[Frontiers in Bioscience S4, 1424-1448, June 1, 2012]
The malaria digestive vacuole
Juliane Wunderlich1, Petra Rohrbach1, John Pius Dalton1
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TABLE OF CONTENTS
1. ABSTRACT
During the development of malaria parasites within human erythrocytes, the fusion of digestive vesicles gives rise to a large digestive vacuole (DV). This organelle, which is maintained at low pH, processes 60-80 percent of the erythrocyte hemoglobin to provide a pool of amino acids that is crucial for parasite growth and development. During proteolysis, heme is released from hemoglobin as a toxic byproduct and is detoxified by biocrystallization to hemozoin. Proteases that contribute to hemoglobin breakdown, as well as other DV-associated proteins, arrive at this site via several different transport pathways. Antimalarial quinoline drugs, such as chloroquine, act by binding to heme and thus prevent its sequestration into hemozoin. Other drugs, such as artemisinin, may cause oxidative damage of DV macromolecules and membranes. The membrane of the DV contains ion pumps and transporters that maintain its low pH but are also pivotal in the development of parasite resistance to several antimalarial drugs. Methods for the isolation of the DV organelle have been developed to study the biogenesis and function of this important organelle.
2. INTRODUCTION
Malaria is a major cause of mortality and morbidity in tropical and subtropical regions of Latin America, Africa, the Indian subcontinent, and Southeast Asia, where approximately 40% of the world's population is at risk of infection. The disease is caused by unicellular, eukaryotic parasites of the genus Plasmodium. Of the five malaria parasites known to infect humans (falciparum, vivax, malariae, ovale and knowlesi), P. falciparum is the most virulent and responsible for an estimated 225 million clinical cases and 781,000 deaths annually (1, 2). Since resistance to a range of antimalarial drugs is spreading throughout parasite populations worldwide and a protective vaccine remains unavailable, there is an urgent need to develop novel therapeutic strategies.
The life cycle of Plasmodium can be divided into three phases: the pre-erythrocytic hepatic stage and the intraerythrocytic asexual cycle in humans or other vertebrate hosts, and the sexual cycle in female Anopheles mosquitoes. Clinical manifestations of malaria are primarily due to the infection of erythrocytes by intracellular parasites (3). The asexual intraerythrocytic cycle begins with the erythrocyte invasion by a merozoite, which is then enclosed within a newly formed parasitophorous vacuole (PV), separate from the erythrocyte cytoplasm. Three intraerythrocytic stages can be distinguished morphologically: (a) the ring stage, which lasts 22-24 h from the time of merozoite invasion in P. falciparum and during which metabolic activity is low, (b) the trophozoite stage, which is 10-12 h in duration and characterized by an acceleration of metabolic processes as well as the consumption of most of the hemoglobin-rich erythrocyte cytoplasm (4), and (c) the schizont stage where parasites undergo 4-5 rounds of binary division in 8-10 h, generating up to 32 merozoites. Schizonts eventually rupture their host cells, releasing the merozoites that invade new erythrocytes and repeat the cycle.
To support its growth and replication, the parasite takes up large amounts of erythrocyte cytosol and digests its primary constituent, hemoglobin, in a specialized acidic organelle called the digestive vacuole (DV) or food vacuole (5, 6). Large pigment-containing vacuoles were first described in electron microscopy studies of thin sections of P. knowlesi in 1956 (7) and a role for this structure in digestion of hemoglobin was first suggested by Rudzinska and Trager in 1957 (8). As in lysosomes of mammalian cells and yeast vacuoles, an acidic pH is maintained and several different proteinases are present in the DV. However, the absence of the typical lysosomal acid phosphatase and glycosidases, which release phosphate groups and smaller sugars from macromolecules, respectively, indicates that the DV of Plasmodium is a specialized organelle that evolved to efficiently degrade hemoglobin. The parasite apparently does not need to degrade macromolecules other than hemoglobin during its intraerythrocytic cycle (9, 10). Proteolytic degradation of hemoglobin, however, results in the release of toxic heme which is neutralized by the parasite primarily by the formation of pigments composed of hemozoin (5, 6, 11). Hemoglobin hydrolysis as well as heme detoxification are thus essential for the parasite's survival in the host erythrocyte (4, 12).
Due to the presence of two plant-type vacuolar proton pumps in the DV membrane, the vacuole has also been compared to the tonoplast, the acidic intracellular vacuole of plant cells (13), in which these H+-pumps act in concert to maintain a low pH. Moreover, the product of the gene PF13_0353, a putative cytochrome b5 reductase localized in the DV membrane, was found to be structurally similar to a plant nitrate reductase (14, 15). The two plant-type H+-pumps are also involved in the acidification of the lumen of acidocalcisomes, small electron-dense granules of some protozoan parasites that store Ca2+ as a phosphate precipitate (16, 17). However, DVs are morphologically and functionally very distinct from acidocalcisomes, which are involved in calcium homeostasis and polyphosphate metabolism (17, 18).
