Transcriptional regulation of apolipoprotein C-III gene expression by the orphan nuclear receptor RORα

Triglyceride-rich remnant lipoproteins are considered as major risk factors contributing to the pathogenesis of atherosclerosis. Because apolipoprotein (apo) C-III is a major determinant of plasma triglyceride and remnant lipoprotein metabolism, it is important to understand how the expression of this gene is regulated. In the present study, we identified the orphan nuclear receptor ROR alpha1 as a regulator of human and mouse apo C-III gene expression. Plasma triglyceride and apo C-III protein concentrations in staggerer (sg/sg) mice, homozygous for a deletion in the ROR alpha gene, were significantly lower than in wild type littermates. The lowered plasma apo C-III levels were associated with reduced apo C-III mRNA levels in liver and intestine of sg/sg mice. Transient transfection experiments in human hepatoma HepG2, human colonic CaCO2, and rabbit kidney RK13 cells demonstrated that overexpression of the human ROR alpha1 isoform specifically increases human apo C-III promoter activity, indicating that ROR alpha1 enhances human apo C-III gene transcription. ROR alpha1 response elements were mapped by promoter deletion analysis and gel shift experiments to two AGGTCA halfsites located at positions -83/-78 (within the C3P site) and -23/-18 (downstream of the TATA box) in the human apo C-III promoter, with the -23/-18 site exhibiting the highest binding affinity. Transfection of site-directed mutated constructs in HepG2 cells indicated that the ROR alpha1 effect is predominantly mediated by the -23/-18 site. This site is conserved in the mouse apo C-III gene promoter. Moreover, ROR alpha binds to the equivalent mouse site and activates constructs containing three copies of the mouse site cloned in front of an heterologous promoter. Taken together, our data identify ROR alpha as a transcriptional regulator of apo C-III gene expression, providing a novel, physiological role for ROR alpha1 in the regulation of genes controlling triglyceride metabolism.