Plasmodium falciparum expresses a multidrug resistance-associated protein Antonios Klokouzas a , Teresa Tiffert b , Donelly van Schalkwyk c , Chung-Pu Wu a , Hendrik W. van Veen a , Margery A. Barrand a , Stephen B. Hladky a, * a Department of Pharmacology, University of Cambridge, Cambridge, CB2 1PD, UK b Department of Physiology, University of Cambridge, Cambridge, CB2 3EG, UK c Department of Pharmacology, University of Cape Town, Observatory 7925, South Africa Received 27 May 2004 Abstract Plasmodium falciparum proteins that efflux toxic metabolic products such as oxidised glutathione (GSSG) are possible targets for anti-malarial drug development. Proteins capable of transporting GSSG and glutathione conjugates include the multidrug resis- tance-associated transporters (MRPs). A gene, PFA0590w, encoding a MRP homologue, has been identified in P. falciparum. Here we show the presence of full-length mRNA (5.5 kb) of this PfMRP in trophozoites by RT-PCR and Northern blotting. A polyclonal anti-PfMRP antibody generated against two unique, hydrophilic peptides in the predicted sequence produced a strong immunore- active protein band of 210–215 kDa on Western blots of schizonts of chloroquine-sensitive and chloroquine-resistant strains, con- firming expression of PfMRP protein. Using confocal microscopy the protein was seen to be localised at the edge of the schizonts with no obvious staining of the food vacuole. We suggest that PfMRP may act as the GSSG transporter in the parasite plasma membrane. Ó 2004 Published by Elsevier Inc. Keywords: Plasmodium falciparum; Drug resistance; ATP-binding cassette transporter; Multidrug resistance-associated protein; Glutathione The identification of potential drug targets in Plasmo- dium falciparum with the aim of developing an effective therapy for the treatment of malaria is one of the key objectives of the Malaria Genome Sequencing Project (see http://www.plasmodb.org). P. falciparum transport proteins that regulate the movement of toxic metabolic products such as oxidised glutathione (GSSG) [1] out of the parasite are one possible target for anti-malarial drug development. In all organisms studied, ATP-bind- ing cassette (ABC) transporters translocate a wide range of molecules across membranes against a concentration gradient [2–6]. Among these, multidrug resistance-asso- ciated (MRP or ABCC) transporters primarily function as organic anion transporters [4] with substrates that include GSSG and glutathione conjugates. At least nine genes containing both an ATP binding cassette and at least one predicted transmembrane span- ning domain have been identified in P. falciparum [7,8]. At present PfMDR1, the protein encoded by the PFE1150w (aka Pfmdr1) gene (NC_004326), is the only one known to be involved in drug resistance [5,9]. PfMDR1 is a homologue of the human P-glycoprotein. The genome of P. falciparum contains another gene, PFA0590w (NC_004325), which has been annotated as a 1822 amino acid full-length ABC transporter (CAB63558). We have previously noted that his putative protein appears to be an MRP homologue [10] here called PfMRP. It is predicted to contain two nucleotide 0006-291X/$ - see front matter Ó 2004 Published by Elsevier Inc. doi:10.1016/j.bbrc.2004.06.135 * Corresponding author. Fax: +44-1223-334040. E-mail address: sbh1@cam.ac.uk (S.B. Hladky). www.elsevier.com/locate/ybbrc Biochemical and Biophysical Research Communications 321 (2004) 197–201 BBRC