FUNCTIONAL MULTIDRUG-RESISTANCE PROTEIN (MRP1) LACKING THE N-TERMINALTRANSMEMBRANE DOMAIN

The human multidrug resistance protein (MRP1) causes drug resistance b y extruding drugs from tumor cells. In addition to an MDR-like core, M RP1 contains an N-terminal membrane-bound region (TMD0) connected to t he core by a cytoplasmic linker (L-0). We have studied truncated MRP1 versions containing either the MDR-like core alone or the core plus li nker L-0, produced in the baculovirus-insect (Sf9) cell system. Their function was examined in isolated membrane vesicles. Full-length MRP1 showed ATP-dependent, vanadate-sensitive accumulation of leukotriene C -4 and N-ethylmaleimide glutathione. In addition, leukotriene C-4-stim ulated, vanadate-dependent nucleotide occlusion was detected. The MDR- like core was virtually inactive. Co-expression of the core with the N -terminal region including L-0 fully restored MRP1 function. Unexpecte dly, a truncated MRP1 mutant lacking the entire TMD, region but still containing L-0 behaved like wild-type MRP1 in vesicle uptake and nucle otide trapping experiments. We also expressed the MRP1 constructs in p olarized canine kidney derived MDCKII cells. Like wild-type MRP1, the MRP1 protein without the TMD, region was routed to the lateral plasma membrane and transported dinitrophenyl glutathione and daunorubicin, T he TMD0L0 and the MRP1 minus TMD0L0 remained in an intracellular compa rtment. Taken together, these experiments strongly suggest that the TM D0 region is neither required for the transport function of MRP1 nor f or its proper routing to the plasma membrane.