DVs are also observed in unicellular organisms other than malaria parasites. Electron microscopy studies demonstrated the presence of a DV in Babesia equi, an intraerythrocytic parasite related to Plasmodium (19). The anaerobic protozoan parasite Entamoeba histolytica, the causative agent of amebiasis, ingests erythrocytes and degrades them with their contained hemoglobin in several acidic DVs using cysteine proteinases (20). The soil-living amoeba Dictyostelium discoideum (21) as well as the apicomplexan parasite Theileria (22), but not Toxoplasma, also contain a large acidic DV, although they do not degrade hemoglobin.
3. STRUCTURE AND FORMATION OF THE DV
The DV of Plasmodium contains hydrolytic enzymes of various mechanistic classes that digest hemoglobin (23, see Sect. 4). Microscopically, hemoglobin degradation is indicated by the alteration of the density of DVs (24). The process of hemoglobin digestion leaves insoluble waste products in the DV due to the release of heme (ferriprotoporphyrin IX), which is neutralized by the formation of coordinated hematin dimers (β-hematin) that biocrystallize to the chemically inert hemozoin, also known as the malaria pigment (11, 25-27). Hemozoin crystals can be observed by light and electron microscopy as they are very dense structures (7). These are lined up along a single axis in intact parasites (Figure 1). By contrast, disordered crystals are observed in isolated DVs, possibly due to isolation or fixation artifacts or the loss of the transmembrane pH gradient between the erythrocyte cytoplasm and the DV during isolation (9). It was proposed that this gradient could establish an electromotive force across the vacuole in intact parasites that causes the iron-containing crystals to align within the generated magnetic field (9).
The DV is not inherited by daughter merozoites but forms de novo after each round of infection (28). The biogenesis, morphology, size and number of DVs vary between developmental stages of the parasite (Figure 2). Multiple small dense vacuoles with a diameter of 150 - 300 nm are characteristic of the early and mid-stage rings (29). They are formed by micropinocytosis of host cell stroma from the parasite surface that results in double membrane-vesicles that function as individual DVs (30). The inner membrane, which develops from the parasitophorous vacuole membrane (PVM), rapidly disappears, allowing more efficient nutrient diffusion into the parasite cytoplasm. Hemoglobin is digested and hemozoin crystals are generated at this early stage (30, 31). In late ring-stage parasites, the cytostome, a cytoskeletal ring for the uptake of erythrocyte cytoplasm, appears (32, 33) and the small pigment-containing vesicles coalesce to form a single large DV (30). A recent study, using thin-section transmission electron microscopy (TEM) and 3-D reconstruction, postulated that early ring-stage parasites take on the shape of a cup that closes and internalizes a large amount of erythrocyte cytoplasm in a single event called the 'Big Gulp'. The authors suggested that the DV is derived from the so-formed vacuole rather than several smaller compartments (34). However, this hypothesis remains controversial (35, 36).
Through the cytostome, erythrocyte cytosol is internalized together with the membrane of the PV and the parasite's plasma membrane (32, 37, 38). This engulfment could increase the surface of exchange between erythrocyte and parasite (39). The outer membrane of the cytostomal system is contiguous with the parasite's surface and its inner membrane originates from the PVM (28). Portions of hemoglobin-rich host cytoplasm are pinched off from its distal end and transported to the DV within small double membrane-vesicles. The outer membrane of these vesicles is probably incorporated into the DV membrane and the inner membrane is subsequently degraded by phospholipases and proteinases inside the vacuole (30, 31, 38, 40). A knockout of the gene encoding the DV aspartic proteinase plasmepsin (Plasmodium pepsin) IV (PfPM4) caused abundant accumulation of electron-dense vesicles in the DV in 30% of the P. falciparum-infected cells as compared to wild-type Dd2 parasites, suggesting that PfPM4 might be involved in the elimination of vesicle membranes within the DV (41). The lining membrane of the DV has been described as a mosaic of different molecular compositions with a patchy distribution of DV markers, such as the 19 kDa fragment of merozoite surface protein 1 (PfMSP119) and the chloroquine resistance transporter (PfCRT, 30, 42, 43), probably due to fusion with membranes from the cytostomal system (30) and Golgi-derived vesicles containing proteinases (23).
