FIGURE 2.

Phenotypic and molecular robustness of the retinoic acid metabolic pathway. Retinoic acid levels were reduced in Xenopus laevis embryos by inhibition of the RALDH activity with DEAB or by CYP26A1 overexpression to render retinoic acid inactive. (A) Control embryo at st. 27. (B) Embryo injected with capped RNA (0.8 ng) encoding the CYP26A1 enzyme. (C) Embryo treated with DEAB (50 μM) from st. 8.5 until st. 27. (D) Embryo treated with DEAB and injected with cyp26a1 mRNA. To exemplify the induced developmental malformations, lines depicting the size of the head domain (blue) and the trunk (red) were drawn on the control embryo (A) and then copied unto the treated embryos (B–D). (E) Distribution of the developmental defect severity induced by the retinoic acid manipulations. Embryos were scored for the induction of moderate (B,C) or severe (D) phenotypes, or normal looking. (F–H) Gene expression changes as a result of retinoic acid manipulation. qPCR analysis of hoxa1.L (F), hoxa1.S (G), and dhrs3.L (H) relative expression levels as a result of the individual or combined retinoic acid manipulations. Statistical significance (Student’s t-test) was calculated compared to the combined treatment group. *p < 0.05; **p < 0.01; ****p < 0.0001; ns, not significant.