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The Mesp bHLH genes play a conserved role during segmental patterning of the mesoderm in the vertebrate embryo by specifying segmental boundaries and anteroposterior (A-P) segmental polarity. Here we use a xenotransgenic approach to compare the transcriptional enhancers that drive expression of the Mesp genes within segments of the presomitic mesoderm (PSM) of different vertebrate species. We find that the genomic sequences upstream of the mespb gene in the pufferfish Takifugu rubripes (Tr-mespb) are able to drive segmental expression in transgenic Xenopus embryos while those from the Xenopus laevis mespb (Xl-mespb) gene drive segmental expression in transgenic zebrafish. In both cases, the anterior segmental boundary of transgene expression closely matches the expression of the endogenous Mesp genes, indicating that many inputs into segmental gene expression are highly conserved. By contrast, we find that direct retinoic acid (RA) regulation of endogenous Mesp gene expression is variable among vertebrate species. Both Tr-mespb and Xl-mespb are directly upregulated by RA, through a complex, distal element. By contrast, RA represses the zebrafish Mesp genes. We show that this repression is mediated, in part, by RA-mediated activation of the Ripply genes, which together with Mesp genes form an RA-responsive negative feedback loop. These observations suggest that variations in a direct response to RA input may allow for changes in A-P patterning of the segments in different vertebrate species.

Somites, the segmented building blocks of the vertebrate embryo, arise one by one in a patterning process that passes wavelike along the anteroposterior axis of the presomitic mesoderm (PSM). We have studied this process in Xenopus embryos by analyzing the expression of the bHLH gene, Thylacine1, which is turned on in the PSM as cells mature and segment, in a pattern that marks both segment boundaries and polarity. Here, we show that this segmental gene expression involves a PSM enhancer that is regulated by retinoic acid (RA) signaling at two levels. RA activates Thylacine1 expression in rostral PSM directly. RA also activates Thylacine1 expression in the caudal PSM indirectly by inducing the expression of MKP3, an inhibitor of the FGF signaling pathway. RA signaling is therefore a major contributor to segmental patterning by promoting anterior segmental polarity and by interacting with the FGF signaling pathway to position segmental boundaries.

The vertebrate central nervous system (CNS) is induced by signals emanating from the dorsal mesoderm, or organizer, that divert the ectoderm away from an epidermal and towards a neural fate. Additional signals from the organizer pattern the neural ectoderm along the anteroposterior axis. We devised highly specific methods utilizing constitutively active or dominant negative receptors to evaluate the role of retinoids in neural patterning. Microinjection of these reagents either augments or reduces retinoid signaling in specific regions of the embryo. We show that increased receptor activity suppresses anterior neural structures while dominant negative receptors lead to anterior enhancement. Similarly, microinjection of the dominant negative receptor leads to the loss of posterior marker genes. We demonstrate that retinoid receptors comprise a critical component in neural posteriorization and are required for proper neuronal differentiation. These results support a quantitative role for retinoid signaling in regionalization of the CNS.