Fig 2

(A–A””) Tg(fli1a:gal4ff;UAS:Kaede) embryonic hearts stopped using BDM and photoconverted at 48 to 50 hpf (left column). Embryos were then returned to normal media and allowed to grow normally until 144 hpf, when the hearts were stopped using BDM again and imaged (middle and right columns). (A) Embryos where the ventricle and the cell at the ventricular side of the AVC is photoconverted. Photoconverted cells can be seen between the 2 luminal layers of the valve at 144 hpf. (Ventricular cells appear to have degraded photoconverted Kaede.) (A’) Embryos where the ventricle is photoconverted. Photoconverted cells can be seen at the luminal side of the valve base at 144 hpf. (Ventricular cells appear to have degraded photoconverted Kaede.) (A”) Embryo where one cell migrating in is photoconverted. Photoconverted cells can be seen between the 2 luminal layers of the valve at 144 hpf. (A”’) Embryos where the deepest cell inside the CJ and the atrial edge of the AVC are photoconverted. At 144 hpf, photoconverted cells can be seen between the 2 luminal layers of the valve, presumably derived from the cell that was deepest inside the CJ at 48 to 50 hpf. Luminal photoconverted cells can also be seen at the tip of the valve, presumably derived from the atrial side of the AVC at 48 to 50 hpf. (A””) Embryos where the atrial side of the AVC is photoconverted. Photoconverted cells can be found at the ventricular luminal layer of the valve leaflet. Asterisks label abluminal cells. Scale bar left column: 50 μm. Scale bar middle and right columns: 20 μm. (B) Model for zebrafish valve leaflet formation, cells are colored to indicate their position and fate over time. At 50 hpf, red cells represent ventricular endocardial cells. Yellow cells represent endocardial cells at the ventricular edge of the AVC. Green cells represent the remaining endocardial cells in the AVC. Blue cells represent atrial endocardial cells in the AVC. In subsequent stages, color schemes are kept to show the position and fate of cells over time. Cells derived from cells at 50 hpf due to cell proliferation are colored the same color as their mothers. Arrows in the top drawing indicate cell movements. Gray box marks period at which delamination can occur (65 to 80 hpf). Delamination itself takes place within 1 hour. At, atrium; AVC, atrioventricular canal; BDM, 2,3-butanedione monoxime; CJ, cardiac jelly; hpf, hours postfertilization; V, ventricle; VIC, valve interstitial cell.