|Budget Amount *help
¥6,400,000 (Direct Cost : ¥6,400,000)
Fiscal Year 1991 : ¥1,800,000 (Direct Cost : ¥1,800,000)
Fiscal Year 1990 : ¥1,800,000 (Direct Cost : ¥1,800,000)
Fiscal Year 1989 : ¥2,800,000 (Direct Cost : ¥2,800,000)
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.