Fig. S4

Expression and distribution of twist1+ cells during development coincides with delamination of tagln/sm22+ membrane from the sclerotome. (A,B) Confocal transverse view images of green unconverted kaede fluorescence in the mid-trunk of Tg(Ola-twist:gal4), Tg(uas:kaede) animals at 1 dpf (A) and 3 dpf (B). Twist+ cells are initially found in the sclerotome region of the somites (A), but later become concentrated around the vicinity of the dorsal aorta (B). Representative images were obtained from n = 3-5 fish; 2 rounds of immunostaining. (C) Confocal lateral view images of unconverted kaede (green) on a red fluorescent dorsal aorta (white) of a Tg(Ola-twist:gal4), Tg(uas:kaede), Tg(kdrl:mCherryCAAX) triple transgenic animal at 2.5 dpf. Images were obtained from 5 fish, from 2 separate experiments and a representative image chosen. (D) Schematic diagram of the Tg(tagln:egfp) transgene, with the transgelin (tagln) promoter driving expression of egfp in the cytoplasm. (E) Selected frames from a confocal time-lapse image series collected from a Tg(tagln:egfp) (in green); Tg(kdrl:mCherryCAAX) (in white) double-transgenic fish shows the movement of a tagln+ membrane (yellow) from the adjacent sclerotome delaminate and associate with the dorsal aorta. The image reconstructions shown are tilted ventral views, looking at a 45o upward tilted angle along the length of the dorsal aorta. Confocal images were acquired every 10 minutes and the displayed stills are shown at approximately 1-hour intervals. Scale bars = 50 μm. In total, 13 tagln+ cells investing the DA were traced to the sclerotome in 6 independent time-lapse experiments.