Hemoglobin degradation occurs predominantly during the trophozoite stage, at which time the DV is enclosed by a single membrane and has a diameter of up to 2.2 �m (30), occupying a significant part of the total volume of the parasite (Figure 1). Late trophozoites show dilated DVs with widely spaced hemozoin, internal membranous structures and an inward folding of the vacuolar membrane (30). During schizogony, the vacuole decreases in size (0.8 - 1.2 �m), loses the folds of its outer membrane as well as the internal membranes (44). Adjacent to the DV, a lipid body of a few hundred nanometers in size forms, whose constituents are probably retrieved from the DV interior, the DV wall, as well as the inner membrane of transport vesicles that fuse with the DV membrane (30). These neutral lipids (44) as well as histidine-rich protein II (HRPII) (45) were suggested to promote β-hematin formation.
In schizonts, large hemozoin crystals are formed, which appear closely packed (30). When the mature schizont ruptures the erythrocyte, merozoites are liberated and leave behind the malarial pigment within a membrane-bound residual body that is released into the blood stream and avidly engulfed by macrophages (30). Released native hemozoin bound to malarial DNA was shown to induce inflammatory responses via binding to Toll-like receptor 9 (TLR9) on the endosomes, resulting in cytokinemia and fever during disease (46). Furthermore, chemically synthesized hemozoin, which is structurally identical to the native crystal, was reported to be sufficient to induce the production of the pro-inflammatory cytokine interleukin-1β (IL-1β) by phagocytes via the activation of the NOD-like receptor containing pyrin domain 3 (NLRP3) inflammasome, which indicates that this metabolic product is critical in malaria pathology (47).
The structure and formation of the DV varies between different Plasmodium species. Whereas only one DV was observed in P. falciparum, P. knowlesi contains several smaller acidic compartments (7). In contrast to other human as well as murine Plasmodium species, the individual DVs fuse together very rapidly in P. falciparum (31). The differences between the cytostomal system of murine parasites such as P. chabaudi, which comprises a long tube connected with a branching tubular network, and that of P. falciparum, showing vesicles budding off from the cytostome (31, 48), are consistent with the phylogenetic relationships between the species (40), i.e., the feeding mechanism of P. falciparum is more similar to avian than either rodent or other human Plasmodium species (33). A cytostome with long tubular structures was also described in Babesia equi (19) and some species of Theileria (22, 49).
4. HEMOGLOBIN DIGESTION
The primary function of the DV in Plasmodium is the degradation of hemoglobin (9), which comprises 95% of the cytosolic erythrocyte protein, corresponding to a concentration of 310 - 350 mg/ml (50). During intraerythrocytic development, the parasite endocytoses large quantities of host cell cytosol and digests 60 to 80% of its hemoglobin (4, 51-53). The most widely suggested purpose of hemoglobin catabolism is the supply of amino acids for the synthesis of new proteins since intraerythrocytic stages of Plasmodium only have a limited capacity of de novo amino acid synthesis (6). This is supported by the observation that radiolabeled amino acids released during hemoglobin degradation are incorporated into plasmodial proteins (54, 55). However, parasite development also depends on amino acids that are rare (Met, Cys, Gln, Glu) or absent (Ile) in hemoglobin, which must be obtained elsewhere (56). Ile has been shown to be of particular significance as P. falciparum can be cultured in medium with Ile as the sole exogenous source of amino acids (4).
According to a study by Krugliak et al. (52), only 16% of the amino acids generated from hemoglobin breakdown are used for protein assembly. This observation suggests that hemoglobin uptake and catabolism could have additional functions, such as the prevention of erythrocyte lysis (57, 58). Since the parasite increases its volume 25-fold in one intraerythrocytic cycle (59), it has been suggested that degradation of host cell cytoplasm might be necessary to make room for parasite growth (57). Furthermore, the diffusion of some hemoglobin-derived amino acids into the host cell (58) could be important for the regulation of intracellular osmolarity of the infected erythrocyte during the maturation and replication of the parasite (52, 60, 61). The use on hemoglobin as a nutrient source is likely to vary with culture conditions (62), given that the entire asexual cycle of P. falciparum, while limited and dependent on constituents of the erythrocyte membrane and stroma, can occur extracellularly (63-66). Interestingly, even axenically grown parasites have a functional DV containing hemozoin crystals (63, 64, 66).
