L-FABP directly interacts with PPARα in cultured primary hepatocytes

作者：Heather Ann Hostetler, Avery L. McIntosh, Barbara P. Atshaves, Stephen M. Storey, Harold Ross Payne, Ann B. Kier, Friedhelm Schroeder · 发表于：Journal of Lipid Research · 年份：2009 · DOI：10.1194/jlr.m900058-jlr200 · 被引用次数：150 · 研究领域：Peroxisome Proliferator-Activated Receptors、Drug Transport and Resistance Mechanisms、Cholesterol and Lipid Metabolism

Although studies with liver type fatty acid binding protein (L-FABP) gene ablated mice demonstrate a physiological role for L-FABP in hepatic fatty acid metabolism, little is known about the mechanisms whereby L-FABP elicits these effects. Studies indicate that L-FABP may function to shuttle lipids to the nucleus, thereby increasing the availability of ligands of nuclear receptors, such as peroxisome proliferator-activated receptor-alpha (PPARalpha). The data herein suggest that such mechanisms involve direct interaction of L-FABP with PPARalpha. L-FABP was shown to directly interact with PPARalpha in vitro through co-immunoprecipitation (co-IP) of pure proteins, altered circular dichroic (CD) spectra, and altered fluorescence spectra. In vitro fluorescence resonance energy transfer (FRET) between Cy3-labeled PPARalpha and Cy5-labeled L-FABP proteins showed that these proteins bound with high affinity (Kd approximately 156 nM) and in close proximity (intermolecular distance of 52A). This interaction was further substantiated by co-IP of both proteins from liver homogenates of wild-type mice. Moreover, double immunogold electron microscopy and FRET confocal microscopy of cultured primary hepatocytes showed that L-FABP was in close proximity to PPARalpha (intermolecular distance 40-49A) in vivo. Taken together, these studies were consistent with L-FABP regulating PPARalpha transcriptional activity in hepatocytes through direct interaction with PPARalpha. Our in vitro and imagin...