ATP-binding cassette transporter isoform C2 localizes to the apical plasma membrane via interactions with scaffolding protein.
ATP-binding cassette transporter isoform C2 (ABCC2) localizes to the apical plasma membrane in polarized cells. Apical localization of ABCC2 in hepatocytes plays an important role in biliary excretion of endobiotics and xenobiotics, but the mechanism by which ABCC2 localizes to the apical membrane has not been conclusively elucidated. Here, we investigate the role of scaffolding proteins on ABCC2 localization with a focus on the function of PDZK1 (post-synaptic density 95/disk large/zonula occludens-1 domain containing 1) in regulating ABCC2 localization. The C-terminal 77 residues of ABCC2 were used to probe interacting proteins from HepG2 cells. Protein mass fingerprinting identified PDZK1 as a major interacting protein. PDZK1 associated with the plasma membrane, most likely at the apical vacuoles of HepG2 cells. Affinity pull-down assays confirmed that the C-terminal NSTKF of ABCC2 bound to the fourth PDZ domain of PDZK1. Removal of this PDZ-binding motif significantly reduced the normal apical localization of ABCC2. In HepG2 cells, overexpression of this fourth domain overcame endogenous PDZK1 and reduced the ABCC2 localization at the apical membrane with a reciprocal increase of intracellular accumulation of mislocalized ABCC2. These results suggest a possible role for an interaction between ABCC2 and PDZK1 in apical localization of ABCC2 in hepatocytes.
Graduate School of Life Science, University of Hyogo, Harima Science Park City, Hyogo, Japan. emys@sci.u-hyogo.ac.jp
Yoshikazu
Y
Emi
Sachiko
S
Nomura
Hiroshi
H
Yokota
Masao
M
Sakaguchi
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism; D016764:Cell Polarity / Q000502:physiology; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D054731:PDZ Domains; D010449:Peptide Mapping; D011485:Protein Binding; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D002352:Carrier Proteins; D008565:Membrane Proteins; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D020033:Protein Isoforms; D011994:Recombinant Proteins; C000715087:ABCC2 protein, human; C484659:PDZK1 protein, human