Molecular Mechanism for Anencephaly

细胞红蛋白（Cygb）最初被发现是一种细胞质蛋白，在纤维化大鼠肝脏活化的星状细胞中显著上调。尽管该蛋白的生物学功能尚不清楚，但该蛋白可能通过其血红素结合域参与细胞内氧浓度的调节。最近，我们培育了带有Cygb基因的转基因青蛙来研究Cygb在体内的功能。利用Kroll和Amaya的方法，构建了一种携带Cygband EGFP cdna的转基因，与精子核混合，并将其微注射到未受精的爪蟾卵中。荧光显微镜下观察EGFP基因的转基因情况。RT-PCR证实转基因胚胎中Cygb mRNA表达水平较高。所得到的转基因胚胎在神经期显示出神经管的高频率暴露。在尾芽期，大多数胚头结构异常，A-P轴缩短。这些表型与人类无脑畸形相似。基于这些结果，我们现在假设Cygb可能参与了非洲爪蟾胚胎早期发育的头部组织。另一方面，我们还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强烈表明Cygb和Mb之间具有相似的功能，因此我们假设肌肉细胞中的Cygb弥补了Mb在两栖动物中氧的储存和细胞内运输中的缺乏。然而，有趣的是，Cygb在心脏中只定位于细胞核，这表明这种珠蛋白在基因调控中起作用。

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.