Functional Characterization of the Human Multidrug Transporter, ABCG2, Expressed in Insect Cells

doi:10.1006/bbrc.2001.5130

Biochemical and Biophysical Research Communications

Volume 285, Issue 1, 6 July 2001, Pages 111-117

https://doi.org/10.1006/bbrc.2001.5130 Get rights and content

Abstract

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.

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^☆: Abbreviations used: MXR, mitoxantrone resistance-associated protein; BCRP, breast cancer resistance protein; ABCP, placenta specific ABC transporter; ABC, ATP binding cassette; Sf9 cells, Spodoptera frugiperda ovarian cells; MDR1, multidrug resistance protein; MRP1, multidrug resistance-associated protein; TMD, transmembrane domain; TAP, transporter associated with antigen processing; Calcein-AM, calcein acetoxy-methylesther; NEM, N-ethylmaleimide; MX, mitoxantrone; FTC, fumitremorgin C; CsA, cyclosporin A.

¹: These authors contributed equally to this work.

²: To whom correspondence should be addressed. Fax: (36-1) 372-4353. E-mail: [email protected].

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Regular ArticleFunctional Characterization of the Human Multidrug Transporter, ABCG2, Expressed in Insect Cells☆

Abstract

Biochim. Biophys. Acta

Biochem. Pharmacol.

Blood

Biochem. Pharmacol.

Biochim. Biophys. Acta

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

Biochemical, cellular, and pharmacological aspects of the multidrug transporter

Annu. Rev. Pharmacol. Toxicol.

Molecular cloning of cDNAs which are highly overexpressed in mitoxantrone-resistant cells: Demonstration of homology to ABC transport genes

Cancer Res.

A multidrug resistance transporter from human MCF-7 breast cancer cells

Proc. Natl. Acad. Sci. USA

A human placenta-specific ATP-binding cassette gene (ABCP) on chromosome 4q22 that is involved in multidrug resistance

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Regular Article
Functional Characterization of the Human Multidrug Transporter, ABCG2, Expressed in Insect Cells☆