Hemoglobin hydrolysis in the Plasmodium DV is thought to be a semi-ordered process mediated by the action of a series of proteases (9). Plasmepsins (aspartic proteases) and falcipains (cysteine proteases) are involved in the initial steps of the pathway (Figure 3). Out of the ten plasmepsins encoded in the genome of P. falciparum (67), four are localized and function in the DV: PfPM1, PfPM2, PfHAP (histoaspartic proteinase) and PfPM4 (68). The plasmepsins PfPM1 and PfPM2 can cleave non-denatured hemoglobin at Phe33 and Leu34 in the hinge region of the α-globin chain, probably causing the globin subunits to unwind and allowing further proteolysis (9, 69). PfHAP and PfPM4 have a significantly lower capacity to digest hemoglobin as compared to PfPM1 or PfPM2 but are capable of actively cleaving globin in vitro (68). The four DV plasmepsins, as well as the falcipains PfFP-2, PfFP-2' and PfFP-3, cleave these large globin fragments into polypeptides (6, 70). PfFP-2 (71) and PfFP-3 (72) are also able to digest native hemoglobin and might thus participate in its initial cleavage.
Whereas P. falciparum and the chimpanzee parasite P. reichenowi contain four DV plasmepsins, whose genes are located adjacent to one another on chromosome 14, all other examined Plasmodium species only have an ortholog of PfPM4. PfPM1, PfPM2 and PfHAP are thus paralogs of PfPM4 that may have arisen through gene duplication (73). The close phylogenetic relationship between P. falciparum and P. reichenowi strengthens with this finding (74, 75). In P. falciparum, the four DV plasmepsins and the falcipains are expressed at different times throughout the life cycle, which indicates that these enzymes have distinct functions at different developmental stages of the parasite. The highest transcript levels of PfPM1 and PfPM4 were detected in ring-stage parasites, while PfPM2, PfHAP, PfFP-2 and PfFP-2' mRNAs were most abundant in trophozoites and PfFP-3 was synthesized at the late trophozoite and early schizont stage (76-78).
While the metalloprotease falcilysin, which is expressed in trophozoites (78), is unable to digest native hemoglobin or denatured globin, it can degrade polypeptide fragments of up to 20 amino acids into short peptides that are 5-10 amino acids in length (79). The DV dipeptidyl aminopeptidase 1 (PfDPAP1), a plasmodial ortholog of mammalian cathepsin C (80, 81) whose expression level is highest at the trophozoite stage (78, 82), might be involved in the cleavage of dipeptides from the N terminus of hemoglobin-derived oligopeptides (83).
How and where amino acids are released from these hemoglobin-derived dipeptides remains to be fully elucidated. Since aminopeptidase activity was detected in the parasite cytoplasm but not in the DV and was found to be optimal at neutral pH, it was suggested that short peptides are actively transported out of this acidic compartment and subsequently cleaved into amino acids by neutral aminopeptidases in the parasite's neutral cytoplasm (83-88), as seen for lysosomes (89). However, recent studies have reported the presence of aminopeptidase P (PfAPP) and M1 alanyl aminopeptidase (PfM1AAP) in the DV (90-92). Whereas the optimum pH of PfM1AAP is 7.4 (88), PfAPP is active at pH 5.0 to 7.5 and might function in the acidic environment of the DV (91). Therefore, peptides generated by the concerted action of various endo- and exopeptidases are probably cleaved into amino acids and small peptides within the DV as well as in the cytoplasm of the parasite. Small peptides that are transported into the parasite's cytoplasm likely undergo terminal degradation by aminopeptidases stationed here, such as PfM1AAP, M17 leucyl aminopeptidase (PfM17LAP) and M18 aspartyl aminopeptidase (PfM18AAP; 93, Figure 3). These enzymes function optimally at neutral pH (84, 87, 94) and are maximally expressed in early trophozoites, whereas peak expression of PfAPP occurs at the late ring stage (95, www.plasmodb.org).
It was suggested that P-glycoprotein homolog 1 (Pgh-1), a member of the ATP-binding cassette (ABC) transporter superfamily in the DV membrane, might be involved in the export of small peptides because pfmdr1 expressed in a heterologous yeast system was able to complement Ste6, which encodes an ABC-transporter that translocates a 12-amino acid peptide across the vacuolar membrane in S. cerevisiae (96). However, Pgh-1 was shown to be an inwardly-directed transporter, which imports solutes into the DV (97). Thus, peptide export from the DV is most likely mediated by another transporter.
The inhibition of plasmepsins and falcipains by a combination of the aspartic protease inhibitor pepstatin and the cysteine protease inhibitor E-64 leads to a complete block of hemoglobin proteolysis and parasite development, indicating that hemoglobin digestion is essential to the survival of the parasites and that these hemoglobinases are potential targets for antimalarial chemotherapy (62, 69, 98). However, knockout studies have shown that no single DV plasmepsin is essential to P. falciparum intracellular growth (41), which suggests that these enzymes are redundant in their functions. Furthermore, falcipains are able to compensate for the loss of all four DV plasmepsins (4, 99), revealing a functional overlap between these two protease classes. The observed redundancy within and between protease families might be a strategy by which the parasite ensures a sufficient level of proteolytic activity (70). Hemoglobin digestion can also be blocked either by inhibiting the cysteine proteinases with leupeptin and E-64 (100), or upon disruption of the gene encoding the cysteine proteinase PfFP-2 (101), both resulting in undigested hemoglobin-rich erythrocyte cytoplasm accumulated in the DV (100, 101). In PfFP-2 knockout parasites, accumulation of undigested hemoglobin only occurs at the trophozoite but not at the schizont stage, when higher expression of PfFP-3 might cause phenotypic rescue (101). As PfFP-3 disruption has not been achieved, in contrast to the other falcipain genes, PfFP-3 might play an important role in parasite survival (102).
