Sterol 12a-hydroxylase (CYP8B1) is involved in cholic acid synthesis and plays a role in intestinal cholesterol absorption and pathogenesis of cholesterol gallstone disease and dyslipidemia. In this study, we investigated the underlying mechanism of a fasting-induced and cholesterol activated nuclear receptor and core clock gene RORa in regulation of circadian rhythm and fasting induction of CYP8B1 expression. In free fed mice, CYP8B1 expression was reduced to the lowest level at the onset of the dark cycle when RORa expression was the lowest. However, fasting stimulated, while re-feeding reduced expression of CYP8B1 mRNA and protein expression. Interestingly, fasting and feeding had little effect on the diurnal rhythm of RORa mRNA expression, but fasting increased, whereas feeding decreased RORa protein levels in mouse liver. Adenovirus-mediated transduction of RORa to mice strongly induced CYP8B1 gene expression, increased 12a-hydroxylated bile acids in bile acid pool and serum and liver cholesterol. Reporter assay and mutagenesis analysis of the CYP8B1 promoter identified a functional RORa response element. Mammalian two-hybrid assay showed strong interaction of RORa with cAMP response element binding protein-binding protein (CBP). Chromatin immune-precipitation assay showed that RORa recruited CBP to the CYP8B1 promoter to stimulate histone acetylation. CAMP-activated protein kinase A phosphorylates RORa and increases its half-life. In conclusion, RORa is a key regulator of circadian expression and fasting induction of CYP8B1 to increase 12a-hydroxylated bile acids in the bile acid pool, and serum and liver cholesterol. This study contributes to our understanding of the molecular mechanism by which bile acid synthesis and composition regulates hepatic metabolic homeostasis. Overall this study sheds light on the mechanism of development of hypercholesterolemia in diabetic patients. Therefore, antagonizing RORa activity may be a therapeutic strategy for treating inflammatory diseases such as non-alcoholic fatty liver disease.
|Advisor:||Stroup, Diane, Chiang, John YL.|
|Commitee:||Diane, Stroup, John, Chiang|
|School:||Kent State University|
|School Location:||United States -- Ohio|
|Source:||DAI-B 75/08(E), Dissertation Abstracts International|
|Keywords:||Bile acid synthesis, CYP8B1, Cytochrome P450 family 8 subfamily B, polypeptide 1, Diurnal rhythm, Non alcoholic fatty liver disease, Rora|
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