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Papers In Press, published online ahead of print September 19, 2000
Oncology and Molecular Endocrinology Research Center, CHUL Research Center, Ste-Foy, Quebec G1V 4G2
Corresponding Author: Sylvain.guerin{at}crchul.ulaval.ca
The accumulation of fibronectin (FN) in response to corneal epithelium injury has been postulated to turn on expression of the FN-binding integrin
J. Biol. Chem, 10.1074/jbc.M002945200
Submitted on April 6, 2000
Revised on August 8, 2000
Accepted on September 19, 2000
Expression of the
5 integrin subunit gene promoter is positively regulated by the extracellular matrix component fibronectin through the transcription factor Sp1 in corneal epithelial cells in vitro
5
1. In this work, we determined whether the activity directed by the 5 gene promoter can be modulated by FN in rabbit corneal epithelial cells (RCEC). The activity driven by CAT/5 promoter bearing plasmids was drastically increased when transfected into RCEC grown on FN-coated culture dishes. The promoter sequence mediating FN-responsiveness was shown to bear a perfect inverted repeat that we designated the Fibronectin Responsive Element (FRE). Analyses in EMSA provided evidence that Sp1 is the predominant transcription factor binding the FRE. Its DNA binding affinity was found to be increased when RCEC are grown on FN-coated dishes. The addition of the MEK kinase inhibitor PD98059 abolished FN-responsiveness suggesting that alteration in the state of phosphorylation of Sp1 likely accounts for its increased binding to the 5 FRE. The FRE also proved sufficient to confer FN-responsiveness to an otherwise unresponsive heterologous promoter. However, site-directed mutagenesis indicated that only the 3? half site of the FRE was required to direct FN-responsiveness. Collectively, binding of FN to its 51 integrin activates a signal transduction pathway that results in the transcriptional activation of the 5 gene likely through altering the phosphorylation state of Sp1.
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