| In vitro: |
| Dev Biol. 2014 Nov 15;395(2):199-208. | | Identification of novel retinoic acid target genes.[Pubmed: 25251699] | 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.
METHODS AND RESULTS:
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.
CONCLUSIONS:
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.
| | J Nutr Biochem. 2014 Sep;25(9):964-76. | | A RARE of hepatic Gck promoter interacts with RARα, HNF4α and COUP-TFII that affect retinoic acid- and insulin-induced Gck expression.[Pubmed: 24973045] | The expression of hepatic glucokinase gene (Gck) is regulated by hormonal and nutritional signals. How these signals integrate to regulate the hepatic Gck expression is unclear. We have shown that the hepatic Gck expression is affected by Vitamin A status and synergistically induced by insulin and retinoids in primary rat hepatocytes.
METHODS AND RESULTS:
We hypothesized that this is mediated by a Retinoic acid responsive element (RARE) in the hepatic Gck promoter. Here, we identified the RARE in the hepatic Gck promoter using standard molecular biology techniques. The single nucleotide mutations affecting the promoter activation by Retinoic acid (RA) were also determined for detail analysis of protein and DNA interactions. We have optimized experimental conditions for performing electrophoresis mobility shift assay and demonstrated the interactions of the Retinoic acid receptor α (RARα), retinoid X receptor α (RXRα), hepatocyte nuclear factor 4α (HNF4α) and chicken ovalbumin upstream promoter-transcription factor II (COUP-TFII) in the rat nuclear extract with this RARE, suggesting their roles in the regulation of Gck expression. Chromatin immunoprecipitation assays demonstrated that recombinant adenovirus-mediated overexpression of RARα, HNF4α and COUP-TFII, but not RXRα, significantly increased their occupancy in the hepatic Gck promoter in primary rat hepatocytes. Overexpression of RARα, HNF4α and COUP-TFII, but not RXRα, also affected the Retinoic acid- and insulin-mediated Gck expression in primary rat hepatocytes.
CONCLUSIONS:
In summary, this hepatic Gck promoter RARE interacts with RARα, HNF4α and COUP-TFII to integrate Vitamin A and insulin signals. |
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Cell. 2018 Jan 11;172(1-2):249-261.e12. doi: 10.1016/j.cell.2017.12.019.IF=36.216(2019)
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