Fig. 6

 Co-expression of p57kip2 or dnPKA with zRab11-FIP4 morpholino. (A, B) The cell cycle was analyzed by BrdU incorporation. Zebrafish embryos were injected with zRab11-FIP4-MO1 in combination with a plasmid to express p57Kip2/EGFP, dnPKA-EGFP or EGFP alone (control) under the regulation of the heat shock promoter. These genes were induced by incubating at 39°C for 1 h (24 to 25 hpf), and proliferating retinal cells were then examined by labeling with BrdU for 6 h (50–56 hpf). Immunostaining patterns with anti-BrdU and GFP are shown in panel A, and GFP-positive cells that had exited from the cell cycle are indicated by arrowheads. L, lens. The ratio of BrdU/GFP double-positive cells to total GFP-positive cells is shown in panel B. The difference between the control and either p57Kip2 or dnPKA-expressing samples is statistically significant (P < 0.001 in each case). (C) A plasmid GFP:HSE:p57Kip2 was coinjected with zRab11-FIP4-MO1 or control-MO into 1- to 4-cell embryos. Differentiation of photoreceptors was examined by immunostaining with zpr-1 at 60 hpf. Zpr-1/GFP double-positive cells are pointed out by arrowheads. Scale bars indicate 50 μm.