Molecular mechanism of morphogenetic movements in amphibian embryos

原肠发育是动物发育过程中最具活力的形态发生运动，启动了动物的身体发育计划。为了分析原肠形成的分子机制，我们在非洲爪哇中通过表达筛选获得了编码POZ/锌指蛋白的新的库珀尔样转录因子香菇(CpG)。CpG的过表达导致原肠形成受到干扰。比较对照胚胎和注射CPG的胚胎11.5期的组成蛋白，发现注射CPG的胚胎中61 kDa的蛋白质点发生了基本的移位，说明注射CPG可以使61 kDa的蛋白质去磷酸化。61 kDa蛋白的部分氨基酸序列(23mer)与非洲爪哇STI1蛋白的氨基酸序列完全一致，STI1蛋白是应激诱导的磷酸化蛋白(STI1)、Hsp70/Hsp90组织蛋白(Hop)或转化敏感蛋白的同源物。STI1/Hop结合Hsp70和Hsp90，调节Hsp90分子伴侣的功能，Hsp90分子伴侣已知与细胞骨架结合。因此，CpG对原肠形成的抑制作用是通过XSTI1的去磷酸化来实现的。这些结果表明，Hsp90分子伴侣参与了形态发生运动的调控。我们还克隆了一个新的基因Xoom，它编码一种跨膜蛋白。将Xoom反义RNA注射到非洲爪哇胚胎中，降低了Xoom蛋白的表达，并导致原肠发育缺陷，但不影响中胚层标记基因的退化和表达。不同实验条件下细胞形态的比较表明，细胞扩散的抑制特异性地发生在反义RNA注射的胚胎的外胚层。细胞学检查显示，注射反义RNA的胚胎外胚层细胞中的F-肌动蛋白脱失。这些结果表明，Xoom通过对肌动蛋白细丝组织的某些调节，在外胚层细胞的外胚层运动中起着重要作用。

Gastrulation is the most dynamic morphogenetic movement and initiates the body plan in animal development. In order to analyze the molecular mechanism of gastrulation, we have isolated a novel kuppel-like transcription factor, champignon (cpg), that encodes POZ/Zinc finger protein by expression screening in Xenopus laevis. Overexpression of cpg resulted in interference with gastrulation. Comparison of the constituent proteins from control embryos and cpg-injected embryos at stage 11.5 revealed that a 61 kDa protein spot was shifted basically in cpg-injected embryos suggesting that the 61 kDa protein was dephosphorylated by cpg-injection. Partial amino acid sequence (23 mer) of the 61 kDa protein accords 100 % with that of putative Xenopus STI1 protein which is a homolog of stress-induced phosphoprptein (STI1), Hsp70/Hsp90-organizing protein (Hop), or transformation-sensitive protein. The STI1/Hop combines Hsp70 with Hsp90 and modulates the function of Hsp90 chaperone which is known to bind to cytoskeletons. Thus the inhibitory role of cpg on gastrulation is mediated by dephosphorylation of XSTI1. These results suggest that the Hsp90 chaperone is involved in the control of morphogenetic movements.We have also cloned a novel gene, Xoom, which encodes a transmembrane protein. Injection of Xoom antisense RNA into Xenopus embryos reduced the Xoom protein and caused gastrulation defects without any influence on the involution and expression of mesodermal marker genes. Comparison of cell shape among various experimental conditions showed that inhibition of cell spreading occurs specifically in the outer ectodermal layer of the antisense RNA-injected embryo. Cytological examination indicated disorgatrization of F-actin in the ectodermal cells of the antisense RNA-injected embryo. These results suggest that Xoom plays an important role in the epibolic movement of ectodermal cells through some regulation of actin filament organizaiion.