Chemical activation of a high-affinity glutamate transporter in human erythrocytes and its implications for malaria-parasite–induced glutamate uptake

Abstract

IntroductionIt has long been recognized that after infection of the human erythrocyte by the malaria parasite Plasmodium falciparum, the host cell membrane undergoes a profound increase in its permeability to a diverse range of low molecular weight solutes including amino acids, sugars, nucleosides, and a range of organic and inorganic anions and cations. [1][2][3][4][5] The increased permeability facilitates the uptake into the infected cell of several key nutrients, 6-8 as well as the uptake of various antiparasitic agents. [8][9][10][11][12] Parasiteinduced pathways also mediate both the efflux of metabolic wastes, 6,13 and a marked change in the ionic composition of the host cell cytosol. 4,14 The question of the number and origin of the pathways underpinning the parasite-induced increase in host erythrocyte permeability has been a vexed one. Subjecting uninfected human erythrocytes to oxidative stress, 15,16 or to protein kinase Amediated protein phosphorylation, [17][18][19] results in the activation of membrane channels that confer on the membrane of the uninfected cell, similar to permeability characteristics seen in the infected cell. This is consistent with at least some part of the parasite-induced permeability being attributed to the activation of endogenous host cell proteins. A recent paper has proposed a central role for an endogenous erythrocyte benzodiazopene receptor/voltagedependent anion channel 20 in forming the parasite-induced pathways. At the same time, there is evidence for the involvement of parasite-encoded proteins. After earlier observations of significant parasite-strain-specific differences in the electrophysiologic characteristics of parasite-induced ion conductances 21,22 and the identification of parasite strains for which the infected erythrocyte has reduced permeability to various cytotoxic agents, 10,11,23 Desai and colleagues have demonstrated a role for parasite-encoded "clag3" proteins in the formation of parasite-induced channels. 24 Whether these proteins themselves form part of the channels is not yet clear.Exposure of cells to arsenite is known both to induce oxidative stress and to perturb intracellular phosphorylation/dephosphorylation pathways. 25 In a study of ferret erythrocytes, Flatman and Creanor demonstrated that treatment of these cells with arsenite resulted in the stimulation of an ion transporter, the Na ϩ K ϩ 2Cl Ϫ cotransporter. 26 In this work, we investigated the effect of arsenite treatment on the transport into human erythrocytes of the acidic amino acid, glutamate. Human erythrocytes have a low basal permeability to glutamate; however, the amino acid readily enters P falciparum-infected erythrocytes. 1,2,27,28 Here we show that arsenite-treatment of (uninfected) human erythrocytes results in the activation of a high-affinity, Na ϩ -dependent glutamate transport pathway, which has the functional characteristics of members of the "excitatory amino acid transporter" (EAAT) family. 29,30 An analysis of the uptake of glutamate into P falciparum-infected ...

Sequestration and metabolism of host cell arginine by the intraerythrocytic malaria parasitePlasmodium falciparum

“…For both neutral (Martin and Kirk, ; Cobbold et al ., ) and acidic (Kirk et al ., ; Winterberg et al ., ) amino acids, the new permeability pathways (NPPs) induced by the parasite in the erythrocyte membrane serve as a major route of entry into the infected cell. However, the NPP inhibitor furosemide had no significant effect on the initial rate of [ 14 C]arginine uptake into iRBCs (Fig.…”

Solute Restriction Reveals an Essential Role forclag3-Associated Channels in Malaria Parasite Nutrient Acquisition

“…Hypoxanthine, a purine source that is not present in RPMI 1640 medium but is commonly added to parasite culture media, was also evaluated. Each of these solutes has documented PSAC permea- (Ginsburg et al, 1985;Gero and Wood, 1991;Asahi et al, 1996;Saliba et al, 1998;Lisk et al, 2006), but some also exhibit uptake by host erythrocyte transporters (Quashie et al, 2010;Winterberg et al, 2012). Microscopic examination of parasites propagated in these single-depletion experiments revealed the greatest detriment with removal of glutamine and hypoxanthine, but short-term growth studies revealed relatively modest effects (Fig.…”

Mice Deficient in the Putative Phospholipid Flippase ATP11C Exhibit Altered Erythrocyte Shape, Anemia, and Reduced Erythrocyte Life Span*

“…Cation Measurement-Intra-and extracellular Na ϩ and K ϩ levels were determined by ultra HPLC (Dionex) linked to a charged aerosol detector (Dionex) as described previously (31). In brief, 10 l of heparinized blood was separated by centrifugation into erythrocyte and plasma fractions.…”