Cbl-transforming Variants Trigger a Cascade of Molecular Alterations That Lead to Epithelial Mesenchymal Conversion

Dispersal of epithelial cells is an important aspect of tumorigenesis, and invasion. Factors such as hepatocyte growth factor induce the breakdown of cell junctions and promote cell spreading and the dispersal of colonies of epithelial cells, providing a model system to investigate the biochemical signals that regulate these events. Multiple signaling proteins are phosphorylated in epithelial cells during hepatocyte growth factor–induced cell dispersal, including c-Cbl, a protooncogene docking protein with ubiquitin ligase activity. We have examined the role of c-Cbl and a transforming variant (70z-Cbl) in epithelial cell dispersal. We show that the expression of 70z-Cbl in Madin-Darby canine kidney epithelial cells resulted in the breakdown of cell–cell contacts and alterations in cell morphology characteristic of epithelial–mesenchymal transition. Structure–function studies revealed that the amino-terminal portion of c-Cbl, which corresponds to the Cbl phosphotyrosine-binding/Src homology domain 2 , is sufficient to promote the morphological changes in cell shape. Moreover, a point mutation at Gly-306 abrogates the ability of the Cbl Src homology domain 2 to induce these morphological changes. Our results identify a role for Cbl in the regulation of epithelial–mesenchymal transition, including loss of adherens junctions, cell spreading, and the initiation of cell dispersal.

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Activation of Cdc42, Rac, PAK, and Rho-Kinase in Response to Hepatocyte Growth Factor Differentially Regulates Epithelial Cell Colony Spreading and Dissociation

Molecular Biology of the Cell ◽

10.1091/mbc.11.5.1709 ◽

2000 ◽

Vol 11 (5) ◽

pp. 1709-1725 ◽

Cited By ~ 195

Author(s):

Isabelle Royal ◽

Nathalie Lamarche-Vane ◽

Louie Lamorte ◽

Kozo Kaibuchi ◽

Morag Park

Keyword(s):

Growth Factor ◽

Epithelial Cell ◽

Hepatocyte Growth Factor ◽

Epithelial Mesenchymal Transition ◽

Rho Kinase ◽

Cell Spreading ◽

Dominant Negative ◽

Phosphatidylinositol 3 Kinase ◽

Mesenchymal Transition ◽

Hepatocyte Growth

Hepatocyte growth factor (HGF), the ligand for the Met receptor tyrosine kinase, is a potent modulator of epithelial–mesenchymal transition and dispersal of epithelial cells, processes that play crucial roles in tumor development, invasion, and metastasis. Little is known about the Met-dependent proximal signals that regulate these events. We show that HGF stimulation of epithelial cells leads to activation of the Rho GTPases, Cdc42 and Rac, concomitant with the formation of filopodia and lamellipodia. Notably, HGF-dependent activation of Rac but not Cdc42 is dependent on phosphatidylinositol 3-kinase. Moreover, HGF-induced lamellipodia formation and cell spreading require phosphatidylinositol 3-kinase and are inhibited by dominant negative Cdc42 or Rac. HGF induces activation of the Cdc42/Rac-regulated p21-activated kinase (PAK) and c-Jun N-terminal kinase, and translocation of Rac, PAK, and Rho-dependent Rho-kinase to membrane ruffles. Use of dominant negative and activated mutants reveals an essential role for PAK but not Rho-kinase in HGF-induced epithelial cell spreading, whereas Rho-kinase activity is required for the formation of focal adhesions and stress fibers in response to HGF. We conclude that PAK and Rho-kinase play opposing roles in epithelial–mesenchymal transition induced by HGF, and provide new insight regarding the role of Cdc42 in these events.

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Hepatocyte Growth Factor (HGF) Attenuates Transforming Growth Factor-β-Induced Epithelial Mesenchymal Transition in Alveolar Epithelial Cells.

