Embryonic retinoic acid synthesis is essential for early mouse post-implantation development

Abstract

A number of studies have suggested that the active derivative of vitamin A, retinoic acid (RA), may be important for early development of mammalian embryos1,2. Severe vitamin A deprivation in rodents results in maternal infertility3, precluding a thorough investigation of the role of RA during embryogenesis. Here we show that production of RA by the retinaldehyde dehydrogenase-2 (Raldh2) enzyme4,5 is required for mouse embryo survival and early morphogenesis. Raldh2 is an NAD-dependent aldehyde dehydrogenase with high substrate specificity for retinaldehyde4,5. Its pattern of expression during mouse development has suggested that it may be responsible for embryonic RA synthesis4,6. We generated a targeted disruption of the mouse Raldh2 gene and found that Raldh2–/– embryos, which die at midgestation without undergoing axial rotation (body turning), exhibit shortening along the anterioposterior axis and do not form limb buds. Their heart consists of a single, medial, dilated cavity. Their frontonasal region is truncated and their otocysts are severely reduced. These defects result from a block in embryonic RA synthesis, as shown by the lack of activity of RA-responsive transgenes, the altered expression of an RA-target homeobox gene and the near full rescue of the mutant phenotype by maternal RA administration. Our data establish that RA synthesized by the post-implantation mammalian embryo is an essential developmental hormone whose lack leads to early embryo death.

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Figure 1: Morphological abnormalities of Raldh2-/- embryos.
Figure 2: Histological analysis of 9.5-dpc embryos.
Figure 3: Whole-mount in situ analysis of Raldh2–/– embryos.
Figure 4: Altered RA-responsive transgene activity and RA-mediated phenotypic rescue of Raldh2 mutants.
Figure 5: Analysis of Otx2 and Hoxa1 transcript expression in Raldh2-/- embryos.

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Acknowledgements

We thank A. Dierich and M. Le Meur for supervising ES cell and animal facility work; N. Messadeq for the scanning electron micrographs; B. Schuhbaur and V.Fraulob for technical assistance; J. Rossant for providing RAREhsplacZ mice; and S.L. Ang (Otx2), P. Bouillet (Meox1), G. Gradwohl (Hand1), J. Grippo (Hoxa1), B. Herrmann (T), B. Hogan (Fgf10), A.P. McMahon (Fgf3) and G. Martin (Fgf8) for in situ hybridization probes. This work was supported by funds from the Centre National de la Recherche Scientifique, the Institut National de la Santé et de la Recherche Médicale, the Collège de France, the Hôpitaux Universitaires de Strasbourg, the Association pour la Recherche sur le Cancer, the Fondation pour la Recherche Médicale and Bristol-Myers Squibb. K.N. was supported by a fellowship from the Université Louis Pasteur (Strasbourg).

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Correspondence to Pierre Chambon.

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