Movies 2-5. Convergence and extension of zebrafish dorsal mesoderm. Time sequences taken from a single four hour, 4D recording of a wild-type embryo, beginning at midgastrula stage (7.3 hpf). The anteroposterior (AP) axis is vertical (anterior towards the top) and the mediolateral (ML) axis horizontal. The field is centered approximately on the dorsal midline. (Movie 2) Views from the recording, from 7.3 to 11.3 hpf. Confocal microscope images of the BODIPY-ceramide labeled cellular field are shown (Cooper et al., 1999b; Glickman, 2000; Thisse et al., 1993). At the first time point the blastoderm margin is evident, separating the cellular blastoderm (upper) from the yolk syncytial layer (YSL, lower). Subsequently, the blastoderm comes to cover the yolk completely by the spreading movement of epiboly (Kimmel et al., 1995; Solnica-Krezel et al., 1996). Brachet’s cleft separates ectoderm to the outside and mesoderm to the inside. The notochord/somite boundaries (axial/paraxial boundaries) appear in the mesoderm, they are barely visible and then become prominent. Convergence narrows the notochord domain to a width of two cells at the last time point. At the lateral side of each boundary lies a distinctive row of somitic adaxial cells. (Movie 3). About 200 cells, represented as spheres, were tracked from the recording of Movie 2. Cells are color coded according to their eventual fates: notochord-forming cells (green), adaxial cells (dark blue), cells forming somite #2 (yellow), other somite-domain cells (red). Scale bar: 50 µm. (Movie 4) Notochord-forming cells (green) shear with overlying floorplate-forming cells (magenta) during extension. The two cell types were collected from separate focal planes of the original 4D recording, and a 30 µm stripe cut at a single AP level. Scale bar: 50 µm. (Movie 5) The notochord domain (green) extends more than the somite domains (red). A 30 µm horizontal stripe of mesoderm cut from the field and is shown.

Movie 6.MIB can account for all or most organized cellular movement within the wild-type notochord domain. The purple lines show movement pathways of cells during 16 minute intervals at the beginning and end (4 hours later) of the recording used for Fig. 3. The yellow lines show the essentially random movements left over after using the values of kC and kE to subtract the components of the cell movements due to MIB at every time point. During the 4 hour period, the change in shape of the field is negligible.

Movies 7-12.Loss of function of ntl disrupts convergence but not extension. The wild-type data are from the same 4D recording used for Fig. 3. (Movies 7, 8) Cellular movement pathways in the notochord/axial (green) and somite domains (red). The axial and somite domains intermix in the ntl mutant (Movie 8) whereas the notochord and somite domains do not intermix in the wild type (Movie 7). The directions of the arrows show the directions of cell movements and the lengths show the speed (the movement history during 40 minutes). Disruption of convergence is evident in the ntl axial domain. Horizontally (ML) oriented tracks are largely missing in ntl mutant. However, extension, vertical lengthening of the field, is prominent in the mutant. (Movies 9, 10) Intermixing does not occur between the notochord/axial domain (green) and the overlying midline epiblast (floorplate domain, magenta) in either the wild type (Movie 9) or ntl mutant (Movie 10). Side views made by 90° rotation of vertical strips of cells in the tracked data sets that are present at the dorsal midline at each time point. (Movies 11, 12) The thickness of the region of tracked dorsal mesodermal cells does not change greatly during convergence and extension in the wild type (Movie 11), and may decrease slightly during extension without convergence in the ntl mutant (Movie 12) axial domain (green). Scale bar: 50 mm.

(Movies 13, 14) Extensive cellular mixing along the ML axis occurs in ntl (Movie 14) but not in wild-type (Movie 13) embryos. The spheres representing tracked cells color coded to produce vertical stripes, 30 µm wide, at the first time point (8 hpf). To facilitate comparison with ntl, only the middle part of the tracked field is shown for the wild type. The stripes all narrow in the wild type, but they mix rather than narrow in ntl–. (Movies 15, 16) Cells move toward the midline in wild type (Movie 15), but move in a disorderly way with respect to the midline in ntl (Movie 16). Red and blue circles show cells moving to the right and left, respectively, green circles show cells with no left-right component to their movement.