No abstract

The ATP binding cassette (ABC) half-transporters ABCG5 and ABCG8 facilitate biliary and intestinal removal of neutral sterols. Here, we identify a binding site for the orphan nuclear receptor liver receptor homolog-1 (LRH-1) at nt 134-142 of the ABCG5/ABCG8 intergenic region necessary for the activity of both the ABCG5 and ABCG8 promoters. Mutating this LRH-1 binding site reduced promoter activity of the human ABCG5/ABCG8 intergenic region more than 7-fold in HepG2 and Caco2 cells. Electrophoretic mobility shift assays with HepG2 nuclear extracts demonstrated specific binding of LRH-1 to the LRH-1 binding motif in the human ABCG5/ABCG8 intergenic region. LRH-1 overexpression increased promoter activity up to 1.6-fold and 3-fold in Caco2 and 293 cells, respectively. Finally, deoxycholic acid repressed the ABCG5 and ABCG8 promoters, consistent with bile acid regulation via the farnesoid X receptor-small heterodimeric partner-LRH-1 pathway. These results demonstrate that LRH-1 is a positive transcription factor for ABCG5 and ABCG8 and, in conjunction with studies on LRH-1 activation of other promoters, identify LRH-1 as a "master regulator" for genes involved in sterol and bile acid secretion from liver and intestine. -Freeman, L. A., A. Kennedy, J. Wu, S. Bark, A. T. Remaley, S. Santamarina-Fojo, and H. B. Brewer, Jr. The orphan nuclear receptor LRH-1 activates the ABCG5/ABCG8 intergenic promoter.

The individual roles of hepatic versus intestinal ABCG5 and ABCG8 in sterol transport have not yet been investigated. To determine the specific contribution of liver ABCG5/G8 to sterol transport and atherosclerosis, we generated transgenic mice that overexpress human ABCG5 and ABCG8 in the liver but not intestine (liver G5/G8-Tg) in three different genetic backgrounds: C57Bl/6, apoE-KO, and low density lipoprotein receptor (LDLr)-KO. Hepatic overexpression of ABCG5/G8 enhanced hepatobiliary secretion of cholesterol and plant sterols by 1.5-2-fold, increased the amount of intestinal cholesterol available for absorption and fecal excretion by up to 27%, and decreased the accumulation of plant sterols in plasma by ϳ25%. However, it did not alter fractional intestinal cholesterol absorption, fecal neutral sterol excretion, hepatic cholesterol concentrations, or hepatic cholesterol synthesis. Consequently, overexpression of ABCG5/G8 in only the liver had no effect on the plasma lipid profile, including cholesterol, HDL-C, and non-HDL-C, or on the development of proximal aortic atherosclerosis in C57Bl/6, apoE-KO, or LDLr-KO mice. Thus, liver ABCG5/G8 facilitate the secretion of liver sterols into bile and serve as an alternative mechanism, independent of intestinal ABCG5/G8, to protect against the accumulation of dietary plant sterols in plasma. However, in the absence of changes in fractional intestinal cholesterol absorption, increased secretion of sterols into bile induced by hepatic overexpression of ABCG5/G8 was not sufficient to alter hepatic cholesterol balance, enhance cholesterol removal from the body or to alter atherogenic risk in liver G5/G8-Tg mice. These findings demonstrate that overexpression of ABCG5/G8 in the liver profoundly alters hepatic but not intestinal sterol transport, identifying distinct roles for liver and intestinal ABCG5/G8 in modulating sterol metabolism.Sitosterolemia, also known as phytosterolemia (1-9) is a rare genetic disorder characterized by elevated plasma and tissue levels of plant, shellfish, and animal sterols and the development of tendon and tuberous xanthomas, hemolytic episodes, arthritis, and premature coronary artery disease. The ATPbinding cassette (ABC) 1 half-transporters ABCG5 and ABCG8 (ABCG5/G8) have been recently identified as the genes defective in sitosterolemia (10 -12). The increased plasma and tissue levels of animal and plant sterols found in sitosterolemic patients have been attributed to enhanced absorption and decreased biliary excretion (1-4, 6, 8, 13-16). Unlike cholesterol, sitosterol accumulation does not inhibit HMGCoA reductase activity in human monocyte-derived macrophages (17). The inadequate down-regulation of HMGCoA reductase activity by plant sterols may promote foam cell formation and explain, in part, the increased risk of atherosclerosis in sitosterolemia (17).Several lines of evidence implicate ABCG5/G8 in intestinal and biliary sterol transport. First, ABCG5/G8 are expressed in the liver and small intestine and, to a lesser degree, in th...