Overexpression of the Integrin-linked Kinase Promotes Anchorage-independent Cell Cycle Progression*
- Galina Radeva,
- Teresa Petrocelli,
- Elke Behrend,
- Chungyee Leung-Hagesteijn,
- Jorge Filmus‡,
- Joyce Slingerland§ and
- Shoukat Dedhar¶
- From the Department of Medical Biophysics, University of Toronto and Cancer Biology Research, Sunnybrook Health Science Centre, Toronto, Ontario M4N 3M5, Canada
Abstract
Cell adhesion to substratum has been shown to regulate cyclin A expression as well as cyclin D- and E-dependent kinases, the latter via the up-regulation of cyclin D1 and the down-regulation of cyclin-Cdk inhibitors p21 and p27, respectively. This adhesion-dependent regulation of cell cycle is thought to be mediated by integrins. Here we demonstrate that stable transfection and overexpression of theintegrin-linked kinase (ILK), which interacts with the β1 and β3 integrin cytoplasmic domains, induces anchorage-independent cell cycle progression but not serum-independent growth of rat intestinal epithelial cells (IEC18). ILK overexpression results in increased expression of cyclin D1, activation of Cdk4 and cyclin E-associated kinases, and hyperphosphorylation of the retinoblastoma protein. In addition, ILK overexpression results in the expression of p21 and p27 Cdk inhibitors with altered electrophoretic mobilities, with the p27 from ILK-overexpressing cells having reduced inhibitory activity. The transfer of serum-exposed IEC18 cells from adherent cultures to suspension cultures results in a rapid down-regulation of expression of cyclin D1 and cyclin A proteins as well as in retinoblastoma protein dephosphorylation. In marked contrast, transfer of ILK-overexpressing cells from adherent to suspension cultures results in continued high levels of expression of cyclin D1 and cyclin A proteins, and a substantial proportion of the retinoblastoma protein remains in a hyperphosphorylated state. These results indicate that, when overexpressed, ILK induces signaling pathways resulting in the stimulation of G1/S cyclin-Cdk activities, which are normally regulated by cell adhesion and integrin engagement.
Footnotes
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↵* This work was supported in part by grants from the National Cancer Institute of Canada and the Medical Research Council of Canada.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵‡ Supported by the Medical Research Council of Canada.
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↵§ Clinician scientist supported by the Ontario Cancer Treatment and Research Foundation.
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↵¶ Terry Fox Cancer Scientist of the National Cancer Institute of Canada. To whom correspondence should be addressed: Dept. of Medical Biophysics, University of Toronto and Cancer Biology Research, Sunnybrook Health Science Centre, 2075 Bayview Ave., Rm S-218, Toronto, Ontario M4N 3M5, Canada.
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↵1 The abbreviations used are: ECM, extracellular matrix; MAPK, mitogen-activated protein kinase; Rb, retinoblastoma; ILK, integrin-linked kinase; PBS, phosphate-buffered saline; FACS, fluorescence-activated cell sorting; PMSF, phenylmethylsulfonyl fluoride; PAGE, polyacrylamide gel electrophoresis; HRP, horseradish peroxidase.
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↵2 C. Wu, S. Y. Keightley, C. Leung-Hagesteijn, G. Radeva, J. McDonald, and S. Dedhar, submitted for publication.
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↵3 D. Hackam, E. Behrend, and S. Dedhar, unpublished observations.
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- Received November 25, 1996.
- Revision received February 28, 1997.
