Abstract
Methotrexate (MTX) is a widely used chemotherapeutic agent, immune suppressant and antimalarial drug. It is a substrate of several human ABC proteins that confer multidrug resistance to cancer cells and determine compartmentalization of a wide range of physiological metabolites and endo or xenobiotics, by their primary active transport across biological membranes. The substrate specificity and tissue distribution of these promiscuous human ABC transporters show a high degree of redundancy, providing robustness to these key physiological and pharmacological processes, such as the elimination of toxins, e.g. methotrexate from the body. A similar network of proteins capable of transporting methotrexate has been recently suggested to exist in Drosophila melanogaster. One of the key players of this putative network is Drosophila Multidrug-resistance Associated Protein (DMRP). DMRP has been shown to be a highly active and promiscuous ABC transporter, capable of transporting various organic anions. Here we provide the first direct evidence that DMRP, expressed alone in a heterologous system lacking other, potentially functionally overlapping D. melanogaster organic anion transporters, is indeed able to transport methotrexate. Our in vitro results support the hypothesized but debated role of DMRP in in vivo methotrexate excretion.
Highlights
Adenosine triphosphate (ATP) binding cassette (ABC) proteins are considered as one of the most abundant protein families, existing in all phyla of life with 49 members in human and 56 in Drosophila melanogaster [1,2,3]
To test the capability of Drosophila Multidrug-resistance Associated Protein (DMRP) to transport methotrexate (MTX), we first investigated its effect on the DMRP mediated hydrolysis of ATP
We investigated the time course of the ATPase activity in the absence and in the presence of 1mM MTX in inside-out vesicles prepared from Sf9 cells overexpressing wild type DMRP (Fig 1A)
Summary
Adenosine triphosphate (ATP) binding cassette (ABC) proteins are considered as one of the most abundant protein families, existing in all phyla of life with 49 members in human and 56 in Drosophila melanogaster [1,2,3]. Most of the ABC proteins are membrane resident solute transporters, the eukaryotic transporters being exclusively exporters [1]. Most of the eukaryotic ABC transporters export a limited number of closely related molecules, but there are a few transporters with an extremely wide spectrum of structurally unrelated substrates. Some of these promiscuous ABC transporters are involved in the multidrug resistance phenotype of cancer cells and are the major obstacle of cancer chemotherapy [4].
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