Inhibitors of the neutral aminopeptidases PfM1AAP and PfM17LAP have been shown to be lethal for P. falciparum in culture as well as for P. chabaudi in vivo, thus providing proof of concept for considering these enzymes as targets for future antimalarial drugs (93, 103). Both aminopeptidases are encoded by single-copy genes and gene knockout experiments (90, 93) as well as a study using enzyme-specific inhibitors (92) have shown that these are of critical importance for parasite survival and are not functionally redundant. However, it is not clear whether the site of action of the aminopeptidase inhibitors is in the DV, the cytoplasm, or both. While strong synergism was observed between inhibitors of aspartic and cysteine proteases using in vitro malaria killing assays, little synergism was observed between aminopeptidase inhibitors and aspartic or cysteine protease inhibitors (104).
During the proteolytic pathway, large quantities of heme are released that can disrupt membranes, inhibit enzymatic processes and initiate oxidative damage (5, 53, 105). The detoxification of this toxic byproduct can be accomplished by peroxidative degradation (53), decomposition by glutathione (106) or conversion into the biologically inert crystalline polymer hemozoin (11, 25-27). Thus, the DV has other functions in addition to hemoglobin digestion, such as detoxification of heme and oxygen radicals (39), storage of non-degradable biomolecules, already evident in hemozoin sequestration, and regulation of intracellular osmolarity of the infected erythrocyte (52, 60, 61).
5. ION HOMEOSTASIS
Ions play an important role in many biochemical reactions and the ability of cells to maintain intracellular levels of essential ions and ionic gradients is critical for a variety of cell functions. In the DV, both pH and calcium (Ca2+) homeostasis have been investigated.
5. 1. pH homeostasis
Many of the enzymes involved in hemoglobin degradation, such as cysteine, aspartic and metalloproteinases, have reported pH optima in the range of 4.5 to 5.5 (9, 69, 107). The DV needs to maintain a similar pH to allow for efficient hemoglobin proteolysis and the biocrystallization of heme to hemozoin (108, 109).
The mechanisms by which the DV pH (pHDV) is maintained remain unclear. It was suggested that inwardly-directed proton pumps (mainly an H+-ATPase and partially an H+-translocating pyrophosphatase) are involved (110-112) that also work against the leak of protons out of the DV (113). Quinoline drugs, such as chloroquine (CQ), quinine (QN) and mefloquine (MQ), may increase pHDV via weak base and non-weak base effects at their biologically active concentrations, effectively preventing the function of parasite acid proteases (9, 114-117). Knowledge of how new drug compounds interfere with pH regulation in the parasite, which is crucial for parasite survival, could give insights in their potential efficacy in malaria treatment.
It has long been debated if, and to what extent, pHDV plays a role in determining levels of CQ accumulation and hence the CQ sensitivity of the parasite (117, 118). Since CQ and many other 4-aminoquinolines are weak bases, their uptake is influenced by passive diffusion along the pH gradient between the extracellular and intracellular compartments (38, 119). It was postulated that an increased pHDV in CQ-resistant (CQR) parasites and thus a reduction in the transvacuolar pH gradient would result in a decreased uptake of CQ into the DV (114, 117, 120).
