Rdh10 mutants deficient in limb field retinoic acid signaling exhibit normal limb patterning but display interdigital webbing

Sanford-Burnham Medical Research Institute, Development and Aging Program, La Jolla, California, CA, USA.
Developmental Dynamics (Impact Factor: 2.38). 05/2011; 240(5):1142-50. DOI: 10.1002/dvdy.22583
Source: PubMed


Retinoic acid (RA) is purported to be required for expression of genes controlling proximodistal (Meis2) or anteroposterior (Shh) limb patterning. Embryos lacking RDH10, the primary enzyme synthesizing retinaldehyde during mouse development, survive until E14.5 with stunted forelimbs but apparently normal hindlimbs. Using embryos carrying the RARE-lacZ RA-reporter transgene, we show that endogenous RA activity in Rdh10(trex/trex) mutants is detected in neuroectoderm but not limbs during initiation and patterning. Treatment of Rdh10 mutants with 25 nM RA restores RARE-lacZ activity to limb mesoderm, validating RARE-lacZ and verifying that RA is absent in mutant limbs. In Rdh10 mutants, hindlimbs exhibit normal Meis2/Shh expression and skeletal patterning, and although forelimbs are growth-retarded their Meis2 expression remains normal. Later in development, Rdh10 mutants lack interdigital RA activity and accordingly fail to exhibit normal loss of interdigital mesenchyme. These findings demonstrate that RA is unnecessary for limb patterning but required later for interdigital tissue loss.

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    • "In line with this, earlier experiments had shown that low doses of an RA implant increase the length of the AER, while higher doses decrease the length of the ridge (Summerbell, 1983; Tickle et al., 1982). On the other hand, it has been noted that there is no proximal expansion in AER-Fgf8 expression in conditional mutants of an RA producing enzyme (Cunningham et al., 2011). However, the same group also showed that RA is necessary for the initiation of forelimbs as otherwise ectopic Fgf8 expression prevents forelimb initiation (Cunningham et al., 2013; Zhao et al., 2009). "
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    • "The embryonic abnormalities and RA reduction observed in Rdh10trex mutant mice are dramatic, but generally not as severe as those caused by loss of RALDH2. Embryos lacking RALDH2 do not survive past E10.5 [8], [27], while most homozyogous Rdh10trex embryos remain viable through E10.5–E11.5 and embryos have been identified surviving as late as E14.5 [23], [28]. The residual production of RA and the less severe phenotype of Rdh10trex embryos compared to Aldh1a2 mutant embryos indicate that some RDH activity remains intact in the Rdh10trex embryos, mediated possibly by hypomorphic activity of the Rdh10trex point mutant enzyme, by other RDH enzymes, or by ADH enzymes. "
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    ABSTRACT: Retinoic Acid (RA) is a small lipophilic signaling molecule essential for embryonic development and adult tissue maintenance. Both an excess of RA and a deficiency of RA can cause pathogenic anomalies, hence it is critical to understand the mechanisms controlling the spatial and temporal distribution of RA. However, our current understanding of these processes remains incomplete. Vitamin A is metabolized to RA via two sequential enzymatic reactions. The first requires retinol dehydrogenase (RDH) activity to oxidize Vitamin A (retinol) to retinal, and the second requires retinaldehyde activity (RALDH) to oxidize retinal into RA. The first reaction has previously been attributed to the alcohol dehydrogenase (ADH) family, whose genes are ubiquitously or redundantly expressed. Consequently, the specificity of RA synthesis was thought to reside exclusively at the level of the second reaction. To better understand the metabolism of Vitamin A into RA during embryogenesis, we generated new mouse models that disrupt this process. Here we describe a new targeted knockout of Rdh10 in which RA synthesis is severely impaired, particularly at critical early embryonic stages. We also introduce a new mutant allele of Aldh1a2. Both mutations produce similar developmental defects resulting in lethality around embryonic day 10.5 (E10.5). The severity of the Rdh10 null phenotype demonstrates that embryonic oxidation of retinol is carried out primarily by RDH10 and that neither ADHs nor other enzymes contribute significantly to this reaction. We also show that reduced RA production results in upregulation of Rdh10. These data demonstrate that RDH10 plays a critical role in mediating the rate limiting RDH step of Vitamin A metabolism and functions as a nodal point in feedback regulation of RA synthesis. Moreover, RDH10-mediated oxidation of retinol plays as important a role in the control and regulation of RA production during embryogenesis as does the subsequent RALDH-mediated reaction.
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