available. Myofibroblasts are major effectors, responsible for extracellular matrix deposition. Inhibiting the proliferation of the myofibroblast is crucial for treatment of fibrosis. Proliferation of myofibroblasts can have many triggering effects that result in fibrosis. In recent years, the Wnt pathway has been studied as an underlying factor as a primary contributor to fibrotic diseases. These efforts notwithstanding, the specific mechanisms by which Wnt-mediated promotes fibrosis reaction remain obscure. The central role of the transforming growth factor-β (TGF-β) and myofibroblast activity in the pathogenesis of fibrosis has become generally accepted. The details of interaction between these two processes are not obvious. The present investigation was conducted to evaluate the level of sustained expression of fibrosis iconic proteins (vimentin, α-SMA and collagen I) and the TGF-β signalling pathway that include smad2/3 and its phosphorylated form p-smad2/3. Detailed analysis of the possible molecular mechanisms mediated by β-catenin revealed epithelial-mesenchymal transition and additionally demonstrated transitions of fibroblasts to myofibroblast cell forms, along with increased activity of β-catenin in regulation of the signalling network, which acts to counteract autocrine TGF-β/smad2/3 signalling. A major outcome of this study is improved insight into the mechanisms by which epithelial and mesenchymal cells activated by TGFβ1-smad2/3 signalling through Wnt/β-catenin contribute to lung fibrosis.
| S-EPMC5542906 | biostudies-other
Project description:Background:Wnt/β-catenin (or canonical) signalling pathway activity is necessary and used independently several times for specification of vegetal fate and endoderm, gut differentiation, maintenance of epithelium in adult intestine and the development of gut-derived organs in various vertebrate and non-vertebrate organisms. However, its conservation in later stages of digestive tract development still remains questionable due to the lack of detailed data, mainly from Spiralia. Results:Here we characterize the Pdu-Tcf gene, a Tcf/LEF orthologue and a component of Wnt/β-catenin pathway from Platynereis dumerilii, a spiralian, marine annelid worm. Pdu-Tcf undergoes extensive alternative splicing in the C-terminal region of the gene generating as many as eight mRNA isoforms some of which differ in the presence or absence of a C-clamp domain which suggests a distinct DNA binding activity of individual protein variants. Pdu-Tcf is broadly expressed throughout development which is indicative of many functions. One of the most prominent domains that exhibits rather strong Pdu-Tcf expression is in the putative precursors of endodermal gut cells which are detected after 72 h post-fertilization (hpf). At day 5 post-fertilization (dpf), Pdu-Tcf is expressed in the hindgut and pharynx (foregut), whereas at 7 dpf stage, it is strongly transcribed in the now-cellularized midgut for the first time. In order to gain insight into the role of Wnt/β-catenin signalling, we disrupted its activity using pharmacological inhibitors between day 5 and 7 of development. The inhibition of Wnt/β-catenin signalling led to the loss of midgut marker genes Subtilisin-1, Subtilisin-2, α-Amylase and Otx along with a drop in β-catenin protein levels, Axin expression in the gut and nearly the complete loss of proliferative activity throughout the body of larva. At the same time, a hindgut marker gene Legumain was expanded to the midgut compartment under the same conditions. Conclusions:Our findings suggest that high Wnt/β-catenin signalling in the midgut might be necessary for proper differentiation of the endoderm to an epithelium capable of secreting digestive enzymes. Together, our data provide evidence for the role of Wnt/β-catenin signalling in gut differentiation in Platynereis.
| S-EPMC5996498 | biostudies-literature
Project description:Tax1 binding protein 3 (Tax1bp3) is a PDZ domain-containing protein that is overexpressed in cancer. Previous studies recognized Tax1bp3 as an inhibitor of β-catenin. Till now it is not known whether Tax1bp3 regulates osteogenic and adipogenic differentiation of mesenchymal progenitor cells. In the current study, the data showed that Tax1bp3 was expressed in bone and was increased in the progenitor cells when induced toward osteoblast and adipocyte differentiation. The overexpression of Tax1bp3 in the progenitor cells inhibited osteogenic differentiation and conversely stimulated adipogenic differentiation, and the knockdown of Tax1bp3 affected the differentiation of the progenitor cells oppositely. Ex vivo experiments using the primary calvarial osteoblasts from osteoblast-specific Tax1bp3 knock-in mice also demonstrated the anti-osteogenic and pro-adipogenic function of Tax1bp3. Mechanistic investigations revealed that Tax1bp3 inhibited the activation of canonical Wnt/β-catenin and bone morphogenetic proteins (BMPs)/Smads signalling pathways. Taken together, the current study has provided evidences demonstrating that Tax1bp3 inactivates Wnt/β-catenin and BMPs/Smads signalling pathways and reciprocally regulates osteogenic and adipogenic differentiation from mesenchymal progenitor cells. The inactivation of Wnt/β-catenin signalling may be involved in the reciprocal role of Tax1bp3.
| S-EPMC10064035 | biostudies-literature