Functional analysis of germ line cells using gene manipulated mouse

我们已经开发了一种简单而高效的系统，在植入阶段之前以非侵入性的方式分离男性和女性胚胎。当用该系统制备的雄卵和雌卵聚集在一起产生性嵌合体时，该系统的优点是可以通过观察X染色体上转基因整合的绿色荧光蛋白基因来容易地指出构成器官的单个细胞的性别。通过对如此制备的性嵌合体的生殖器官的分析，我们在表型男性睾丸中发现了许多女性起源的精原细胞。这些基因的基因组印迹模式被发现，尽管它们的原始性别是女性的，但它们的基因组印记模式是男性类型的。与Nakatsuji教授的团队合作，还发现这些细胞在发育的早期进入减数分裂，但另一方面，一些起源于雌性的细胞在有性嵌合体动物的睾丸中变成了卵状的巨大细胞，这就是…更多的雌性细胞发育成雄性类型的精原细胞。通过产生这些性别嵌合体小鼠，阐明了生殖细胞性分化的复杂性，同时也为分析生殖系细胞的雌雄分化机制提供了工具。Pig-a是一个X连锁基因，是糖基磷脂酰肌醇(GPI)锚定生物合成的关键成分，该基因的缺失导致数百种GPI锚定蛋白的缺失。Kondoh博士已经培育了一种转基因小鼠，它以GPI锚定的形式表达增强的绿色荧光蛋白，并对GPI锚定阳性细胞进行了原位监测。结果表明，血管紧张素转换酶具有二肽基肽酶以外的磷脂酶活性。提示睾丸血管紧张素转换酶是必需的，因为它可以从精子表面释放GPI锚定蛋白，以形成可育精子。结果表明，有必要重新评价血管紧张素转换酶参与生殖生物学的必要性。较少

We have developed an easy and very efficient system to separate male and female embryos before implantation stage with non invasive manner. When male and female eggs prepared by this system were aggregated to produce sex chimera, the advantage of this system is the easiness of pointing out the sex of individual cells that form organs by observing the transgenically incorporated green fluorescent protein gene in X chromosome. By analyzing the genital organs in thus prepared sex chimeras, we have found many spermatogonia like cells of female origin in phenotypically male testes. The pattern of the genomic imprinting of these genes was found out to be male type in spite of their original sex was female.With the collaboration to Prof. Nakatsuji's group, it was also found out that these "female spermatogonia" cells entered meiosis in early stages of development However, on the other hand, some cells originated from female became huge cells like eggs in the testis of sex chimera animals, whi … More le the majority of the female cells developed into male type spermatogonia. The complexity of the sex differentiation of germ cells was elucidated by producing these sex chimera mice and simultaneously provided a tool to analyze the mechanism of male-female differentiation in germ line cells.Pig-a, an X-linked gene, is a key component of glycosylphosphatidylinositol (GPI) anchor biosynthesis based on the fact that lack of this gene causes deficiencies of hundreds of GPI-anchored proteins. Dr. Kondoh has produced a transgenic mouse line that expressed enhanced green fluorescent protein in GPI-anchored form and monitored GPI-anchor-positive cells in situ. It was found out that ACE has phospholipase activity other than the dipeptidyl-peptidase. It was suggested that testicular ACE is indispensable because it may release GPI anchored proteins from sperm surface to make fertile sperm. The results indicated that the necessity of re-evaluation of the involvement of ACE in reproductive biology. Less