Enzymatic Metabolism of Vitamin A in Developing Vertebrate Embryos

Abstract

Embryonic development is orchestrated by a small number of signaling pathways, one of which is the retinoic acid (RA) signaling pathway. Vitamin A is essential for vertebrate embryonic development because it is the molecular precursor of the essential signaling molecule RA. The level and distribution of RA signaling within a developing embryo must be tightly regulated; too much, or too little, or abnormal distribution, all disrupt embryonic development. Precise regulation of RA signaling during embryogenesis is achieved by proteins involved in vitamin A metabolism, retinoid transport, nuclear signaling, and RA catabolism. The reversible first step in conversion of the precursor vitamin A to the active retinoid RA is mediated by retinol dehydrogenase 10 (RDH10) and dehydrogenase/reductase (SDR family) member 3 (DHRS3), two related membrane-bound proteins that functionally activate each other to mediate the interconversion of retinol and retinal. Alcohol dehydrogenase (ADH) enzymes do not contribute to RA production under normal conditions during embryogenesis. Genes involved in vitamin A metabolism and RA catabolism are expressed in tissue-specific patterns and are subject to feedback regulation. Mutations in genes encoding these proteins disrupt morphogenesis of many systems in a developing embryo. Together these observations demonstrate the importance of vitamin A metabolism in regulating RA signaling during embryonic development in vertebrates.

Keywords: Dhrs3; Rdh10; development; embryo; metabolism; retinoic acid; retinoid; review; signaling; vitamin A.

Figure 1

Retinoid compounds differ with respect to the polar end group at the terminus of the chain. The compound all-trans-retinol (ROL) can be reversibly converted to all-trans-retinal (RAL) or to retinyl esters (RE). The carotenoid β-carotene can be converted to RAL. RAL can be irreversibly oxidized to retinoic acid (RA).

Figure 2

Canonical RA signaling activity occurs when RA regulates gene transcription by binding as ligand to RAR nuclear transcription factors at RARE sites. (A) RARs function as heterodimer with RXRs. Binding of RA (represented by orange hexagon) to the ligand-binding domain of a RAR heterodimerized with a RXR can activate transcription; (B) RAR–RXR heterodimers bind to DNA at RARE sites. Examples of classical RARE binding sites consist of two directly repeated hexamer motifs separated by spacer nucleotide sequences of different lengths. Examples of DR5, DR2, and DR1 RAREs from important RA-regulated developmental genes are shown.

Figure 3

RA is not present uniformly throughout an embryo, and RA signaling activity does not occur equally in all cells. The distribution of canonical ligand-dependent RA signaling in mouse embryos can be visualized by activity of a RARE-lacZ transgenic reporter [55]. (A) At embryonic day 8.5 (E8.5) RA signaling occurs predominantly in the trunk and the forebrain. RA negative tissues include the heart and most of head; (B) At E9.5 RA signaling remains strong in trunk. RA negative tissues include pharyngeal arch 1, which gives rise to the maxilla and mandible and most of the midbrain and hindbrain; (C) At E11.5 strong RA signaling activity is present in the eye, in the developing nasal region and throughout the developing spinal cord of the trunk. Forelimbs and hindlimbs are negative for RA signaling at this stage; (D) At E12.5 RA signaling is detected in the eye, in distinct regions of the trunk, and in interdigital regions of the forelimb and hindlimb. The gut, which is outside the body wall at this stage, is strongly positive for RA signal. RA-negative tissues include the tail and digits of the limb; (E) RA signaling occurs in many developing organs including the E13.5 heart. E, embryonic day; FB, forebrain; FL, forelimb; FNP, frontonasal prominence; G, gut; H, heart; HL, hindlimb; LA, left atrium; LV, left ventricle; NP, nasal prominence; PA1, pharyngeal arch 1; RA right atrium; RV, right ventricle; T, tail.

Figure 4

RA production in embryo cells is regulated by enzymatic metabolism and catabolism. Embryo cells obtain retinoids from maternal sources via circulation. In the blood serum the hydrophobic ROL is bound by RBP4. ROL enters embryo cells predominantly through membrane diffusion. ROL can also be transported into cells via RBP4 delivery to the membrane transporter STRA6. Inside the cell hydrophobic ROL can be associated with a membrane compartment, where it can exist as free retinoid. Hydrophobic ROL can be present in the aqueous cytosol if it is bound by RBP1. ADH enzymes do not participate in metabolism of ROL in embryo tissues under normal conditions because the cytoplasmic enzymes cannot act on RBP1-bound ROL. RDH10, a membrane bound SDR, acts on free ROL, possibly from a membrane pool, or alternatively, delivered from RBP1. A related membrane SDR, DHRS3 catalyzes the reduction of RAL to ROL. RDH10 and DHRS3 are oxidoreductases that act reciprocally and enhance the activity of each other. RDH10 functions primarily as an oxidase and DHRS3 acts chiefly as a reductase, the redox directions being defined by relative abundance of cofactors NAD⁺ and NADPH. An alternate mechanism to generate RAL is cleavage of β-carotene by BCO1. RAL is irreversibly converted to RA by members of the ALDH1A family. Once RA is generated by oxidation of RAL, it can have several fates. RA can bind to CRABP2, which can deliver the RA into the nucleus where it can bind to RARs. RA can also be eliminated by action of CYP26 family enzymes, or it can move to neighboring cells, possibly by diffusion or transport. ER, endoplasmic reticulum; mit, mitochondrion.