Figure 1—figure supplement 1.

Figure 1—figure supplement 1. Evolution of the apical/luminal membrane throughout time.

(A) Two EHT pol+ cells visualized using a Tg(Kdrl:Gal4;UAS:RFP;4xNR:eGFP-podxl2) embryo and time-lapse sequence (initiated at 55 hpf) obtained with spinning disk confocal microscopy. The images (single z-planes) are extracted from the same time lapse sequence than the one used for Figure 1C. Green channel (eGFP-podxl2) only is shown. Green arrows point at the evolution of the connection between the aortic/eht cell lumens at t=0, 10, 30 min. Asterisks label the lumen delimited by the luminal/apical membranes. Scale bar = 8 µm. (B) Model summarizing and interpreting the temporal evolution of the luminal/apical membrane (in green, the asterisks mark the lumen of the apparent vacuole-like intracellular membrane structures) after the release of an EHT pol+ cell from the aortic floor. Step 1, the pseudo-vacuole filled with fluid and delimited by eGFP-podxl2 as visualized at t=65 min in (A) (and also at 45 min in Figure 1C, 2 asterisks) is consumed partly via budding (after sorting of eGFP-podxl2); these budding profiles can be seen on the left image (pink arrowheads) corresponding, for eht cell 1, to the plane 16 of the time point t=80 min of the time lapse sequence (see the Z-stack in Figure 1—video 2). Step 2, after sorting and budding, the cell remains with pseudo-endocytic Podxl2 containing membranes and the remaining vacuolar structures filled up with fluid regress, putatively by chasing water as illustrated in step 3. Step 4, the eht cell remains with pseudo-endocytic Podxl2 containing membranes that label newly born precursors of HSPCs. Note that since the cell remains in contact with the aortic floor while the pseudo-vacuole is regressing, the vacuole-like intracellular membrane proximal to aortic cells may never undergo fission (see Figure 1A, steps 4–4’) but gets consumed via budding and flattening upon water chase (steps 1–3 that are similar to Figure 1A, steps 3–3’). Scale bar = 8 µm.