Miyake Akimitsu, Saito Taiju, Kashiwagi Noboru, Ando Daisuke, Yamamoto Akitsugu, Suzuki Tohru, Nakatsuji Norio, Nakatsuji Takako
Department of Marine Biology, Graduate School of Marine Science and Technology, Tokai University, Japan.
Int J Dev Biol. 2006;50(7):619-25. doi: 10.1387/ijdb.062172am.
The vasa genes are expressed in the germ cell lineage in many organisms, but their expression patterns show large variations. Recent studies suggest that vasa transcripts are involved in germ cell lineage development. In this paper, we isolated the vasa cDNA clone from a teleost, shiro-uo, Leucopsarion petersii and examined its expression pattern during embryogenesis. Then, we examined the functional significance of vasa mRNA during the formation of primordial germ cells (PGCs). The amino acid sequence of shiro-uo VASA is 61.1% identical to that of zebrafish. In whole-mount in situ hybridization, vasa transcripts appeared at the 4- and 8-cell stages as four spots at both ends of two cleavage planes between the lower tier of blastomeres and the yolk cell mass. At the 16-cell stage, eight spots were observed. After the blastula stage, shiro-uo vasa transcripts showed similar localization as in the zebrafish. Ultrastructural analysis of 4-cell stage embryos revealed the presence of a subcellular organelle that resembled 'nuage' in the germ cell lineage observed in the embryos of various organisms. We carried out micromanipulation of 4- or 8-cell stage embryos to remove the vasa mRNA-containing spots and then measured the number of the vasa-expressing PGCs in the genital ridge of the manipulated embryos. The numbers decreased when all of the four spots were removed, indicating that the vasa-containing spots at early cleavage stages have important functions in the development of PGCs.
瓦萨基因在许多生物的生殖细胞谱系中表达，但其表达模式存在很大差异。最近的研究表明，瓦萨转录本参与生殖细胞谱系的发育。在本文中，我们从一种硬骨鱼——日本七鳃鳗（Leucopsarion petersii）中分离出瓦萨cDNA克隆，并研究了其在胚胎发育过程中的表达模式。然后，我们研究了瓦萨mRNA在原始生殖细胞（PGC）形成过程中的功能意义。日本七鳃鳗VASA的氨基酸序列与斑马鱼的氨基酸序列有61.1%的同源性。在全胚胎原位杂交中，瓦萨转录本在4细胞和8细胞阶段出现在两个卵裂平面两端的四个点上，位于卵裂球下层和卵黄细胞团之间。在16细胞阶段，观察到八个点。囊胚期后，日本七鳃鳗瓦萨转录本的定位与斑马鱼相似。对4细胞期胚胎的超微结构分析显示，存在一种亚细胞器，类似于在各种生物胚胎中观察到的生殖细胞谱系中的“生殖质”。我们对4细胞或8细胞期胚胎进行显微操作，去除含有瓦萨mRNA的点，然后测量操作后胚胎生殖嵴中表达瓦萨的PGC数量。当所有四个点都被去除时，数量减少，这表明早期卵裂阶段含有瓦萨的点在PGC的发育中具有重要功能。
