- 作者：Youwen Yang ; Hans Michael Haitchi ; Julie Cakebread ; David Sammut ; Anna Harvey ; Robert M. Powell ; John W. Holloway ; Peter Howarth ; Stephen T. Holgate ; Donna E. Davies
- 关键词：Promoter ; CpG island ; methylation ; expression ; ADAM33 ; epithelial-mesenchymal transition
- 刊名：Journal of Allergy and Clinical Immunology
- 出版年：2008
- 出版时间：June 2008
- 年：2008
- 卷：121
- 期：6
- 页码：1393-1399.e14
- 全文大小：3313 K
Background
A disintegrin and metalloprotease 33 (ADAM33) polymorphism is strongly associated with asthma and bronchial hyperresponsiveness. Although considered to be a mesenchymal cell–specific gene, recent reports have suggested epithelial expression of ADAM33 in patients with severe asthma.Objectives
Because dysregulated expression of ADAM33 can contribute to disease pathogenesis, we characterized the mechanism or mechanisms that control its transcription and investigated ADAM33 expression in bronchial biopsy specimens and brushings from healthy and asthmatic subjects.
Methods
The ADAM33 promoter and CpG island methylation were analyzed by using bioinformatics, luciferase reporters, and bisulfite sequencing of genomic DNA. Epithelial-mesenchymal transition was induced by using TGF-β1. ADAM33 mRNA was scrutinized in bronchial biopsy specimens and brushings by using reverse transcriptase–quantitative polymerase chain reaction, melt-curve analysis, and direct sequencing.
Results
The predicted ADAM33 promoter (−550 to +87) had promoter transcriptional activity. Bisulfite sequencing showed that the predicted promoter CpG island (−362 to +80) was hypermethylated in epithelial cells but hypomethylated in ADAM33-expressing fibroblasts. Treatment of epithelial cells with 5-aza-deoxycytidine caused demethylation of the CpG island and induced ADAM33 expression. In contrast, phenotypic transformation of epithelial cells through a TGF-β–induced epithelial-mesenchymal transition was insufficient to induce ADAM33 expression. ADAM33 mRNA was confirmed in bronchial biopsy specimens, but no validated signal was detected in bronchial brushings from healthy or asthmatic subjects.
Conclusion
The ADAM33 gene contains a regulatory CpG island within its promoter, the methylation status of which tightly controls its expression in a cell type–specific manner. ADAM33 repression is a stable feature of airway epithelial cells, irrespective of disease.