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Growth factor induced activation of Rho and Rac GTPases and actin cytoskeletal reorganization in human lens epithelial cells.

Publication ,  Journal Article
Maddala, R; Reddy, VN; Epstein, DL; Rao, V
Published in: Mol Vis
July 17, 2003

PURPOSE: To determine the involvement of the Rho GTPases-mediated signaling pathway in growth factor-stimulated actomyosin cytoskeletal organization and focal adhesion formation in lens epithelial cells. METHODS: Serum starved human lens epithelial cells (SRA01/04) were treated with different growth factors including epidermal growth factor (EGF), basic-fibroblast growth factor (b-FGF), platelet derived growth factor (PDGF), transforming growth factor beta (TGF-beta), insulin-like growth factor 1 (IGF-1), lysophosphatidic acid (LPA), and thrombin. Growth factor stimulated activation of Rho and Rac GTPases were evaluated by GTP-loading pull-down assays. Changes in actin cytoskeletal organization and focal adhesions were determined by fluorescence staining using FITC-phalloidin and anti-vinculin antibody/rhodamine-conjugated secondary antibody, respectively. Fluorescence images were recorded using either confocal or fluorescence microscopy. RESULTS: Rho GTPase activity was significantly augmented in human lens epithelial cells treated with EGF, b-FGF, TGF-beta, IGF-1, and LPA. Rac GTPase activation, in contrast, was significantly enhanced in response to only EGF or b-FGF. Serum starved human lens epithelial cells exhibited a strong increase in cortical actin stress fibers and integrin-mediated focal adhesions in response to b-FGF, PDGF, TGF-beta, thrombin, and LPA. While EGF induced a striking increase in membrane ruffling and a marginal increase on focal adhesion formation, IGF-1 had no effect on either. Pretreatment of lens epithelial cells with C3-exoenzyme (an irreversible inhibitor of Rho-GTPase), lovastatin (an isoprenylation inhibitor), or the Rho kinase inhibitor Y-27632 abolished the ability of the different growth factors to elicit actin stress fiber and focal adhesion formation. EGF induced membrane ruffling, however, was not suppressed by Y-27632 and C3-exoenzyme. CONCLUSIONS: These results demonstrate that different growth factors induce actin cytoskeleton reorganization and formation of cell-ECM interactions in lens epithelial cells and this response of growth factors appears to be mediated, at least in part, through the Rho/Rho kinase-mediated signaling pathway. The ability of growth factors to trigger activation of Rho and Rac GTPases along with actomyosin cytoskeletal reorganization and formation of focal adhesions might well play a crucial role in lens epithelial cell proliferation, migration, elongation and survival.

Duke Scholars

Published In

Mol Vis

EISSN

1090-0535

Publication Date

July 17, 2003

Volume

9

Start / End Page

329 / 336

Location

United States

Related Subject Headings

  • rho GTP-Binding Proteins
  • rac GTP-Binding Proteins
  • Signal Transduction
  • Ophthalmology & Optometry
  • Microscopy, Fluorescence
  • Microscopy, Confocal
  • Lens, Crystalline
  • Humans
  • Growth Substances
  • Fluorescent Antibody Technique, Indirect
 

Citation

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Maddala, R., Reddy, V. N., Epstein, D. L., & Rao, V. (2003). Growth factor induced activation of Rho and Rac GTPases and actin cytoskeletal reorganization in human lens epithelial cells. Mol Vis, 9, 329–336.
Maddala, Rupalatha, Venkat N. Reddy, David L. Epstein, and Vasantha Rao. “Growth factor induced activation of Rho and Rac GTPases and actin cytoskeletal reorganization in human lens epithelial cells.Mol Vis 9 (July 17, 2003): 329–36.

Published In

Mol Vis

EISSN

1090-0535

Publication Date

July 17, 2003

Volume

9

Start / End Page

329 / 336

Location

United States

Related Subject Headings

  • rho GTP-Binding Proteins
  • rac GTP-Binding Proteins
  • Signal Transduction
  • Ophthalmology & Optometry
  • Microscopy, Fluorescence
  • Microscopy, Confocal
  • Lens, Crystalline
  • Humans
  • Growth Substances
  • Fluorescent Antibody Technique, Indirect