Retinoic Acid Promotes Limb Induction through Effects on Body Axis Extension but Is Unnecessary for Limb Patterning

Development and Aging Program, Burnham Institute for Medical Research, 10901 North Torrey Pines Road, La Jolla, CA 92037, USA.
Current biology: CB (Impact Factor: 9.57). 07/2009; 19(12):1050-7. DOI: 10.1016/j.cub.2009.04.059
Source: PubMed


Retinoic acid (RA) is thought to be a key signaling molecule involved in limb bud patterning along the proximodistal or anteroposterior axes functioning through induction of Meis2 and Shh, respectively. Here, we utilize Raldh2-/- and Raldh3-/- mouse embryos lacking RA synthesis to demonstrate that RA signaling is not required for limb expression of Shh and Meis2. We demonstrate that RA action is required outside of the limb field in the body axis during forelimb induction but that RA is unnecessary at later stages when hindlimb budding and patterning occur. We provide evidence for a model of trunk mesodermal RA action in which forelimb induction requires RA repression of Fgf8 in the developing trunk similar to how RA controls somitogenesis and heart development. We demonstrate that pectoral fin development in RA-deficient zebrafish embryos can be rescued by an FGF receptor antagonist SU5402. In addition, embryo ChIP assays demonstrate that RA receptors bind the Fgf8 promoter in vivo. Our findings suggest that RA signaling is not required for limb proximodistal or anteroposterior patterning but that RA inhibition of FGF8 signaling during the early stages of body axis extension provides an environment permissive for induction of forelimb buds.

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Available from: Ovidiu Sirbu, Mar 18, 2014
    • "For the mouse Aldh1a2 KOs, although the cardiac phenotypes were initially interpreted as supporting the atrial-ventricular patterning model from cardiac morphology, revisiting the analysis of the cardiac defects with additional cardiac progenitor markers revealed that these mutants also display a posterior expansion of the cardiac progenitors (Ryckebusch et al., 2008; Sirbu et al., 2008). Studies initially in zebrafish and later in mice have suggested that the posterior expansion of the cardiac progenitors is potentially at the expense of neighboring forelimbs progenitors (Waxman et al., 2008; Zhao et al., 2009; Sorrell and Waxman, 2011; Cunningham et al., 2013). Despite the genetic data supporting a conserved requirement for RA in restricting cardiomyocyte specification, there are differences in interpretation as to whether or not there is strictly an expansion of the FHF, the earlier differentiating population of cardiomyocytes , and/or the second SHF, a later differentiating population of cardiomyocytes (Ryckebusch et al., 2008; Sirbu et al., 2008). "
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    ABSTRACT: Appropriate levels of retinoic acid (RA) signaling are critical for normal heart development in vertebrates. A fascinating property of RA signaling is the thoroughness by which positive and negative feedback are employed to promote proper embryonic RA levels. In the present short review, we first cover the advancement of hypotheses regarding the impact of RA signaling on cardiac specification. We then discuss our current understanding of RA signaling feedback mechanisms and the implications of recent studies, which have indicated improperly maintained RA signaling feedback can be a contributing factor to developmental malformations. Developmental Dynamics 244:513-523, 2015. © 2014 Wiley Periodicals, Inc.
    Developmental Dynamics 11/2014; 244(3). DOI:10.1002/dvdy.24232 · 2.38 Impact Factor
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    • "Retinoic acid (RA) produced by retinaldehyde dehydrogenase 2 (Raldh2; Aldh1a2) functions as a diffusible signal controlling vertebrate development (Duester, 2008; Niederreither and Dollé, 2008). Loss of RA synthesis in RA-deficient chick embryos and mouse Raldh2 −/− embryos results in ectopic Fgf8 expression extending from the caudal progenitor zone into the trunk that disrupts somitogenesis and neurogenesis (Diez del Corral et al., 2003; Molotkova et al., 2005; Vermot et al., 2005; Vermot and Pourquié, 2005; Sirbu and Duester, 2006) as well as forelimb initiation (Zhao et al., 2009; Cunningham et al., 2013). Thus, RA functions as a diffusible signal that downregulates Fgf8 as cells exit the caudal progenitor zone. "
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    ABSTRACT: Retinoic acid (RA) generated in the mesoderm of vertebrate embryos controls body axis extension by downregulating Fgf8 expression in cells exiting the caudal progenitor zone. RA activates transcription by binding to nuclear RA receptors (RARs) at RA response elements (RAREs), but it is unknown whether RA can directly repress transcription. Here, we analyzed a conserved RARE upstream of Fgf8 that binds RAR isoforms in mouse embryos. Transgenic embryos carrying Fgf8 fused to lacZ exhibited expression similar to caudal Fgf8, but deletion of the RARE resulted in ectopic trunk expression extending into somites and neuroectoderm. Epigenetic analysis using chromatin immunoprecipitation of trunk tissues from E8.25 wild-type and Raldh2(-/-) embryos lacking RA synthesis revealed RA-dependent recruitment of the repressive histone marker H3K27me3 and polycomb repressive complex 2 (PRC2) near the Fgf8 RARE. The co-regulator RERE, the loss of which results in ectopic Fgf8 expression and somite defects, was recruited near the RARb RARE by RA, but was released from the Fgf8 RARE by RA. Our findings demonstrate that RA directly represses Fgf8 through a RARE-mediated mechanism that promotes repressive chromatin, thus providing valuable insight into the mechanism of RA-FGF antagonism during progenitor cell differentiation.
    Development 08/2014; 141(15):2972-7. DOI:10.1242/dev.112367 · 6.46 Impact Factor
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    • "Fgf8 expression is initiated upon limb but outgrowth in spite of the inhibitory role of RA, and the simulations predict that receptor binding limits diffusion of RA from the flank initially, once RA signalling enhances the expression of RA receptors, as indeed observed in experiments (Noji et al., 1991; Tabin, 1991). According to the model, receptor saturation eventually permits RA to diffuse further distally and to form a gradient that could regulate aspects of proximal-distal limb bud development (Figure 4C) and that could define the proximal part (stylopod -> humerus) of the proximal-distal axis as suggested by two recent studies (Cooper et al., 2011a; Roselló-Díez et al., 2011) and challenged by the Duester group (Cunningham et al., 2013; Zhao et al., 2009). "
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