Functional characterization of the human multidrug transporter, ABCG2, expressed in insect cells.
ABCG2 (also called MXR (3), BCRP (4), or ABCP (5) is a recently-identified ABC half-transporter, which causes multidrug resistance in cancer. Here we report that the expression of the ABCG2 protein in Sf9 insect cells resulted in a high-capacity, vanadate-sensitive ATPase activity in isolated membrane preparations. ABCG2 was expressed underglycosylated, and its ATPase activity was stimulated by daunorubicin, doxorubicin, mitoxantrone, prazosin and rhodamine 123, compounds known to be transported by this protein. ABCG2-ATPase was inhibited by low concentrations of Na-orthovanadate, N-ethylmaleimide and cyclosporin A. Verapamil had no effect, while Fumitremorgin C, reversing ABCG2-dependent cancer drug resistance, strongly inhibited this ATPase activity. The functional expression of ABCG2 in this heterologous system indicates that no additional partner protein is required for the activity of this multidrug transporter, probably working as a homodimer. We suggest that the Sf9 cell membrane ATPase system is an efficient tool for examining the interactions of ABCG2 with pharmacological agents.
Ozvegy
C
C
Institute of Enzymology, Hungarian Academy of Sciences, Budapest, H-1113, Hungary.
Litman
T
T
Szakács
G
G
Nagy
Z
Z
Bates
S
S
Váradi
A
A
Sarkadi
B
B
D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000502:physiology*; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D001943:Breast Neoplasms / Q000473:pathology; D002462:Cell Membrane / Q000201:enzymology; D003001:Cloning, Molecular; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D006801:Humans; D009363:Neoplasm Proteins; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D018411:Spodoptera; D014407:Tumor Cells, Cultured
C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D009363:Neoplasm Proteins; D011994:Recombinant Proteins; D000251:Adenosine Triphosphatases