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Nucleotide‐induced conformational changes in the human multidrug resistance protein MRP1 are related to the capacity of chemotherapeutic drugs to accumulate or not in resistant cells
Author(s) -
Manciu Liliana,
Chang Xiu-Bao,
Riordan John R.,
Buyse Frédéric,
Ruysschaert Jean Marie
Publication year - 2001
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(01)02270-0
Subject(s) - daunorubicin , hek 293 cells , doxorubicin , multiple drug resistance , chemistry , anthracycline , intracellular , atp binding cassette transporter , efflux , biochemistry , conformational change , transporter , biology , receptor , genetics , chemotherapy , cancer , leukemia , breast cancer , gene , antibiotics
Intracellular accumulation of anthracycline derivatives was measured in a human embryonic kidney cell line (HEK) and a resistant subline (HEK/multidrug resistance protein (MRP1)) overexpressing MRP1 at the plasma membrane surface. Two compounds (daunorubicin and doxorubicin) were rejected outside the multidrug‐resistant cells. On the contrary, three compounds (4′‐deoxy‐4′‐iodo‐doxorubicin, 4‐demethoxy‐daunorubicin and 3′‐(3‐methoxymorpholino)doxorubicin) accumulated equally within sensitive HEK cells and resistant HEK/MRP1 cells. Our main objective here was to characterize the MRP1 conformational changes mediated by the binding of these anthracycline derivatives and to determine whether these conformational changes are related to MRP1‐mediated drug transport. MRP1 was reconstituted in lipid vesicles as previously described [Manciu, L., Chang, X.B., Riordan, J.R. and Ruysschaert, J.‐M. (2000) Biochemistry 39, 13026–13033]. The reconstituted protein was shown to conserve its ATPase and drug transport activity. Acrylamide quenching of Trp fluorescence was used to monitor drug‐dependent conformational changes. Binding of drugs (4‐demethoxy‐daunorubicin and 3′‐(3‐methoxymorpholino)doxorubicin) which accumulate in resistant cells immobilizes MRP1 in a conformational state that is insensitive to ATP binding whereas drugs rejected outside the resistant cells (daunorubicin, doxorubicin) favor a conformational change which may be a required step in the transport process.