Department of Life Science, Tokyo Institute of Technology, Nagatsuta, Midori-ku, Yokohama 226, Japan, , , , , , JP.
Dev Genes Evol. 1996 Dec;206(5):297-304. doi: 10.1007/s004270050056.
Cytoplasmic determinants that specify the fate of endoderm, muscle and epidermis cells are known to be localized in specific areas of fertilized eggs of ascidians. The presence of such cytoplasmic determinants in unfertilized eggs was demonstrated in previous studies, but no information has yet been proved about their distribution. To investigate the distribution of cytoplasmic determinants in unfertilized eggs, we devised a method for distinguishing the polarity of unfertilized eggs using vital staining and we performed cytoplasmic-transfer experiments by fusing blastomeres and cytoplasmic fragments from various identified regions of unfertilized eggs. Cytoplasmic fragments, that contained cortical and subcortical material, from five different positions along the animal-vegetal axis were prepared, and they were fused with a4.2 (presumptive-epidermis) or A4.1 (non-epidermis) blastomeres. The ectopic development of endoderm, muscle and epidermis cells that was promoted by the transplanted cytoplasm was assessed by examining the expression of alkaline phosphatase (ALP), myosin and epidermis-specific antigen, respectively. Differentiation of endoderm and muscle was observed at higher frequencies as cytoplasmic fragments closer to the vegetal pole were transplanted. Conversely, formation of epidermis was observed at higher frequencies as cytoplasmic fragments closer to the animal pole were transplanted. The results suggest that, in cortical and subcortical regions of unfertilized ascidian eggs, endoderm and muscle determinants are widely distributed along a gradient, with maximum activity at the vegetal pole, whilst epidermis determinants are also distributed along a gradient but with maximum activity at the animal pole.
已知海鞘受精卵的细胞质决定因素决定内胚层、肌肉和表皮细胞的命运，这些决定因素被定位在受精卵的特定区域。之前的研究表明，未受精卵中也存在这种细胞质决定因素，但尚未证明其分布情况。为了研究未受精卵中细胞质决定因素的分布，我们设计了一种使用活体染色来区分未受精卵极性的方法，并通过融合来自未受精卵不同鉴定区域的胚胎细胞和细胞质片段来进行细胞质转移实验。我们从动物-植物轴的五个不同位置制备了包含皮质和皮层下材料的细胞质片段，并将其与 4.2（假定表皮）或 A4.1（非表皮）胚胎细胞融合。通过分别检测碱性磷酸酶 (ALP)、肌球蛋白和表皮特异性抗原的表达，评估了移植细胞质促进的内胚层、肌肉和表皮细胞的异位发育情况。随着移植的细胞质更接近植物极，内胚层和肌肉的分化频率更高；相反，随着移植的细胞质更接近动物极，表皮的形成频率更高。这些结果表明，在未受精卵的皮质和皮层下区域，内胚层和肌肉决定因素沿着一个梯度广泛分布，在植物极处活性最高，而表皮决定因素也沿着一个梯度分布，但在动物极处活性最高。
