Abnormal muscle development in the Mesp quadruple mutant. (A-C) Immunostaining of Engrailed (4D9) and Prox1 in the wild type (A) and Mesp quadruple mutant (B). Embryos were fixed at 24hpf. Graph showing the numbers of MPs, identified as the 4D9 and Prox1 double-positive cells, in each somite in the wild-type and Mesp quadruple mutant embryos (C). The number of MPs at 24hpf was counted in the 5-6 somites located in the most posterior part of the trunk region. A total of 16 somites were counted from 3 wild-type embryos and 23 somites from 4 mutant embryos. (D,E) Expression of smyhc in the wild-type and Mesp quadruple mutant embryos at 24hpf. After whole-mount in situ hybridization, the embryos were transversely sectioned at the middle level of the yolk tube. (F,G) Expression of cxcl12a in the wild-type and Mesp quadruple mutant embryos at 24hpf (100%, n=14). (H,I) Expression of pax7 in wild-type and Mesp quadruple mutant embryos in the most posterior region of yolk tube. Embryos were fixed at the 20-somite stage. Scale bar: 50µm. (J-L) Cell-autonomous function of Mesp in superficial horizontal myoseptum formation. (J) Schema for the transplantation experiment. The wild-type cells were transplanted at the marginal zone of Mesp quadruple mutant embryos at the blastula stage. Although the expression of cxcl12a is rescued in the transplanted embryo at 24hpf (K,L), this expression is restricted to the donor cells (K′′,L′′). K and L show the rescued phenotype in different embryos. Although the rescued expression of cxcl12a was observed in a total of 42 cells in 10 embryos, this rescue occurred only in the donor cells in all cases.
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