In Silico and In Vitro Identification of MicroRNAs That Regulate Hepatic Nuclear Factor 4α Expression
Open Access
- 9 January 2012
- journal article
- Published by American Society for Pharmacology & Experimental Therapeutics (ASPET) in Drug Metabolism and Disposition
- Vol. 40 (4), 726-733
- https://doi.org/10.1124/dmd.111.040329
Abstract
Hepatic nuclear factor 4α (HNF4A) is a nuclear transcription factor that regulates the expression of many genes involved in drug disposition. To identify additional molecular mechanisms that regulate HNF4A, we identified microRNAs (miRNAs) that target HNF4A expression. In silico analyses suggested that HNF4A is targeted by many miRNAs. We conducted in vitro studies to validate several of these predictions. With use of an HNF4A 3′-untranslated region (UTR) luciferase reporter assay, five of six miRNAs tested significantly down-regulated (∼20–40%) the luciferase activity. In HepG2 cells, miR-34a and miR-449a also down-regulated the expression of both the HNF4A protein and an HNF4A target gene, PXR (∼30–40%). This regulation appeared without reduction in HNF4A mRNA expression, suggesting that they must be blocking HNF4A translation. Using additional bioinformatic algorithms, we identified polymorphisms that are predicted to alter the miRNA targeting of HNF4A. Luciferase assays indicated that miR-34a and miR-449a were less effective in regulating a variant (rs11574744) than the wild-type HNF4A 3′-UTR. In vivo, subjects with the variant HNF4A had lower CYP2D6 enzyme activity, although this result was not statistically significant (p = 0.16). In conclusion, our findings demonstrate strong evidence for a role of miRNAs in the regulation of HNF4A.Keywords
This publication has 40 references indexed in Scilit:
- Conserved Seed Pairing, Often Flanked by Adenosines, Indicates that Thousands of Human Genes are MicroRNA TargetsCell, 2005
- Human MicroRNA TargetsPLoS Biology, 2004
- Polymorphisms associated with circulating sex hormone levels in postmenopausal women.JNCI Journal of the National Cancer Institute, 2004
- MicroRNA-Directed Cleavage of HOXB8 mRNAScience, 2004
- MicroRNAs: Genomics, Biogenesis, Mechanism, and FunctionCell, 2004
- Prediction of Mammalian MicroRNA TargetsCell, 2003
- Regulatory Mechanisms Controlling Human Hepatocyte Nuclear Factor 4α Gene ExpressionMolecular and Cellular Biology, 2001
- Hepatocyte Nuclear Factor 4α (Nuclear Receptor 2A1) Is Essential for Maintenance of Hepatic Gene Expression and Lipid HomeostasisMolecular and Cellular Biology, 2001
- LightCycler technology for the quantitation of bcr/abl fusion transcripts.1999
- The maturity-onset diabetes of the young (MODY1) transcription factor HNF4α regulates expression of genes required for glucose transport and metabolismProceedings of the National Academy of Sciences of the United States of America, 1997