The first measurements of steady-state pHDV were performed by spectrofluorometry using the pH-sensitive fluorescein-dextran and gave a value of approximately 5.2 (120). This was followed by Ginsburg et al. using the membrane-permeable fluorochrome acridine orange (AO), where a pHDV value of approximately 4.2 was determined in a spectrofluorometric setting (121). More than ten years later, values of 5.2 − 5.6 were reported using AO and epifluorescence microscopy (122, 123). These measurements were later determined unreliable since AO was shown to be very photosensitive and causes lipid peroxidation leading to DV membrane lysis (124, 125). Using the ratiometric pH-sensitive dye DM-Nerf and confocal fluorescence microscopy, the pHDV values of 5.2−5.6 were confirmed (126). However, these results could not be reproduced by other groups in subsequent studies using the same fluorochrome. In 2006, Hayward et al. determined pHDV using spectrofluorometry and reported pH values that ranged from 3.7 to 6.5 depending on the fluorochrome used for measurement and what concentration was applied (118). Furthermore, the group reported comparable pHDV values for CQ-sensitive (CQS) and CQR parasites (118), challenging previous studies (122, 123, 126, 127). This result was confirmed by two subsequent studies (128, 129), showing comparable pHDV values for CQS and CQR parasites. Whereas Hayward et al. (118) and Klonis et al. (128) analyzed parasite populations of different strains using spectrofluometry and flow cytometry, respectively, Kuhn et al. (129) studied single live parasite-infected erythrocytes by confocal fluorescence microscopy and calibrated the cytoplasmic and digestive vacuolar fluorescence ratios for each cell by selecting defined regions of interest (Figure 4). This method allows for site-specific pH measurements in the intact parasite. Using P. falciparum expressing pHluorin, a ratiometric pH-sensitive GFP (130), pH values of 7.15 � 0.07 in the cytoplasm and 5.18 � 0.05 in the DV of the parasite were determined, respectively (129). These pH determinations were independent of intracellular pHluorin concentrations and the non-invasive nature of the technique was shown to preserve the physiological conditions of the parasite (125, 129). This method was validated by confirming the measured pH values of the cytoplasm using the pH-sensitive fluorochromes SNARF-1 AM and SNARF-5F AM (129).
The similar pHDV values of different sensitive and resistant parasite strains indicate that mechanisms other than altered drug uptake and/or pH-dependent changes in heme solubility, turn-over or biocrystallization rates are responsible for the CQR phenotype as well as the reduced drug accumulation observed in CQR parasites (131-133). A more favored explanation is that alterations in carrier-mediated drug efflux reduce the CQ concentration in the DV of CQR parasites (21, 133-136). It was suggested that mutations in the DV membrane proteins Pgh-1 and PfCRT (42, 137, 138, see Sect. 6) cause CQ resistance (138, 139).
5. 2. Calcium homeostasis
Calcium is a well-known second messenger and Ca2+ ions have been shown to be important for parasite development (140). In P. falciparum, Ca2+ signaling is involved in invasion (141-143), maturation (144), synchronization (140), exflagellation and gamete formation (145, 146). For the intraerythrocytic parasite, Ca2+ is not readily available since the Ca2+ concentration in the host erythrocyte cytoplasm is low. For this reason, the parasite must build up a regulatory Ca2+ pool to maintain its Ca2+ homeostasis (147, 148), a function normally associated with ATP-dependent endoplasmic reticulum (ER) pumps in eukaryotes (149). The DV was initially suggested to act as a dynamic Ca2+ store from fluorescence studies using the non-ratiometric green fluorescent Ca2+ indicator Fluo-4 (18) but confocal fluorescence studies using the ratiometric Ca2+ indicator Fura-Red showed that this organelle is unlikely to serve as a Ca2+ pool (150). X-ray spectral analysis and elemental mapping confirmed no increase in Ca2+ levels within this organelle (150). The intense Fluo-4 fluorescence observed from the DV was thought to be due to the pH dependence of the dye and its active transport into the DV through Pgh-1 rather than an increased Ca2+ content (97, 150). It is more likely that the parasitophorous vacuole (148), the acidocalcisomes (151) the mitochondria (147, 152) and/or the ER (153, 154) carry out the function of a Ca2+ store and regulator.
6. TRANSPORTERS IN THE DV MEMBRANE
The maintenance of ion homeostasis in the parasite, as well as nutrient uptake, is mediated by integral membrane proteins that are able to transport ions and other molecules across a biological membrane. Since these processes are important for survival, growth and replication of the parasite, transport proteins of P. falciparum are promising drug targets although only a few of them have been characterized to date (155). Using a genomic database approach, the 'permeome' of the malaria parasite was characterized and more than 100 transport proteins encoded by P. falciparum were examined for a range of organic and inorganic substrates (156). Table 1 shows a list of proteins that are known to be localized in the DV membrane and function as transporters in P. falciparum.
