Expression and function of pvcrt-o, a Plasmodium vivax ortholog of pfcrt, in Plasmodium falciparum and Dictyostelium discoideum.
Chloroquine resistance in Plasmodium vivax threatens the use of this drug as first-line treatment for millions of people infected each year worldwide. Unlike Plasmodium falciparum, in which chloroquine resistance is associated with mutations in the pfcrt gene encoding a digestive vacuole transmembrane protein, no point mutations have been associated with chloroquine resistance in the P. vivax ortholog gene, pvcrt-o (also called pvcg10). However, the question remains whether pvcrt-o can affect chloroquine response independent of mutations. Since P. vivax cannot be cultured in vitro, we used two heterologous expression systems to address this question. Results from the first system, in which chloroquine sensitive P. falciparum parasites were transformed with pvcrt-o, showed a 2.2-fold increase in chloroquine tolerance with pvcrt-o expression under a strong promoter; this effect was reversed by verapamil. In the second system, wild type pvcrt-o or a mutated form of the gene was expressed in Dictyostelium discoideum. Forms of PvCRT-o engineered to express either lysine or threonine at position 76 produced a verapamil-reversible reduction of chloroquine accumulation in this system to approximately 60% of that in control cells. Our data support an effect of PvCRT-o on chloroquine transport and/or accumulation by P. vivax, independent of the K76T amino acid substitution.
Sá
Juliana Martha
JM
Departamento de Parasitologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, Av. Lineu Prestes 1374, São Paulo 05508-900, SP, Brazil.
Yamamoto
Marcio M
MM
Fernandez-Becerra
Carmen
C
de Azevedo
Mauro Ferreira
MF
Papakrivos
Janni
J
Naudé
Bronwen
B
Wellems
Thomas E
TE
Del Portillo
Hernando A
HA
D000818:Animals; D000962:Antimalarials / Q000494:pharmacology*; D002738:Chloroquine / Q000378:metabolism / Q000494:pharmacology*; D004023:Dictyostelium / Q000737:chemistry / Q000187:drug effects / Q000235:genetics*; D004351:Drug Resistance; D011992:Endosomes / Q000378:metabolism; D004912:Erythrocytes / Q000469:parasitology; D015870:Gene Expression; D017125:Genes, Protozoan; D006801:Humans; D020128:Inhibitory Concentration 50; D026901:Membrane Transport Proteins / Q000032:analysis / Q000235:genetics; D010963:Plasmodium falciparum / Q000737:chemistry / Q000187:drug effects / Q000235:genetics*; D010966:Plasmodium vivax / Q000235:genetics*; D011401:Promoter Regions, Genetic; D015800:Protozoan Proteins / Q000032:analysis / Q000235:genetics*; D014162:Transfection; D014617:Vacuoles / Q000737:chemistry
D000962:Antimalarials; D026901:Membrane Transport Proteins; C416369:PfCRT protein, Plasmodium falciparum; D015800:Protozoan Proteins; D002738:Chloroquine