Abstract
Chloroquine resistance in the malaria parasite Plasmodium falciparum has made malaria increasingly difficult to control. Chloroquine-resistant parasites accumulate less chloroquine than their chloroquine-sensitive counterparts; however, the mechanism underlying this remains unclear. The primary site of accumulation and antimalarial action of chloroquine is the internal acidic digestive vacuole of the parasite, the acidity of which is maintained by inwardly-directed H+ pumps, working against the (outward) leak of H+. In this study we have investigated the leak of H+ from the digestive vacuole of the parasite by monitoring the alkalinisation of the vacuole following inhibition of the H+-pumping V-type ATPase by concanamycin A. The rates of alkalinisation observed in three chloroquine-resistant strains were two- to fourfold higher than those measured in three chloroquine-sensitive strains. On addition of chloroquine there was a dramatic increase in the rate of alkalinisation in the chloroquine-resistant strains, whereas chloroquine caused the rate of alkalinisation to decrease in the chloroquine-sensitive strains. The chloroquine-associated increase in the rate of alkalinisation seen in chloroquine-resistant parasites was inhibited by the chloroquine-resistance reversal agent verapamil. The data are consistent with the hypothesis that in chloroquine-resistant parasites chloroquine effluxes from the digestive vacuole, in association with H+, via a verapamil-sensitive pathway.
Highlights
The emergence, followed by the spread throughout most malariaendemic regions, of Plasmodium falciparum parasites resistant to the antimalarial drug chloroquine (CQ) has worsened the global malaria situation (Trape, 2001)
CQ resistance is associated with a significant reduction in CQ accumulation by the parasite (Fitch, 1970; Krogstad et al, 1987; Yayon et al, 1984), and has been attributed to mutations in PfCRT (Fidock et al, 2000; Sidhu et al, 2002), a member of the ‘drug/metabolite transporter’ superfamily (Martin and Kirk, 2004; Tran and Saier, 2004) that is localised to the digestive vacuole (DV) membrane (Cooper et al, 2002; Fidock et al, 2000)
We have focused on the DV and present evidence that in CQR parasites the presence of CQ gives rise to a substantial outward H+ leak, consistent with
Summary
The emergence, followed by the spread throughout most malariaendemic regions, of Plasmodium falciparum parasites resistant to the antimalarial drug chloroquine (CQ) has worsened the global malaria situation (Trape, 2001). The DV localisation of PfCRT, together with the findings that the majority of the CQ accumulated by CQsensitive (CQS) parasites is within the DV (Bray et al, 2006), and that DVs isolated from CQ-resistant (CQR) parasites accumulate less CQ than those from CQS parasites (Saliba et al, 1998a), are consistent with the hypothesis that mutations in PfCRT cause CQ resistance by reducing the intravacuolar CQ concentration. How they do so is the subject of ongoing debate (Sanchez et al, 2007a)
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