RESUMEN
Mutations in the Plasmodium falciparum chloroquine (CQ) resistance transporter (PfCRT) are major determinants of verapamil (VP)-reversible CQ resistance (CQR). In the presence of mutant PfCRT, additional genes contribute to the wide range of CQ susceptibilities observed. It is not known if these genes influence mechanisms of chemosensitization by CQR reversal agents. Using quantitative trait locus (QTL) mapping of progeny clones from the HB3 × Dd2 cross, we show that the P. falciparum multidrug resistance gene 1 (pfmdr1) interacts with the South-East Asia-derived mutant pfcrt haplotype to modulate CQR levels. A novel chromosome 7 locus is predicted to contribute with the pfcrt and pfmdr1 loci to influence CQR levels. Chemoreversal via a wide range of chemical structures operates through a direct pfcrt-based mechanism. Direct inhibition of parasite growth by these reversal agents is influenced by pfcrt mutations and additional loci. Direct labelling of purified recombinant PfMDR1 protein with a highly specific photoaffinity CQ analogue, and lack of competition for photolabelling by VP, supports our QTL predictions. We find no evidence that pfmdr1 copy number affects CQ response in the progeny; however, inheritance patterns indicate that an allele-specific interaction between pfmdr1 and pfcrt is part of the complex genetic background of CQR.
Asunto(s)
Cloroquina/farmacología , Plasmodium falciparum/efectos de los fármacos , Plasmodium falciparum/genética , Antimaláricos/farmacología , Resistencia a Medicamentos , Dosificación de Gen , Proteínas de Transporte de Membrana/genética , Proteínas de Transporte de Membrana/metabolismo , Mutación , Plasmodium falciparum/metabolismo , Proteínas Protozoarias/genética , Proteínas Protozoarias/metabolismo , Sitios de Carácter Cuantitativo , Verapamilo/farmacologíaRESUMEN
We utilize the recent successful overexpression of recombinant Plasmodium falciparum multi-drug resistance transporter, purification and reconstitution of the protein, and a novel high affinity chloroquine analogue to probe hypothesized interaction between the transporter and quinoline drugs. Results suggest that PfMDR1 binding sites for chloroquine, mefloquine, and quinine overlap, that P. falciparum chloroquine resistance transporter has intrinsically higher affinity for chloroquine relative to P. falciparum multi-drug resistance transporter, and that there is an isoform specific competition between the two transporters for binding of quinoline antimalarial drugs.