Erythrocyte peripheral type benzodiazepine receptor/voltage-dependent anion channels are upregulated by Plasmodium falciparum

Abstract

Plasmodium falciparum relies on anion channels activated in the erythrocyte membrane to ensure the transport of nutrients and waste products necessary for its replication and survival after invasion. The molecular identity of these anion channels, termed "new permeability pathways" is unknown, but their currents correspond to up-regulation of endogenous channels displaying complex gating and kinetics similar to those of ligand-gated channels. This report demonstrates that a peripheral-type benzodiazepine receptor, including the voltage dependent anion channel, is present in the human erythrocyte membrane. This receptor mediates the maxi-anion currents previously described in the erythrocyte membrane. Ligands that block this peripheral-type benzodiazepine receptor reduce membrane transport and conductance in P falciparum-infected erythrocytes. These ligands also inhibit in vitro intraerythrocytic growth of P falciparum. These data support the hypothesis that dormant peripheral-type benzodiazepine receptors become the "new permeability pathways" in infected erythrocytes after upregulation by P falciparum. These channels are obvious targets for selective inhibition in anti-malarial therapies, as well as potential routes for drug delivery in pharmacologic applications. (Blood. 2011; 118(8):2305-2312) IntroductionThe most severe form of malaria in humans is caused by parasite Plasmodium falciparum, infecting 225 million people and causing 781 000 deaths in 2009 (World Health Organization, 2010). Erythrocyte invasion by P falciparum provides the parasite access to a plentiful source of nutrients in a locale that is largely shielded from host immune defenses. After invasion, the invading parasite uses a variety of strategies to adapt to the intraerythrocytic environment. To ensure the transport of nutrients and waste products necessary for its replication and survival, P falciparum relies on broad specificity anion channels activated in the erythrocyte membrane after invasion. 1 Initially, this transport was attributed to "new" permeability pathways (NPPs) 2 exported by the parasite to the host membrane. 3 However, later studies revealed that the current is because of up-regulation of endogenous channels 4 and that the diversity of anion channel activities recorded in these studies correspond to different kinetic modalities of a unique type of maxi-anion channel. 5 This channel displays complex gating and kinetics similar to those of ligand-gated channels. 5 Anions are transported through the human erythrocyte membrane by a 2-component system: a large electroneutral exchanger mediated by band 3 and a 4 orders of magnitude smaller electrogenic component estimated at approximately 10 S/cm 2 corresponding presumably to a small number of channels. 6 Remarkably, the molecular identification and characterization of this conductive pathway has not yet been achieved. Neither genomic nor proteomic studies have provided meaningful clues to the composition of this pathway. 7 Considering the small amount of protein a few hundred c...

Patch-Clamp Analysis of Membrane Transport in Erythrocytes

“…VDAC are components of the Peripheral Benzodiazepine Receptor (PBR), composed of at least three components: VDAC, an Adenine Nucleotide Transporter (ANT) and a translocator protein (TSPO) that probably modulates VDAC activity (Veenman et al, 2008). Presence of binding sites for specific ligands of this receptor in the RBC membrane had already been suggested (Canat et al, 1993;Olson et al, 1988), and in this study evidence for presence of transcripts in differentiating erythrocytes and of proteins in mature erythrocytes was given (Bouyer et al, 2011a).…”

“…The channel showed a preference for SCN -ions, since some of these substates could be seen in serum-free conditions when cells were bathed in solutions where 10mM of Cl -ions was replaced by SCN -ions. The identity of this channel remained undetermined until a second study by our group showed that a Voltage Dependent Anion Channel (VDAC) was present in the human RBC membrane (Bouyer et al, 2011a). VDAC are components of the Peripheral Benzodiazepine Receptor (PBR), composed of at least three components: VDAC, an Adenine Nucleotide Transporter (ANT) and a translocator protein (TSPO) that probably modulates VDAC activity (Veenman et al, 2008).…”

“…We recently showed that the main component of NPPs was the PBR complex including the VDAC channel, since the specific PBR ligands could prevent both parasite growth in vitro, NPP-mediated sorbitol permeability and whole cell anion currents (Figure 6) of infected cells (Bouyer et al, 2011a). Fig.…”

“…6. Anion channel activity in Plasmodium falciparum-infected human RBCs 3 cells A, B and C were studied using the whole-cell configuration, and serial perfusion were performed showing the typical serum effect and the inhibitory effect of PBR ligands and NPPB, as described in (Bouyer et al, 2011a). Stimulation was made with 500ms ramps between +100mV and -100mV, with -10mV increments The identification of PBR/VDAC activity in the Plasmodium-infected RBC membrane is an important step in the description of the pathophysiology of malaria.…”

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

“…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.…”