Abstract
Electrophoretic mobility shift assay (EMSA) is an invaluable tool to study interaction of proteins with DNA. Estrogens are major female hormones and modulate biological function through estrogen receptor (ER). ER regulates its target gene expression via the classical mechanism in which ER directly binds to its target gene promoter or the nonclassical mechanism involving tethering of ER to other transcription factors (such as AP-1 proteins). Here, we describe the EMSA to examine the nonclassical mechanism of ER action in regulation of a gene CYP2B6 by using competition and supershift assays.
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References
Fried M, Crothers DM (1981) Equilibria and kinetics of lac repressor-operator interactions by polyacrylamide gel electrophoresis. Nucleic Acids Res 9(23):6505–6525
Koh KH, Jurkovic S, Yang K et al (2012) Estradiol induces cytochrome P450 2B6 expression at high concentrations: implication in estrogen-mediated gene regulation in pregnancy. Biochem Pharmacol 84(1):93–103
Pedram A, Razandi M, O'Mahony F, Harvey H, Harvey BJ, Levin ER (2013) Estrogen reduces lipid content in the liver exclusively from membrane receptor signaling. Sci Signal 6(276):ra36
Bjornstrom L, Sjoberg M (2005) Mechanisms of estrogen receptor signaling: convergence of genomic and nongenomic actions on target genes. Mol Endocrinol 19(4):833–842
Safe S, Kim K (2008) Non-classical genomic estrogen receptor (ER)/specificity protein and ER/activating protein-1 signaling pathways. J Mol Endocrinol 41(5):263–275
Larouche K, Bergeron MJ, Leclerc S, Guerin SL (1996) Optimization of competitor poly(dI-dC).poly(dI-dC) levels is advised in DNA-protein interaction studies involving enriched nuclear proteins. Biotechniques 20(3):439–444
Lambertini E, Tavanti E, Torreggiani E, Penolazzi L, Gambari R, Piva R (2008) ERalpha and AP-1 interact in vivo with a specific sequence of the F promoter of the human ERalpha gene in osteoblasts. J Cell Physiol 216(1):101–110
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Koh, K.H., Jeong, H. (2016). Electrophoretic Mobility Shift Assay (EMSA) and Supershift Assay of Cytochrome P450 2B6 in Response to Estrogen. In: Eyster, K.M. (eds) Estrogen Receptors. Methods in Molecular Biology, vol 1366. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3127-9_5
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DOI: https://doi.org/10.1007/978-1-4939-3127-9_5
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3126-2
Online ISBN: 978-1-4939-3127-9
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