Abstract
The epithelial-mesenchymal transition (EMT) is involved in neoplastic metastasis, and the RON protein may be involved. In the present study, we determined the role and the mechanisms of action of RON in EMT in Madin-Darby canine kidney (MDCK) cells by Western blot and cell migration analysis. Activation of RON by macrophage stimulating protein (MSP) results in cell migration and initiates changes in the morphology of RON-cDNA-transfected MDCK cells. The absence of E-cadherin, the presence of vimentin and an increase in Snail were observed in RE7 cells, which were derived from MDCK cells transfected with wt-RON, compared with MDCK cells. Stimulation of RE7 cells with MSP resulted in increased migration (about 69% of the wounded areas were covered) as well as increased activation of extracellular signal-regulated kinase 1/2 (Erk1/2) and glycogen synthase kinase-3β (GSK-3β; the percent of the activation ratio was 143.6/599.8% and 512.4%, respectively), which could be inhibited with an individual chemical inhibitor PD98059 (50 μM) specific to MAPK/ERK kinase (the percent inhibition was 98.9 and 81.2%, respectively). Thus, the results indicated that RON protein could mediate EMT in MDCK cells via the Erk1/2 pathway. Furthermore, GSK-3β regulates the function of Snail in controlling EMT by this pathway.
Highlights
Epithelial-mesenchymal transition (EMT) is a biologic process that allows a polarized epithelial cell, which normally interacts with the basement membrane via its basal surface, to undergo multiple biochemical changes that enable the epithelial cell to assume a mesenchymal cell phenotype
Through a series of experiments, we demonstrated that RON could mediate EMT in Madin-Darby canine kidney (MDCK) cells through the extracellular signal-regulated kinase 1/2 (Erk1/2) pathway
Goat or rabbit IgG antibodies specific for Pan-Erk, phosphor-Erk1/2 (p44/42), E-cadherin, vimentin, Snail, Twist, regular or phosphor glycogen synthase kinase-3β (GSK-3β) and AKT were purchased from Cell Signaling Technology (USA) and Santa Cruz Biotechnology (USA), respectively
Summary
Epithelial-mesenchymal transition (EMT) is a biologic process that allows a polarized epithelial cell, which normally interacts with the basement membrane via its basal surface, to undergo multiple biochemical changes that enable the epithelial cell to assume a mesenchymal cell phenotype. This, in turn, provides the cell with enhanced migratory capacity, invasiveness, and increased resistance to apoptosis [1]. The completion of an EMT is signaled by the degradation of the underlying basement membrane and the formation of a mesenchymal cell that can migrate away from the epithelial layer from which it originated. In EMT, epithelial cells acquire fibroblast-like properties and show reduced intercellular adhesion and increased motility. This process is associated with the functional loss of E-cadherin [2,3]. Down-regulation of E-cadherin is the key step towards the invasive phase of carcinoma, and dominant transcriptional repression is largely responsible for the loss of E-cadherin expression [4,5,6,7,8]
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