Differential Modulation of the Ileal Bile Acid Transporter ASBT by Fatty Acids

The ileal ASBT bile acid transporter is a critical mediator of bile acid and cholesterol homeostasis and is modulated by fatty acids via S-acylation. We investigated mechanisms underlying differential regulation of ASBT by saturated and polyunsaturated fatty acids. Using a prenylation mimetic with a CAAX box at the C-terminus of ASBT, we showed that transport activity in transfected HEK293 cells increased significantly, suggesting that direct attachment of a saturated fatty acid stimulates ASBT function. Furthermore, siRNA-mediated attenuation of DHHC21 acyltransferase expression reduced ASBT S-acylation and its function, demonstrating DHHC21 as a positive regulator of ASBT function. Conversely, treatment with omega-3 polyunsaturated fatty acids (n-3 PUFAs), like eicosapentaenoic acid (EPA), reduced ASBT function in HEK293 cells. Supplementation of fish oil, as a source of n-3 PUFAs, to a high-fat, high-cholesterol diet fed to C57BL6/J mice protected them against diet-induced hepatic steatosis, along with reduced bile acid absorption consistent with the inhibition of ASBT. This was supported by increased fecal bile acids, decreased ileal bile acid-stimulated gene expression, and increased hepatic bile acid synthesis-related gene expression. Metabolic labeling with alkyne-EPA in HEK293 cells showed, for the first time, direct binding of this n-3 PUFA to ASBT. Unlike palmitate modification, EPA binding was highly resistant to hydroxylamine retrieval, indicating a mechanism distinct from S-acylation. Our data unraveled a novel role for DHHC21 in regulating ASBT and provided evidence for distinct mechanisms by which saturated palmitate and polyunsaturated EPA fatty acids bind ASBT. The direct effect on ASBT and inhibition of bile acid absorption may partly contribute to the protective effects of n-3 PUFAs on the liver.