The endogenous retinoid metabolite S‐4‐oxo‐9‐cis‐13,14‐dihydro‐retinoic acid activates retinoic acid receptor signalling both in vitro and in vivo

Abstract

Retinoic acid receptor (RAR) and retinoid X receptor are ligand-induced transcription factors that belong to the nuclear receptor family. The receptors are activated by small hydrophobic compounds, such as all-trans-retinoic acid and 9-cis-retinoic acid, respectively. Interestingly, these receptors are also targets for a number of exogenous compounds. In this study, we characterized the biological activity of the 9-cissubstituted retinoic acid metabolite, S-4-oxo-9-cis-13,14-dihydro-retinoic acid (S-4o9cDH-RA). The endogenous levels of this metabolite in wild-type mice and rats were found to be higher than those of alltrans-retinoic acid, especially in the liver. Using cell-based luciferase reporter systems, we showed that S-4o9cDH-RA activates the transcription of retinoic acid response element-containing genes in several cell types, both from a simple 2xDR5 element and from the promoter of the natural retinoid target gene RARb2. In addition, quantitative RT-PCR analysis demonstrated that S-4o9cDH-RA treatment significantly increases the endogenous mRNA levels of the RAR target gene RARb2. Utilizing a limited proteolytic digestion assay, we showed that S-4o9cDH-RA induces conformational changes to both RARa and RARb in the same manner as does all-trans-retinoic acid, suggesting that S-4o9cDH-RA is indeed an endogenous ligand for these receptors. These in vitro results were corroborated in an in vivo system, where S-4o9cDH-RA induced morphological changes similar to those of all-trans-retinoic acid in the developing chicken wing bud. When locally applied to the wing bud, S-4o9cDH-RA induced digit pattern duplications in a dose-dependent fashion. The results illustrate that S-4o9cDH-RA closely mimics alltrans-retinoic acid with regard to pattern respecification. Finally, using quantitative RT-PCR analysis, we showed that S-4o9cDH-RA induces the transcription of several retinoic acid-regulated genes in chick wing buds, including Hoxb8, RARb2, shh, Cyp26 and bmp2. Although

From carrot to clinic: an overview of the retinoic acid signaling pathway

“…The endogenous levels of S-4o9cDH-RA were found to be higher than those of all-trans RA in mice and rats, and the compound is able to activate transcription of RAR target genes [120]. In vivo, S-4o9cDH-RA induces morphological changes in the chicken wing bud similar to those induced by all-trans RA [120]. Additional RAR ligands include b-cryptoxanthin and lutein, which act as RAR ligands in a yeast two-hybrid system and bind RA with lower affinity than all-trans RA [121].…”

“…The retinol metabolite 14-hydroxy-4,14-retro retinol was also shown to be active in vivo [10]. Another recently identified and characterized retinoid is the 9-cis-substituted RA metabolite S-4-oxo-9-cis-13,14-dihydro-RA (S-4o9cDH-RA) [120]. The endogenous levels of S-4o9cDH-RA were found to be higher than those of all-trans RA in mice and rats, and the compound is able to activate transcription of RAR target genes [120].…”

“…Another recently identified and characterized retinoid is the 9-cis-substituted RA metabolite S-4-oxo-9-cis-13,14-dihydro-RA (S-4o9cDH-RA) [120]. The endogenous levels of S-4o9cDH-RA were found to be higher than those of all-trans RA in mice and rats, and the compound is able to activate transcription of RAR target genes [120]. In vivo, S-4o9cDH-RA induces morphological changes in the chicken wing bud similar to those induced by all-trans RA [120].…”

Activation of Retinoic Acid Receptors by Dihydroretinoids

“…These studies led to the discovery of several retinoids with potential biological activity, but only a few enzymes have been identified that could produce them in vivo (Buck et al, 1991;Heyman et al, 1992;Schuchardt et al, 2009). We originally identified and characterized the enzyme known as retinol saturase (RetSat) that catalyzes the saturation of all-trans-retinol (atROL) through a stereospecific reaction that generates (R)-all-trans-13,14-dihydroretinol [(R)-DROL] (Moise et al, 2004(Moise et al, , 2007(Moise et al, , 2008.…”

Enzymatic intracrine regulation of white adipose tissue

“…Oxidized metabolites of RA. RA is metabolized by cytochrome P450 enzymes and its oxidized metabolites also activate RAR [141, 142]; however, the function of these RA derivatives in adipose tissue has not been reported.…”