Complex Regulation of cyp26a1 Creates a Robust Retinoic Acid Gradient in the Zebrafish Embryo

Abstract

Positional identities along the anterior–posterior axis of the vertebrate nervous system are assigned during gastrulation by multiple posteriorizing signals, including retinoic acid (RA), fibroblast growth factors (Fgfs), and Wnts. Experimental evidence has suggested that RA, which is produced in paraxial mesoderm posterior to the hindbrain by aldehyde dehydrogenase 1a2 (aldh1a2/raldh2), forms a posterior-to-anterior gradient across the hindbrain field, and provides the positional information that specifies the locations and fates of rhombomeres. Recently, alternative models have been proposed in which RA plays only a permissive role, signaling wherever it is not degraded. Here we use a combination of experimental and modeling tools to address the role of RA in providing long-range positional cues in the zebrafish hindbrain. Using cell transplantation and implantation of RA-coated beads into RA-deficient zebrafish embryos, we demonstrate that RA can directly convey graded positional information over long distances. We also show that expression of Cyp26a1, the major RA-degrading enzyme during gastrulation, is under complex feedback and feedforward control by RA and Fgf signaling. The predicted consequence of such control is that RA gradients will be both robust to fluctuations in RA synthesis and adaptive to changes in embryo length during gastrulation. Such control also provides an explanation for the fact that loss of an endogenous RA gradient can be compensated for by RA that is provided in a spatially uniform manner.

Cyp26 enzymes function in endoderm to regulate pancreatic field size

“…The size of the pancreatic domain is defined not by limiting RA in the mesoderm, but rather by restricting RA signaling within the endoderm. This mechanism ensures that mesodermal RA remains available for signaling to other tissues, such as the ectoderm, where it has important roles in hindbrain patterning (13,15,16).…”

“…The 3 zebrafish cyp26 genes play redundant functions in the developing hindbrain, where they regulate RA-dependent gene expression (15). Reduction of zebrafish Cyp26 activity, using the cyp26a1 mutant giraffe (gir rw716 ), by morpholino knockdown or using pharmacological inhibitors, leads to excess RA signaling and hence changes in hindbrain patterning (13,15,16). Similarly, double mutant analysis of mouse Cyp26a1 and Cyp26c1 has indicated redundant roles in hindbrain patterning.…”

Fate of retinoic acid–activated embryonic cell lineages

“…Conditions of RA excess through maternal administration (Marshall et al, 1992;Simeone et al, 1995, and references therein) or Cyp26 knock-down Sakai et al, 2001;Emoto et al, 2005;Hernandez et al, 2007), and of RA deficiency through mutation of Rdh10 or Raldh2 (Niederreither et al, 2000;Sandell et al, 2007), have severe effects on hindbrain development and can lead to changes in rhombomeric molecular identities. Several models have been proposed to account for the effects of endogenous RA in controlling hindbrain development, mainly relying on the establishment of a posterior-toanterior RA diffusion gradient, and/or on the sequential action of CYP26 enzymes to generate RA-depleted areas in specific rhombomere territories (Gavalas, 2002;Maden, 2002;Maves and Kimmel, 2005;Sirbu et al, 2005;Hernandez et al, 2007;White et al, 2007;White and Schilling, 2008).…”

“…There has been some debate as to whether RA may act as a morphogen, i.e., may be released in the form of concentration gradients across developing morphogenetic fields (e.g., in the limb bud; Thaller and Eichele, 1987;Noji et al, 1991). Recent work has provided evidence that a RA gradient is operating across the hindbrain field in the zebrafish embryo (White et al, 2007;White and Schilling, 2008). The control of RA distribution within embryonic tissues results from an interplay between enzymes involved in its synthesis from retinol (mainly RDH10) and/or retinaldehyde (RALDH1, 2, and 3), and enzymes that metabolize it into hydroxylated derivatives and trigger its catabolism (CYP26A1, B1, and C1)-each of these enzymes being expressed in a stage and tissue-specific manner.…”

Interaction of retinoic acid and scl controls primitive blood development

“…In the zebrafish (Danio rerio), retinaldehyde dehydrogenase 2 (raldh2) is principally responsible for biosynthesis, whereas cyp26a1 is the main RA catabolic enzyme. 3,4 Retinoids bind to retinoic acid receptors (RARs) and retinoid X receptors, which dimerize to bind to RA responsive elements and activate gene transcription. Perturbing the delicate balance of RA signaling during development has profound effects on many tissues, including brain, heart, eye, limbs, and pancreas (reviewed by Duester 1 ), making retinoids highly teratogenic compounds.…”