Various cellular and molecular events are involved in palatogenesis, including apoptosis, epithelial- mesenchymal transition (EMT), cell proliferation, and cell migration. Smad2 and Snail, which are well-known key mediators of the transforming growth factor beta (Tgf-β) pathway, play a crucial role in the regulation of palate development. Regulatory effects of microRNA 200b (miR- 200b) on Smad2 and Snail in palatogenesis have not yet been elucidated. The aim of this study is to determine the relationship between palate development regulators miR-200b and Tgf-β-mediated genes. Expression of miR-200b, E-cadherin, Smad2, and Snail was detected in the mesenchyme of the mouse palate, while miR-200b was expressed in the medial edge epithelium (MEE) and palatal mesenchyme. After the contact of palatal shelves, miR- 200b was no longer expressed in the mesenchyme around the fusion region. The binding activity of miR-200b to both Smad2 and Snail was examined using a luciferase assay. MiR-200b directly targeted Smad2 and Snail at both cellular and molecular levels. The function of miR-200b was determined by overexpression via a lentiviral vector in the palatal shelves. Ectopic expression of miR-200b resulted in suppression of these Tgf-β-mediated regulators and changes of apoptosis and cell proliferation in the palatal fusion region. These results suggest that miR-200b plays a crucial role in regulating the Smad2, Snail, and in apoptosis during palatogenesis by acting as a direct non-coding, influencing factor. Furthermore, the molecular interactions between miR-200b and Tgf-β signaling are important for proper palatogenesis and especially for palate fusion. Elucidating the mechanism of palatogenesis may aid the design of effective gene-based therapies for the treatment of congenital cleft palate.
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Acknowledgments We are grateful to Dr. Cho, Eui-Sic for critical reading of this manuscript. We also thank Dr. Kim, Jong-Soo for providing technical advice to perform luciferase assay. This work was supported by the National Research Foundation of Korea(NRF) grant funded by the Korea government(MEST) (No. 2011-0015661).
All Science Journal Classification (ASJC) codes
- Molecular Biology
- Medical Laboratory Technology
- Cell Biology