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LIVER AND BILIARY TRACT

Protein-protein interactions and membrane localization of the human organic solute transporter

¹Department of Pediatrics, Mount Sinai School of Medicine, and ²Department of Orthopaedic Surgery, New York University, New York, New York

Submitted 3 October 2006 ; accepted in final form 23 February 2007

Two proteins that mediate bile acid export from the ileal enterocyte, organic solute transporter (OST)- and -, have recently been identified. It is unclear whether these two proteins associate directly and how they interact to mediate transport function and membrane localization. In this study, the protein-protein interactions, transport functions, and membrane localization of human (h)OST- and - proteins were examined. The results demonstrated that coexpression of hOST- and - in transfected cells resulted in a three- to fivefold increase of the initial rate of taurocholate influx or efflux compared with cells expressing each protein individually and nontransfected cells. Confocal microscopy demonstrated plasma membrane colocalization of hOST- and - proteins in cells cotransfected with hOST- and - cDNAs. Protein-protein interactions between hOST- and - were demonstrated by mammalian two-hybrid and coimmunoprecipitation analyses. Truncation of the amino-terminal 50 amino acid extracellular residues of hOST- abolished its interaction with hOST- and led to an intracellular accumulation of the two proteins and to only background levels of taurocholate transport. In contrast, carboxyl-terminal 28 amino acid truncated hOST- still interacted with hOST-, and majority of this cytoplasmic tail-truncated protein was expressed on the basolateral membrane when it was stably cotransfected with hOST- protein in Madin-Darby canine kidney cells. In summary, hOST- and - proteins are physically associated. The intracellular carboxyl-terminal domain of hOST- is not essential for this interaction with hOST-. The extracellular amino-terminal fragment of hOST- may contain important information for the assembly of the heterodimer and trafficking to the plasma membrane.

bile acid transporter