Figure 5

Later development of M. stichopi embryos including muscle formation. Nuclear labeling with Propidium Iodide (magenta), muscle fibers with BODIPY FL-Phallacidin (green) and anti-tyrosinated tubulin (yellow). Left row Maximum Intensity Projections, right row optical sections. A) Embryo two weeks after fertilization with ~180 cells labeled with BODIPY FL-phallacidin. A’) shows the inner cell mass (encircled by dotted line) in an optical section of the embryo shown in A). B) Embryo with ~500 cells three weeks after fertilization. B’) shows the nuclei close to the cell membrane of each cell. C) 4-week old embryo composed out of approximately 700 cells. C’) Optical section of C), with actin filaments visible that indicate the beginning of the formation of muscle fibers (arrows). D) Dorsal view on 5–6 week old embryo composed out of ~800 cells. The actin fibers of the myocytes are visible in all areas of the embryo. D’) Optical section of D) with subepidermal signal of BODIPY FL-phallacidin visible in multiple areas of the embryo. E) The labeling of tyrosinated-tubulin in 6–7 week old embryo shows the cilia in the epidermis of the embryo (yellow), dorsal view. The phallacidin labeling of the musculature has become more prominent but is still irregular. E’) Optical cross section through another embryo in the same age as E) The propidium iodide labeled nuclei and the musculature, dorsal view. F) The 7–8 week old embryo shows regularly arranged muscle fibers corresponding to the future pattern of the ring-musculature. F’) most nuclei are located at the apical pole of the epidermal cells (white arrows). Other nuclei are located also at the base of the epidermis (red arrows), likely the nuclei of the neural precursors of the basiepidermal nerve net. Scale bar 30 μm in all images except F’ 10 μm.