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Endothelial Cell–Specific Expression of Roundabout 4 Is Regulated by Differential DNA Methylation of the Proximal Promoter
| Content Provider | Scilit |
|---|---|
| Author | Okada, Yoshiaki Funahashi, Nobuaki Tanaka, Toru Nishiyama, Yuji Yuan, Lei Shirakura, Keisuke Turjman, Alexis S. Kano, Yoshihiro Naruse, Hiroki Suzuki, Ayano Sakai, Miki Zhixia, Jiang Kitajima, Kenji Ishimoto, Kenji Hino, Nobumasa Kondoh, Masuo Mukai, Yohei Nakagawa, Shinsaku García-Cardeña, Guillermo Aird, William C. Doi, Takefumi |
| Copyright Year | 2014 |
| Description | Journal: Arteriosclerosis, thrombosis, and vascular biology Objective— The molecular basis of endothelial cell (EC)–specific gene expression is poorly understood. Roundabout 4 (Robo4) is expressed exclusively in ECs. We previously reported that the 3-kb 5′-flanking region of the human Robo4 gene contains information for lineage-specific expression in the ECs. Our studies implicated a critical role for GA-binding protein and specificity protein 1 (SP1) in mediating overall expression levels. However, these transcription factors are also expressed in non-ECs. In this study, we tested the hypothesis that epigenetic mechanisms contribute to EC-specific Robo4 gene expression. Methods and Results— Bisulfite sequencing analysis indicated that the proximal promoter of Robo4 is methylated in non-ECs but not in ECs. Treatment with the DNA methyltransferase inhibitor 5-aza-2′-deoxycytidine increased Robo4 gene expression in non-ECs but not in ECs. Proximal promoter methylation significantly decreased the promoter activity in ECs. Electrophoretic mobility shift assays showed that DNA methylation of the proximal promoter inhibited SP1 binding to the −42 SP1 site. In DNase hypersensitivity assays, chromatin condensation of the Robo4 promoter was observed in some but not all nonexpressing cell types. In Hprt (hypoxanthine phosphoribosyltransferase)-targeted mice, a 0.3-kb proximal promoter directed cell-type–specific expression in the endothelium. Bisulfite sequencing analysis using embryonic stem cell–derived mesodermal cells and ECs indicated that the EC-specific methylation pattern of the promoter is determined by demethylation during differentiation and that binding of GA-binding protein and SP1 to the proximal promoter is not essential for demethylation. Conclusions— The EC-specific DNA methylation pattern of the Robo4 proximal promoter is determined during cell differentiation and contributes to regulation of EC-specific Robo4 gene expression. |
| Related Links | https://www.ahajournals.org/doi/pdf/10.1161/ATVBAHA.114.303818 https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5378492/pdf https://www.ahajournals.org/doi/reader/10.1161/ATVBAHA.114.303818 |
| Ending Page | 1538 |
| Page Count | 8 |
| Starting Page | 1531 |
| ISSN | 10795642 |
| e-ISSN | 15244636 |
| DOI | 10.1161/atvbaha.114.303818 |
| Journal | Arteriosclerosis, thrombosis, and vascular biology |
| Issue Number | 7 |
| Volume Number | 34 |
| Language | English |
| Publisher | Ovid Technologies (Wolters Kluwer Health) |
| Publisher Date | 2014-07-01 |
| Access Restriction | Open |
| Subject Keyword | Journal: Arteriosclerosis, thrombosis, and vascular biology Endocrinology and Metabolism Endothelial Cells Dna Methylation |
| Content Type | Text |
| Resource Type | Article |
| Subject | Cardiology and Cardiovascular Medicine |