10.1164/ajrccm-conference.2009.179.1_meetingabstracts.a5293 ◽

2009 ◽

Author(s):

MS Krishnaveni ◽

B Zhou ◽

BC Willis ◽

ED Crandall ◽

Z Borok

Keyword(s):

Growth Factor ◽

Epithelial Cells ◽

Hepatocyte Growth Factor ◽

Transforming Growth Factor ◽

Epithelial Mesenchymal Transition ◽

Transforming Growth Factor Β ◽

Alveolar Epithelial Cells ◽

Mesenchymal Transition ◽

Alveolar Epithelial ◽

Hepatocyte Growth

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Crk Associates with a Multimolecular Paxillin/GIT2/β-PIX Complex and Promotes Rac-dependent Relocalization of Paxillin to Focal Contacts

Molecular Biology of the Cell ◽

10.1091/mbc.e02-08-0497 ◽

2003 ◽

Vol 14 (7) ◽

pp. 2818-2831 ◽

Cited By ~ 71

Author(s):

Louie Lamorte ◽

Sonia Rodrigues ◽

Veena Sangwan ◽

Christopher E. Turner ◽

Morag Park

Keyword(s):

Growth Factor ◽

Hepatocyte Growth Factor ◽

Adherens Junctions ◽

Cell Spreading ◽

Dependent Manner ◽

Amino Terminal ◽

Focal Contacts ◽

Src Homology ◽

Hepatocyte Growth ◽

Lamellipodia Formation

We have previously demonstrated that the CrkII and CrkL adapter proteins are required for the spreading of epithelial colonies and the breakdown of adherens junctions in response to hepatocyte growth factor. When overexpressed, CrkII and CrkL promote lamellipodia formation, cell spreading, and the loss of epithelial adherens junctions in the absence of hepatocyte growth factor. The exact mechanism by which Crk proteins elicit these changes is unclear. We show that the overexpression of CrkII or CrkL, but not Src homology 2 or amino-terminal Src homology 3 domain mutant Crk proteins, promotes the relocalization of Paxillin to focal contacts throughout the cell and within lamellipodia in a Rac-dependent manner. In stable cell lines overexpressing CrkII, enhanced lamellipodia formation and cell spreading correlate with an increased association of CrkII with Paxillin, GIT2 (an ARF-GAP) and β-PIX (a Rac1 exchange factor). Mutants of Paxillin that fail to associate with Crk or GIT2, or do not target to focal adhesions inhibit Crk-dependent cell spreading and lamellipodia formation. We conclude from these studies that the association of Crk with Paxillin is important for the spreading of epithelial colonies, by influencing the recruitment of Paxillin to focal complexes and promoting the enhanced assembly of Paxillin/GIT2/β-PIX complexes.

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Mesenchymal Stem Cells Attenuates TGF-β1-Induced EMT by Increasing HGF Expression in HK-2 Cells

Iranian Journal of Public Health ◽

10.18502/ijph.v50i5.6108 ◽

2021 ◽

Author(s):

Jun-Jun Wei ◽

Li Tang ◽

Liang-Liang Chen ◽

Zhen-Hua Xie ◽

Yu Ren ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Growth Factor ◽

Hepatocyte Growth Factor ◽

Transforming Growth Factor Beta ◽

Transforming Growth Factor ◽

Epithelial Mesenchymal Transition ◽

Mesenchymal Transition ◽

Hepatocyte Growth ◽

Tgf Β1

Background: Mesenchymal stem cells (MSCs) have recently shown promise for the treatment of various types of chronic kidney disease models. However, the mechanism of this effect is still not well understood. Our study is aimed to investigate the effect of MSCs on transforming growth factor beta 1 (TGF-β1)-induced epithelial mesenchymal transition (EMT) in renal tubular epithelial cells (HK-2 cells) and the underlying mechanism related to the reciprocal balance between hepatocyte growth factor (HGF) and TGF-β1. Methods: Our study was performed at Ningbo University, Ningbo, Zhejiang, China between Mar 2017 and Jun 2018. HK-2 cells were initially treated with TGF-β1，then co-cultured with MSCs. The induced EMT was assessed by cellular morphology and the expressions of alpha-smooth muscle actin (α-SMA) and EMT-related proteins. MTS assay and flow cytometry were employed to detect the effect of TGF-β1 and MSCs on HK-2 cell proliferation and apoptosis. SiRNA against hepatocyte growth factor (siHGF) was transfected to decrease the expression of HGF to identify the role of HGF in MSCs inhibiting HK-2 cells EMT. Results: Overexpressing TGF-β1 decreased HGF expression, induced EMT, suppressed proliferation and promoted apoptosis in HK-2 cells; but when co-cultured with MSCs all the outcomes were reversed. However, after treated with siHGF, all the benefits taken from MSCs vanished. Conclusion: TGF-β1 was a motivating factor of kidney cell EMT and it suppressed the HGF expression. However, MSCs provided protection against EMT by increasing HGF level and decreasing TGF-β1 level. Our results also demonstrated HGF is one of the critical factor in MSCs anti- fibrosis.

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