P-glycoprotein homolog 1 (Pgh-1), also known as multi-drug resistance protein 1 (PfMDR1), is a member of the ABC (ATP-binding cassette) transporter superfamily and an ortholog of mammalian P-glycoproteins that mediate multi-drug resistance in cancer cells (157). It is encoded by the mdr homolog pfmdr1 (158, 159), predominantly localizes to the DV membrane (160, 161), with its ATP-binding domain facing the cytoplasm (160, 162), and is expressed during the intraerythrocytic cycle of Plasmodium, with a peak of pfmdr1 mRNA abundance at the schizont stage (163). This transporter is known to play a role in resistance to several antimalarial drugs (139). A strong association has been observed between pfmdr1 amplification, which leads to increased Pgh-1 expression, and MQ treatment failure (164). In vitro cross-resistance to QN, lumefantrine (LF), halofantrine (HF) and artemisinin (ART) was also reported (165-167). A higher pfmdr1 copy number was also associated with an increased susceptibility to CQ (159, 164, 168, 169). Furthermore, since the point mutation N86Y was frequently found in parasites showing high-level CQ resistance (170-173), it is believed that pfmdr1 mutations can enhance the degree of CQ resistance albeit are insufficient to confer complete resistance to this drug (139, 170, 172, 174). It is not fully understood how Pgh-1 mediates these effects although it might directly transport drugs across the DV membrane or indirectly affect their partitioning by altering the transport of other substrates (175). Pgh-1 might transport some drugs, including MQ, HF and ART, into the DV where they may be less harmful for the parasite (97). By contrast, increased CQ or QN import and accumulation in the DV could lead to a decreased CQ resistance level since these antimalarials are known to act in this acidic compartment (97, 166). One study showed that Chinese hamster ovary (CHO) cells expressing wild-type Pgh-1 exhibit higher intracellular CQ accumulation and are therefore more sensitive to the drug compared to cells expressing a mutant Pgh-1 bearing the amino acid changes S1034C and N1042D, which have been associated with the CQ resistance phenotype in P. falciparum (176). Moreover, the transport of chemically and structurally unrelated drugs into the lumen of the Plasmodium DV via Pgh-1 can be inhibited by the P-glycoprotein inhibitors XR-9576 (tariquidar) and ONT-093 (97).
The chloroquine resistance transporter (PfCRT) is a well-studied marker for the DV membrane whose expression peaks at the ring and trophozoite stage (42, 76, 138). It is a member of the drug/metabolite transporter (DMT) superfamily (177, 178) and can extrude CQ from the DV, thereby preventing the drug from accumulating to toxic levels (179-181). Polymorphisms within the pfcrt gene are linked to CQ treatment failure as well as in vitro CQ resistance (138, 173). In particular, the mutation K76T is conserved in CQ-resistant parasites and used as a molecular marker of CQ resistance (170, 182). In some CQR parasite strains carrying a mutation at position 76 in PfCRT, CQ transport out of the DV has been shown to be inhibited by verapamil (183). Different pfcrt alleles were also shown to influence the in vitro susceptibility of parasites to other antimalarial agents including QN, quinidine (QD), amodiaquine (AQ), HF, MQ and ART (42, 184-186). Furthermore, QN, QD, AQ, MF, primaquine (PQ), quinacrine (QC) and small peptides containing aromatic amino acids were capable of competing with CQ for transport via mutated PfCRT (135, 180). Thus, mutant PfCRT proteins might transport a wide range of clinically important antimalarial drugs as well as peptides out of the DV (Figure 5).
Two discrete H+ pumps are present in the DV membrane: the V-type (vacuolar) H+-ATPase and the V-type H+-translocating pyrophosphatase (H+-PPase) (111, 112). The H+-ATPase is a heteromultimeric enzyme complex whose structure is well conserved among animals, plants, fungi and some bacteria. It is composed of a cytosolic V1 domain, which contains the ATP catalytic site, and a transmembrane V0 domain. In P. falciparum, the V1 domain is composed of three A subunits, three B subunits, two G subunits and the subunits C, D, E, F and H, while the V0 domain consists of the subunits a, c, c" and d (112). The genes encoding these subunits are transcribed throughout all stages of the parasite's life cycle and do not differ in the sequence of their coding regions between CQS and CQR P. falciparum-strains (187-189), suggesting that the H+-ATPase is not involved in the CQ resistance phenotype (187). However, this H+ pump has a major role in the acidification of DV and the maintenance of the transvacuolar pH gradient (111, 190, 191). Its function is dependent on ATP (111), and is sensitive to concanamycin A (111), bafilomycin A1 (127) and N-ethylmaleimide (127). Interestingly, the amino acid sequences deduced from the cloned H+-ATPase genes of the parasite were more similar to those of plant origin than those of animal origin (112).
