Integrin 3 1-dependent -catenin phosphorylation links epithelial Smad signaling to cell contacts

Pulmonary and Critical Care Division, Department of Medicine, and Cardiovascular Research Institute, University of California, San Francisco, San Francisco, CA 94143, USA.
The Journal of Cell Biology (Impact Factor: 9.83). 02/2009; 184(2):309-22. DOI: 10.1083/jcb.200806067
Source: PubMed


Injury-initiated epithelial to mesenchymal transition (EMT) depends on contextual signals from the extracellular matrix, suggesting a role for integrin signaling. Primary epithelial cells deficient in their prominent laminin receptor, alpha3beta1, were found to have a markedly blunted EMT response to TGF-beta1. A mechanism for this defect was explored in alpha3-null cells reconstituted with wild-type (wt) alpha3 or point mutants unable to engage laminin 5 (G163A) or epithelial cadherin (E-cadherin; H245A). After TGF-beta1 stimulation, wt epithelial cells but not cells expressing the H245A mutant internalize complexes of E-cadherin and TGF-beta1 receptors, generate phospho-Smad2 (p-Smad2)-pY654-beta-catenin complexes, and up-regulate mesenchymal target genes. Although Smad2 phosphorylation is normal, p-Smad2-pY654-beta-catenin complexes do not form in the absence of alpha3 or when alpha3beta1 is mainly engaged on laminin 5 or E-cadherin in adherens junctions, leading to attenuated EMT. These findings demonstrate that alpha3beta1 coordinates cross talk between beta-catenin and Smad signaling pathways as a function of extracellular contact cues and thereby regulates responses to TGF-beta1 activation.

