Analysis of the molecular mechanism of vertebral column formation by a cross-species genome engineering approach between mice and zebrafish

2014 Fiscal Year Final Research Report

• Project/Area Number: 24300154
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Partial Multi-year Fund
• Section: 一般
• Research Field: Laboratory animal science
• Research Institution: Osaka University
• Principal Investigator: KOKUBU Chikara 大阪大学, 医学(系)研究科(研究院), 准教授 (70379238)
• Co-Investigator(Kenkyū-buntansha): SHUKUNAMI Chisa 広島大学, 大学院医歯薬保健学研究科, 教授 (60303905) ; HIRAKI Yuji 京都大学, 再生医科学研究所, 教授 (40144498)
• Project Period (FY): 2012-04-01 – 2015-03-31
• Keywords: 比較ゲノム / 軟骨形成 / 発生・分化 / 発現制御

Outline of Final Research Achievements

The structure and function of the Pax1 genomic region, which is essential for vertebral column formation, was compared between the mouse and zebrafish by a cross-species genome engineering approach. Transgenic reporter assays revealed that a mouse sclerotome (vertebral anlage)-specific enhancer mPf1 (0.8 kb) did not show enhancer activity in zebrafish embryos, while its zebrafish ortholog zPf1 (0.7 kb) drove stronger sclerotomal expression in mice than the isogenic mPf1. Another sclerotomal enhancer mXe1 (1.7 kb), which is evolutionary conserved in terrestrial vertebrates but not in aquatic vertebrates, drove robust reporter expression in the zebrafish sclerotome. Intriguingly, forced expression of Pax1 driven by the Xe1 enhancer transgene inhibited cartilage formation in zebrafish. The inhibitory role of Pax1 against cartilage formation was further analyzed in detail by forced expression experiments in chicken embryos and primary cultured cells.

Free Research Field

実験動物学