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Analysis on the function of a protein encoded by a thyroid hormone responsive gene, ZAKI-4, in order to elucidate the molecular mechanizm of thyroid hormone action on brain development

分析甲状腺激素反应基因ZAKI-4编码蛋白的功能，阐明甲状腺激素对大脑发育作用的分子机制

基本信息

批准号：
11470225
负责人：
MURATA Yoshiharu
金额：
$ 7.42万
依托单位：
Nagoya University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

项目摘要

It is well known that thyroid hormone (T_3) plays a pivotal role in brain development, however, its molecular mechanism remains to be elucidated. We previously identified a thyroid hormone-responsive gene, ZAKI-4. ZAKI-4 encodes a protein (ZAKI-4 protein) with a molecular mass of 26kd. Recently, it has been reported that ZAKI-4 belongs to a family protein that inhibits calcineurin activity in vivo. Sicne calcineurin is known to play important roles in brain development and function, we hypothesized, that ZAKI-4 protein is one of key factors to mediate T_3 action during brain development and designed the present research projects to elucidate the in vivo function of ZAKI-4 protein. For this purpose we tried to produce mice in which the expression of ZAKI-4 is completely eliminated. As the first step to knockout ZAKI-4 gene, we determined the genomic organization of mouse ZAKI-4 gene and a targeting vector was constructed. We are currently carrying out homologous recombination in embryon … More ic stem (ES) cells derived from a mouse train (129/Sv). In the present study, the spatial and ontogenic expression of ZAKI-4 mRNA and the effect of hypothyroidism on the expression were also examined in brain of rodents. In cerebral cortex, the expression of ZAKI-4 mRNA was observed as early as embryonic day 18 and the expression increased during the brain development. This increase was significantly blunted tn hypothyroid rats. On the other hand the expression was constant and was not affected by hypothyroidism in cerebellum. In situ hybridization revealed that ZAKI-4 mRNA was widely expressed in the entire brain. It was noted that ZAKI-4 was highly expressed in the area such as olfactory bulb, hipocampus, cerebral cortex and cerebellum, where calcineurin is abundt. These results demonstrated that the expression of ZAKI-4 mRNA was indeed regulated by thyroid status in vivo and would suggest that thyroid hormone excerts its biological function during brain development at least in part by regulating ZAKI-4 expression and thus by controling calcineruin activity. Less

众所周知，甲状腺激素（T_3）在大脑发育中发挥着关键作用，但其分子机制仍有待阐明。我们之前发现了一种甲状腺激素反应基因 ZAKI-4。 ZAKI-4编码一种分子量为26kd的蛋白质（ZAKI-4蛋白）。最近有报道称ZAKI-4属于抑制体内钙调神经磷酸酶活性的家族蛋白。众所周知，钙调神经磷酸酶在大脑发育和功能中发挥着重要作用，我们假设ZAKI-4蛋白是大脑发育过程中介导T_3作用的关键因素之一，并设计了本研究项目来阐明ZAKI-4蛋白的体内功能。为此，我们尝试培育出 ZAKI-4 表达完全消除的小鼠。作为敲除ZAKI-4基因的第一步，我们确定了小鼠ZAKI-4基因的基因组结构并构建了靶向载体。我们目前正在小鼠胚胎干 (ES) 细胞 (129/Sv) 中进行同源重组。在本研究中，还检测了啮齿类动物大脑中 ZAKI-4 mRNA 的空间和个体表达以及甲状腺功能减退对其表达的影响。在大脑皮层中，早在胚胎第18天就观察到ZAKI-4 mRNA的表达，并且在大脑发育过程中表达增加。这种增加在甲状腺功能减退大鼠中显着减弱。另一方面，小脑中的表达是恒定的并且不受甲状腺功能减退症的影响。原位杂交显示ZAKI-4 mRNA在整个大脑中广泛表达。值得注意的是，ZAKI-4在嗅球、海马、大脑皮层和小脑等富含钙调神经磷酸酶的区域高表达。这些结果表明，ZAKI-4 mRNA 的表达确实受到体内甲状腺状态的调节，并表明甲状腺激素在大脑发育过程中至少部分通过调节 ZAKI-4 表达从而控制钙调蛋白活性来发挥其生物学功能。较少的