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

authored by

Jan Philipp Schuchardt, David Wahlström, Joëlle Rüegg, Norbert Giese, Madalina Stefan, Henning Hopf, Ingemar Pongratz, Helen Hakansson, Gregor Eichele, Katarina Pettersson, Heinz Nau

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-cis-substituted 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 all-trans-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 RAR. In addition, quantitative RT-PCR analysis demonstrated that S-4o9cDH-RA treatment significantly increases the endogenous mRNA levels of the RAR target gene RAR. Utilizing a limited proteolytic digestion assay, we showed that S-4o9cDH-RA induces conformational changes to both RAR and RARin 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 all-trans-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, RAR, shh, Cyp26 and bmp2. Although S-4o9cDH-RA was less potent when compared with all-trans-retinoic acid, the findings clearly demonstrate that S-4o9cDH-RA has the capacity to bind and activate nuclear retinoid receptors and regulate gene transcription both in vitro and in vivo.

Organisation(s)

Institute of Food Science and Human Nutrition
Nutrition Physiology and Human Nutrition Section

External Organisation(s)

Karolinska Institutet
Technische Universität Braunschweig
Max Planck Institute for Biophysical Chemistry (Karl Friedrich Bonhoeffer Institute)
University of Veterinary Medicine of Hannover, Foundation

Type

Article

Journal

FEBS Journal

Volume

276

Pages

3043-3059

No. of pages

17

ISSN

1742-464X

Publication date

06.2009

Publication status

Published

Peer reviewed

Yes

ASJC Scopus subject areas

Biochemistry, Molecular Biology, Cell Biology

Electronic version(s)

https://doi.org/10.1111/j.1742-4658.2009.07023.x (Access: Unknown)