Figure 1

U0126 application during a specific time-window decreases ERK phosphorylation specifically. A: Diphospho-Erk staining in the trunk of wt 24 hpf embryos.B: U0126 eliminates ERK1/2 phosphorylation (green) in the trunk and tail of 24 hpf zebrafish embryos. Notice that U0126-treated embryos have been overexposed (can be judged by the autofluorescence of the yolk sac extension) and still fail to show specific staining. C: U0124 does not eliminate the p-Erk staining. D: PD98059 also does not eliminate p-Erk staining when given just below the level of lethal toxicity (25 μM). E: Somewhat weaker but still significant p-Erk staining in embryos treated with U0126 starting at 10 hpf. F: Western-blot analysis demonstrated a slight but incomplete reduction in pErk1/2 levels in 10-somite embryos.G: Stage and dose-dependency of U0126 application. There is a dramatic drop in the percentage of affected embryos when treatment is applied after epiboly is completed. H: Washout experiments reveal the necessity of U0126 to be present until at least the 18 somite stage to produce the full phenotype.