Myelin is produced by oligodendrocyte lineage cells in the central nervous system. Numerous extracellular and intracellular molecules influence myelination, but how these diverse cues are integrated at the level of transcription to control myelination remains poorly understood. Nuclear Receptors (NRs) are transcription factors that bind a wide variety of lipophilic molecules and activate or repress transcriptional programs that control differentiation and maturation of cells. One NR signaling cascade important for myelination is mediated by Retinoid X Receptors (RXRs), which dimerize with other NR family members, potentially influencing multiple NR signaling cascades. Signaling interactions between RXRs and other NRs have been tested with in vitro cell assays, which fail to replicate the complex intercellular relationships and signaling interactions that occur in vivo. The zebrafish embryo is an excellent model to study myelination in vivo as embryos develop ex utero, myelination of the spinal cord occurs during the first week of life and each pair of zebrafish produce hundreds of embryos, allowing multiple drugs or combinations of drugs to be tested simultaneously for their impact on myelination. As such, we have developed a novel zebrafish model which quantifies the promoter response of a critical myelin gene, myelin protein zero (mpz), to RXR and NR ligands using a fluorescent or nanoLuciferase-based reporter system.
The mpz reporter embryo responds to known positive and negative regulators of myelination, consistent with changes in myelin mRNAs as quantified by qPCR. Initial studies of known RXR agonists and antagonists indicate that endogenous ligands such as 9-cis retinoic acid (9cRA) or Docosahexaenoic acid (DHA) have no significant effect on myelination in vivo, while synthetic molecules such as Fluorobexarotene, SR11237, UVI3003, PA 452, HX 630 and HX 531 can have significant dose-dependent effects on mpz promoter activity. These studies suggest that RXR-mediated signaling in vivo is more complicated than previously predicted and that investigating RXRs in an in vivo context will be essential for understanding the underlying mechanisms of action. Future experiments will use the mpz reporter to identify RXR and NR-mediated signaling interactions using an in vivo screening approach with the goal of identifing other pro-myelinating signaling interactions.