Localization of a substrate specificity domain in the multidrug resistance protein.
Multidrug resistance protein (MRP) confers resistance to a number of natural product chemotherapeutic agents. It is also a high affinity transporter of some physiological conjugated organic anions such as cysteinyl leukotriene C(4) and the cholestatic estrogen, 17beta-estradiol 17(beta-D-glucuronide) (E(2)17betaG). We have shown that the murine orthologue of MRP (mrp), unlike the human protein, does not confer resistance to common anthracyclines and is a relatively poor transporter of E(2)17betaG. We have taken advantage of these functional differences to identify region(s) of MRP involved in mediating anthracycline resistance and E(2)17betaG transport by generating mrp/MRP hybrid proteins. All hybrid proteins conferred resistance to the Vinca alkaloid, vincristine, when transfected into human embryonic kidney cells. However, only those in which the COOH-terminal third of mrp had been replaced with the corresponding region of MRP-conferred resistance to the anthracyclines, doxorubicin, and epirubicin. Exchange of smaller segments of the COOH-terminal third of the mouse protein by replacement of either amino acids 959-1187 or 1188-1531 with those of MRP produced proteins capable of conferring some level of resistance to the anthracyclines tested. All hybrid proteins transported cysteinyl leukotriene C(4) with similar efficiencies. In contrast, only those containing the COOH-terminal third of MRP transported E(2)17betaG with an efficiency comparable with that of the intact human protein. The results demonstrate that differences in primary structure of the highly conserved COOH-terminal third of mrp and MRP are important determinants of the inability of the murine protein to confer anthracycline resistance and its relatively poor ability to transport E(2)17betaG.
Stride
B D
BD
Department of Biochemistry, Queen's University, Kingston K7L 3N6, Canada.
Cole
S P
SP
Deeley
R G
RG
D018528:ATP-Binding Cassette Transporters / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D018943:Anthracyclines / Q000494:pharmacology*; D001692:Biological Transport; D004317:Doxorubicin / Q000494:pharmacology; D018432:Drug Resistance, Multiple; D015251:Epirubicin / Q000494:pharmacology; D004958:Estradiol / Q000031:analogs & derivatives* / Q000378:metabolism; D006801:Humans; D017997:Leukotriene C4 / Q000378:metabolism; D051379:Mice; D008958:Models, Molecular; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D013045:Species Specificity; D013379:Substrate Specificity; D014750:Vincristine / Q000494:pharmacology
D018528:ATP-Binding Cassette Transporters; D018943:Anthracyclines; D004317:Doxorubicin; D015251:Epirubicin; D004958:Estradiol; D017997:Leukotriene C4; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D011993:Recombinant Fusion Proteins; D014750:Vincristine; C025483:estradiol-17 beta-glucuronide