Functional Characterization of <em>ABCB4</em> Mutations Found in Low Phospholipid-Associated Cholelithiasis (LPAC)

Functional Characterization of <em>ABCB4</em> Mutations Found in Low Phospholipid-Associated Cholelithiasis (LPAC)

Multidrug resistance 3 (MDR3) is expressed on the canalicular membrane of the hepatocytes and plays an important role in protecting the liver from bile acids. Altered <em>ABCB4</em> gene expression can lead to a rare hepatic disease, low phospholipid-associated cholelithiasis (LPAC). In this study, we characterized 3 <em>ABCB4</em> mutations in LPAC patients using various in vitro assay systems. We first measured the ability of each mutant to transport paclitaxel and then the mechanisms by which these mutations might change MDR3 transport activity were determined using immunoblotting, cell surface protein biotinylation, and immunofluorescence. Through a membrane vesicular transport assay, we observed that the uptake of paclitaxel was significantly reduced in membrane vesicles expressing 2 <em>ABCB4</em> mutations, F165I and S320F. Both mutants showed significantly decreased total and cell surface MDR3 expression. These data suggest two missense mutations of <em>ABCB4</em> may alter function of MDR3 and ultimately can be determined as LPAC-causing mutations.