Molecular Mechanisms of Muscle Cell Differentiation in Embryos Ascidian

研究了海鞘胚胎肌细胞分化的分子机制，认为海鞘胚胎具有自主分化的潜力。首先，我们分离了肌球蛋白重链和肌型肌动蛋白的CDNA克隆。这些肌肉特异性基因的表达大约在原肠胚形成时期开始，并且仅限于分化自体细胞。海鞘胚胎合成几种不同的肌动蛋白mrna。其中一个克隆HrcMA4包含一个1137 bp的开放阅读框和一个100 bp的非编码区，后面是一个poly(a)尾巴。然后，我们分离出含有HrMA4基因的基因组片段。该基因的表征表明HrMA4被两个内含子打断。有几个潜在的调控元件的准确表达的基因。此外，HrMA4和其他几个肌肉肌动蛋白基因形成了一个簇。其次，我们尝试分离MyoDl和myogenic等肌生成因子的基因。到目前为止，我们已经用PCR方法获得了一个候选的dna片段。最后，我们通过产生几种单克隆抗体，分析了被认为含有致肌因子的肌浆的大分子。其中一种抗体被注射到受精卵后会阻碍肌肉的发育。然而，抗原多肽的表征表明它与细胞骨架蛋白有关。

Molecular mechanisms of muscle cell differentiation of ascidian embryos have been investigated with respect to an extreme potential of autonomous differentiation of the cells.First, we have isolated CDNA clones for myosin-heavy chain and museletype actins. Expression of these muscle-specific genes beans around the time of gastrulation, and is restricted to differentiating sele cells. Ascidian embryos synthesize several different actin mRNAs. One of the clones, HrcMA4 contains an open reading frame of 1137 bp and a 100 bp noncoding region followed by a poly(A)tail. Then, we have isolated genomic fragments that contain HrMA4 gene. Characterization of the gene shows that HrMA4 is interrupted by two introns. There are several potential regulatory elements for accurate expression of the gene. In addition, it becomes clear that HrMA4 and several other muscle-actin genes form a cluster.Second, we have tried to isolate genes for myogenic factors such as MyoDl and myogenic. S6 far, we have obtained by PCR method a candidate DNA-fragment for the genes.Finally, we have analyzed macromolecules of the myoplasm, which is thought to contain the myogenic factors, by producing several monoclonal antibodies. One of the antibodies blocks muscle development when it is injected into fertilized eggs. Characterization of the antigenic polypeptide, however, has shown that it is related to cytoskeletal proteins.