Research article

DNA demethylation-dependent enhancement of toll-like receptor-2 gene expression in cystic fibrosis epithelial cells involves SP1-activated transcription

Takashi Furuta^{* 1} , Tsuyoshi Shuto^{* 1} , Shogo Shimasaki¹ , Yuko Ohira¹ , Mary Ann Suico¹ , Dieter C Gruenert^2,3,4 and Hirofumi Kai¹

¹Department of Molecular Medicine, Graduate School of Pharmaceutical Science, Global COE "Cell Fate Regulation Research and Education Unit", Kumamoto University, Kumamoto 862-0973, Japan

²California Pacific Medical Center Research Institute, 475 Brannan St, Suite 220, San Francisco, CA 94115, USA

³Department of Laboratory Medicine, University of California, San Francisco, San Francisco, CA 94143, USA

⁴Department of Medicine, University of Vermont, Burlington, VT 05405, USA

author email corresponding author email* Contributed equally

BMC Molecular Biology 2008, 9:39doi:10.1186/1471-2199-9-39

Published:	21 April 2008

Abstract

Background

The clinical course of cystic fibrosis (CF) is characterized by recurrent pulmonary infections and chronic inflammation. We have recently shown that decreased methylation of the toll-like receptor-2 (TLR2) promoter leads to an apparent CF-related up-regulation of TLR2. This up-regulation could be responsible, in part, for the CF-associated enhanced proinflammatory responses to various bacterial products in epithelial cells. However, the molecular mechanisms underlying DNA hypomethylation-dependent enhancement of TLR2 expression in CF cells remain unknown.

Results

The present study indicates that there is a specific CpG region (CpG#18-20), adjacent to the SP1 binding site that is significantly hypomethylated in several CF epithelial cell lines. These CpGs encompass a minimal promoter region required for basal TLR2 expression, and suggests that CpG#18-20 methylation regulates TLR2 expression in epithelial cells. Furthermore, reporter gene analysis indicated that the SP1 binding site is involved in the methylation-dependent regulation of the TLR2 promoter. Inhibition of SP1 with mithramycin A decreased TLR2 expression in both CF and 5-azacytidine-treated non-CF epithelial cells. Moreover, even though SP1 binding was not affected by CpG methylation, SP1-dependent transcription was abolished by CpG methylation.

Conclusion

This report implicates SP1 as a critical component of DNA demethylation-dependent up-regulation of TLR2 expression in CF epithelial cells.