Analysis of the Bile Salt Export Pump (ABCB11) Interactome Employing Complementary Approaches

• Published in: PloS one 2016-07, Vol.11 (7), p.e0159778-e0159778
• Main Authors: Przybylla, Susanne, Stindt, Jan, Kleinschrodt, Diana, Schulte Am Esch, Jan, Häussinger, Dieter, Keitel, Verena, Smits, Sander H, Schmitt, Lutz
• Format: Article
• Language: English
• Subjects: ATP Binding Cassette Subfamily B Member 11; ATP-Binding Cassette Transporters - genetics; ATP-Binding Cassette Transporters - metabolism; Baking yeast; Bile; Biological transport; Biology and Life Sciences; Cell Line; Cholestasis; DNA, Complementary - genetics; Exports; Hepatocytes; Humans; Liver; Localization; Mass spectrometry; Mass spectroscopy; Medicine and Health Sciences; Membrane trafficking; Physical Sciences; Physiological aspects; Plasmids; Protein Binding; Proteins; Radixin; Research and Analysis Methods; Rodents; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Salts; Two-Hybrid System Techniques
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0159778

The bile salt export pump (BSEP, ABCB11) plays an essential role in the formation of bile. In hepatocytes, BSEP is localized within the apical (canalicular) membrane and a deficiency of canalicular BSEP function is associated with severe forms of cholestasis. Regulation of correct trafficking to the canalicular membrane and of activity is essential to ensure BSEP functionality and thus normal bile flow. However, little is known about the identity of interaction partners regulating function and localization of BSEP. In our study, interaction partners of BSEP were identified in a complementary approach: Firstly, BSEP interaction partners were co-immunoprecipitated from human liver samples and identified by mass spectrometry (MS). Secondly, a membrane yeast two-hybrid (MYTH) assay was used to determine protein interaction partners using a human liver cDNA library. A selection of interaction partners identified both by MYTH and MS were verified by in vitro interaction studies using purified proteins. By these complementary approaches, a set of ten novel BSEP interaction partners was identified. With the exception of radixin, all other interaction partners were integral or membrane-associated proteins including proteins of the early secretory pathway and the bile acyl-CoA synthetase, the second to last, ER-associated enzyme of bile salt synthesis.