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Molecular Mechanism for Anencephaly

无脑畸形的分子机制

基本信息

批准号：
17591142
负责人：
OBARA Masanobu
金额：
$ 2.11万
依托单位：
HIROSHIMA UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-17591142/
关键词：
cytoglobin heme-protein transgenic forg anencephaly neural tube disclosure molecular embryogenesis in vitro fertilization head organizer genes

项目摘要

Cytoglobin (Cygb) is originally found as a cytoplasmic protein that is dramatically upregulated in activated stellate cells in fibrotic rat liver. Although the biological function of this protein still remains to be unclear, the protein may be involved in the regulation of intracellular oxygen concentrations via its heme-binding domain.Recently, we generated transgenic frogs bearing Cygb gene to investigate the function of Cygb in vivo. A transgene bearing cDNAs of Cygband EGFP was constructed, mixed with sperm nuclei, and microinjected into unfertilized Xenopus eggs by the method developed by Kroll and Amaya. The transgenesis of EGFP gene was monitored through a fluorescent microscope. A high level of mRNA expression of Cygb in transgenic embryos was confirmed by RT-PCR. The resulting transgenic embryos at neural stage showed the disclosure of neural tubes at a high frequency. At the tail bud stage, most embryos showed anomaly in head structures as well as the shortening of the A-P axis. These phenotypes were similar to those of human anencephaly. Based on these results, we now hypothesize that Cygb may be involved in the head organization during early development of Xenopus leavis embryos.On the other hand, we also conducted molecular evolutional study with Xenopus leavis and Iguana iguana. The study on Cygb gene expression and protein distribution strongly indicates a similar function between Cygb and Mb, thus we hypothesize that Cygb in muscle cells compensates for the lack of Mb for the storage and the intracellular transportation of oxygen in the Amphibia. However, interestingly, Cygb in the heart was exclusively localized in nuclei, which suggests a role of this globin in gene regulation.

细胞珠蛋白（Cygb）最初被发现是一种细胞质蛋白，在纤维化大鼠肝脏的活化星状细胞中显著上调。虽然该蛋白的生物学功能尚不清楚，但它可能通过血红素结合结构域参与细胞内氧浓度的调节。构建带有Cygband EGFP cDNA的转基因，与精子核混合，并通过Kroll和Amaya开发的方法显微注射到未受精的爪蟾卵中。通过荧光显微镜监测EGFP基因的转化。RT-PCR结果表明，Cygb基因在转基因胚中有高水平的表达。神经期转基因胚胎中神经管的出现频率较高。在尾芽期，大多数胚胎出现头部结构异常，A-P轴缩短。这些表型与人类无脑畸形相似。基于这些结果，我们推测Cygb可能参与了非洲爪蟾胚胎早期发育过程中的头部组织。另一方面，我们也对非洲爪蟾和鬣蜥进行了分子进化研究。对Cygb基因表达和蛋白分布的研究表明，Cygb和Mb具有相似的功能，因此我们推测Cygb在两栖类动物肌肉细胞中可以弥补Mb的缺乏，用于氧的储存和胞内运输。然而，有趣的是，心脏中的Cygb仅定位于细胞核中，这表明这种球蛋白在基因调控中的作用。