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Available from: Xiaopeng Li, Aug 15, 2014
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    • "This morphologic transformation was observed in the absence of BDNF addition in independent clones expressing TrkB, and was neither observed in the empty vector clones nor in the clones expressing kinase-inactive TrkB (Figure 4A and B). In order to exclude the possibility that unwanted alterations that inhibited declustering had occurred in any of the cell clones during selection or passaging, cells were also grown on fibronectin which leads to cell dispersal and EMT in primary alveolar epithelial cells as reported previously [32]. Culturing the cell clones on fibronectin revealed a similar extent of cell dispersal in all clones arguing against passaging-induced alterations (Figure 4A). "
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    ABSTRACT: TrkB mediates the effects of brain-derived neurotrophic factor (BDNF) in neuronal and nonnneuronal cells. Based on recent reports that TrkB can also be transactivated through epidermal growth-factor receptor (EGFR) signaling and thus regulates migration of early neurons, we investigated the role of TrkB in migration of lung tumor cells. Early metastasis remains a major challenge in the clinical management of non-small cell lung cancer (NSCLC). TrkB receptor signaling is associated with metastasis and poor patient prognosis in NSCLC. Expression of this receptor in A549 cells and in another adenocarcinoma cell line, NCI-H441, promoted enhanced migratory capacity in wound healing assays in the presence of the TrkB ligand BDNF. Furthermore, TrkB expression in A549 cells potentiated the stimulatory effect of EGF in wound healing and in Boyden chamber migration experiments. Consistent with a potential loss of cell polarity upon TrkB expression, cell dispersal and de-clustering was induced in A549 cells independently of exogeneous BDNF. Morphological transformation involved extensive cytoskeletal changes, reduced E-cadherin expression and suppression of E-cadherin expression on the cell surface in TrkB expressing tumor cells. This function depended on MEK and Akt kinase activity but was independent of Src. These data indicate that TrkB expression in lung adenoma cells is an early step in tumor cell dissemination, and thus could represent a target for therapy development.
    PLoS ONE 06/2014; 9(6):e100944. DOI:10.1371/journal.pone.0100944 · 3.23 Impact Factor
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    • "We observed an increase of miR-21 in isolated lung epithelial cells in which EMT occurred in vivo during bleomycin-induced lung injury. To clarify whether culture condition that induce EMT also increased miR-21 in alveolar epithelial type II cells cultured in vitro, we cultured mouse alveolar epithelial type II cells and induced EMT using endogenous and exogenous TGF-β as previously reported [23,24]. For cultured mouse alveolar type II cells, lung cell suspensions were prepared by intratracheal instillation of Dispase and agarose, followed by mechanical disaggregation of the lungs according to the method of Corti et al. [21]. "
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    ABSTRACT: The excess and persistent accumulation of fibroblasts due to aberrant tissue repair results in fibrotic diseases such as idiopathic pulmonary fibrosis. Recent reports have revealed significant changes in microRNAs during idiopathic pulmonary fibrosis and evidence in support of a role for microRNAs in myofibroblast differentiation and the epithelial-mesenchymal transition in the context of fibrosis. It has been reported that microRNA-21 is up-regulated in myofibroblasts during fibrosis and promotes transforming growth factor-beta signaling by inhibiting Smad7. However, expression changes in microRNA-21 and the role of microRNA-21 in epithelial-mesenchymal transition during lung fibrosis have not yet been defined. Lungs from saline- or bleomycin-treated C57BL/6 J mice and lung specimens from patients with idiopathic pulmonary fibrosis were analyzed. Enzymatic digestions were performed to isolate single lung cells. Lung epithelial cells were isolated by flow cytometric cell sorting. The expression of microRNA-21 was analyzed using both quantitative PCR and in situ hybridization. To induce epithelial-mesenchymal transition in culture, isolated mouse lung alveolar type II cells were cultured on fibronectin-coated chamber slides in the presence of transforming growth factor-beta, thus generating conditions that enhance epithelial-mesenchymal transition. To investigate the role of microRNA-21 in epithelial-mesenchymal transition, we transfected cells with a microRNA-21 inhibitor. Total RNA was isolated from the freshly isolated and cultured cells. MicroRNA-21, as well as mRNAs of genes that are markers of alveolar epithelial or mesenchymal cell differentiation, were quantified using quantitative PCR. The lung epithelial cells isolated from the bleomycin-induced lung fibrosis model system had decreased expression of epithelial marker genes, whereas the expression of mesenchymal marker genes was increased. MicroRNA-21 was significantly upregulated in isolated lung epithelial cells during bleomycin-induced lung fibrosis and human idiopathic pulmonary fibrosis. MicroRNA-21 was also upregulated in the cultured alveolar epithelial cells under the conditions that enhance epithelial-mesenchymal transition. Exogenous administration of a microRNA-21 inhibitor prevented the increased expression of vimentin and alpha-smooth muscle actin in cultured primary mouse alveolar type II cells under culture conditions that induce epithelial-mesenchymal transition. Our experiments demonstrate that microRNA-21 is increased in lung epithelial cells during lung fibrosis and that it promotes epithelial-mesenchymal transition.
    Respiratory research 09/2013; 14(1):95. DOI:10.1186/1465-9921-14-95 · 3.09 Impact Factor
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    • "In addition, we have previously shown that Fgf10 secreted by the PSMCs modulates the differentiation of Clara cells into goblet cells [22], which is a hallmark of the asthmatic airway. Future experiments will be needed to determine if loss of mesenchymal c-Myc may also reduce proliferation of (myo)fibroblasts in the bleomycin model of pulmonary fibrosis, in which Wnt signaling plays an important role [55], [56], [57], [58], [59], [60], [61]. If so, targeting c-Myc might be an effective and selective way to treat fibroproliferative lung diseases in general. "
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    ABSTRACT: During lung development, Fibroblast growth factor 10 (Fgf10), which is expressed in the distal mesenchyme and regulated by Wnt signaling, acts on the distal epithelial progenitors to maintain them and prevent them from differentiating into proximal (airway) epithelial cells. Fgf10-expressing cells in the distal mesenchyme are progenitors for parabronchial smooth muscle cells (PSMCs). After naphthalene, ozone or bleomycin-induced airway epithelial injury, surviving epithelial cells secrete Wnt7b which then activates the PSMC niche to induce Fgf10 expression. This Fgf10 secreted by the niche then acts on a subset of Clara stem cells to break quiescence, induce proliferation and initiate epithelial repair. Here we show that conditional deletion of the Wnt target gene c-Myc from the lung mesenchyme during development does not affect proper epithelial or mesenchymal differentiation. However, in the adult lung we show that after naphthalene-mediated airway epithelial injury c-Myc is important for the activation of the PSMC niche and as such induces proliferation and Fgf10 expression in PSMCs. Our data indicate that conditional deletion of c-Myc from PSMCs inhibits airway epithelial repair, whereas c-Myc ablation from Clara cells has no effect on airway epithelial regeneration. These findings may have important implications for understanding the misregulation of lung repair in asthma and COPD.
    PLoS ONE 08/2013; 8(8):e71426. DOI:10.1371/journal.pone.0071426 · 3.23 Impact Factor
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