Savory Joanne G A, Edey Caitlin, Hess Bradley, Mears Alan J, Lohnes David
Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, ON, Canada.
Children's Hospital of Eastern Ontario Research Institute, University of Ottawa, Ottawa, ON, Canada.
Dev Biol. 2014 Nov 15;395(2):199-208. doi: 10.1016/j.ydbio.2014.09.013. Epub 2014 Sep 22.
Retinoic acid is required for diverse ontogenic processes and as such identification of the genes and pathways affected by retinoic acid is critical to understanding these pleiotropic effects. The presomitic mesoderm of the E8.5 mouse embryo is composed of undifferentiated cells that are depleted of retinoic acid, yet are competent to respond to the retinoid signal. We have exploited these properties to use this tissue to identify novel retinoic acid-responsive genes, including candidate target genes, by treating E8.5 embryos with retinoic acid and assessing changes in gene expression in the presomitic mesoderm by microarray analysis. This exercise yielded a cohort of genes that were differentially expressed in response to exogenous retinoic acid exposure. Among these were a number of previously characterized retinoic acid targets, validating this approach. In addition, we recovered a number of novel candidate target genes which were confirmed as retinoic acid-responsive by independent analysis. Chromatin immunoprecipitation assays revealed retinoic acid receptor occupancy of the promoters of certain of these genes. We further confirmed direct retinoic acid regulation of the F11r gene, a new RA target, using tissue culture models. Our results reveal a significant number of potential RA targets implicated in embryonic development and offer a novel in vivo system for better understanding of retinoid-dependent transcription.
视黄酸是多种个体发育过程所必需的，因此，鉴定受视黄酸影响的基因和信号通路对于理解这些多效性作用至关重要。E8.5小鼠胚胎的体节中胚层由未分化的细胞组成，这些细胞缺乏视黄酸，但能够对视黄酸信号作出反应。我们利用这些特性，通过用视黄酸处理E8.5胚胎，并通过微阵列分析评估体节中胚层基因表达的变化，利用该组织鉴定新的视黄酸反应基因，包括候选靶基因。这项工作产生了一组因外源性视黄酸暴露而差异表达的基因。其中有许多先前已鉴定的视黄酸靶标，验证了这种方法的有效性。此外，我们还发现了一些新的候选靶基因，通过独立分析证实它们对视黄酸有反应。染色质免疫沉淀分析揭示了视黄酸受体对其中某些基因启动子的占据情况。我们使用组织培养模型进一步证实了新的视黄酸靶标F11r基因受视黄酸的直接调控。我们的结果揭示了大量与胚胎发育相关的潜在视黄酸靶标，并提供了一个新的体内系统，以更好地理解视黄酸依赖性转录。