V-type H+-PPases are highly conserved monomeric membrane proteins with an apparent molecular mass of 56-79 kDa and a 15- or 16-transmembrane topology. They are found in plant vacuoles, plasma membranes of phototrophic bacteria and various membranes of some protozoa. They translocate protons across a membrane by using potential energy derived from hydrolysis of the phosphoanhydride bond of pyrophosphate (111, 192, 193). The properties of Plasmodium H+-PPases are similar to those of plants as well as of other protozoa including Trypanosoma cruzi (194), T. brucei (195), and Leishmania donovani (196). As in plants, two different H+-PPases are encoded in the genome of P. falciparum: PfVP1, an ortholog of the canonical plant type I H+-PPase, and PfVP2, an ortholog of the plant type II H+-PPase (197). While both proteins are expressed in intraerythrocytic parasites, the mRNA level of PfVP1 is higher than that of PfVP2 and is greatest in early trophozoite- and schizont-stage parasites (197). PfVP1 is predominantly localized in the DV membrane and to a lesser extent in the plasma membrane (13, 112, 197). The enzyme requires Mg2+ and modest concentrations of K+ (50 mM) for its activity (13, 16, 197) and can be inhibited by sodium fluoride (NaF, 111) or aminomethylenediphosphonate (AMDP, 197).
The co-localization of vacuolar H+-ATPases and H+-PPases in the DV membrane in P. falciparum resembles the situation in the tonoplast of plant cells (13) and in the acidocalcisomes of a number of protozoan parasites (16) where the two H+ pumps acidify the same compartment (112). As a byproduct of several major biosynthetic pathways, pyrophosphate is an abundant energy source and since intraerythrocytic malaria parasites rely mainly on glycolysis for ATP production (195), it is thought that its use is a mechanism to conserve ATP for vital functions in the cell (193). However, it was also postulated that the H+-ATPases play the major role in the acidification of the DV lumen since H+-PPases may not be capable of doing this alone (111). Due to their pivotal involvement in energy conservation and membrane transport in malaria parasites, and particularly because of the apparent absence of H+-PPases in animals, these enzymes are considered important new antimalarial drug targets (193).
The existence of H+-coupled transporters in the DV membrane such as a Na+/H+ antiporter has been suggested (18) but remains to be shown experimentally. Furthermore, it was proposed that a Ca2+/H+ antiporter and a P-type Ca2+-ATPase, which is sensitive to thapsigargin and cyclopiazonic acid, function as transport proteins in the DV membrane (18), but have later been shown to be located to mitochondria (152) and a perivacuolar cytoplasmic region (198). Additionally, P. falciparum contains the gene PFA0375c that codes for a lipid/sterol-H+: symporter with strong homology to Niemann-Pick type-C proteins, which mediate the efflux of lipids and cholesterol from lysosomes (www.plasmoDB.org, 199, 200). This transporter may be involved in the H+-coupled extrusion of lipids and sterols from the DV (156). Another protein - PFE1185w - could be an H+-driven effluxer of Fe2+ from the DV into the parasite's cytoplasm (156, www.plasmoDB.org) since it shows close homology to endosomal Fe2+ NRAMP2 transporters that are involved in the transferrin cycle (201). The function and localization of these proteins remain to be established.
7. PROTEIN TRAFFICKING TO THE DV
The trafficking pathway of parasite-derived proteinases that are responsible for hemoglobin proteolysis in the DV of P. falciparum takes advantage of the parasite's feeding mechanism (23). The premature enzymes contain putative N-terminal signal peptides for translocation into the ER and translation by ER-bound ribosomes, as well as a prodomain that bears additional targeting motif(s) for subsequent transport to the DV (28). These motifs are probably responsible for the targeting of these proteins to the DV (28) since green fluorescent proteins (GFPs) attached to a signal peptide can target the DV, while those without this targeting signal localize to the cytoplasm of the parasite (80, 129, 202, 203).
The proenzymes, as well as several other DV-targeted Plasmodium proteins, enter the secretory pathway of the parasite via the ER (6, 204) but it is unclear whether they transit a post-ER compartment such as the Golgi apparatus since they are not glycosylated (205). For the proteins that target the DV, differences in solubility and prodomain structure have been reported (28) and several trafficking routes have been hypothesized (Figure 6): for example, proteins can traffic directly from the ER to the DV membrane (Figure 6-I, e.g. PfCRT). Other proteins are transported from the ER to the cytostomal system, where they accumulate before being trafficked to the DV via double-membrane vesicles along with their substrate hemoglobin. The transport from the ER to the cytostomal system occurs either directly (Figure 6-II, e.g. PfPM2), via the parasite's plasma membrane (Figure 6-III, e.g. PfPM1, PfFP-2 and PfFP-3), via the PV (Figure 6-IV, e.g. PfDPAP1) or via the host erythrocyte's cytosol (Figure 6-V, e.g. PfHRP2). Additional proteins or substrates for nutrition could also be engulfed by the cytostome, together with those carrying a targeting sequence.
The premature DV plasmepsins are integral membrane proteins and are inserted into the ER via a transmembrane domain in their proregions. Pro-PfPM1 is initially transported to the parasite's plasma membrane before it is incorporated into the cytostomal system, possibly by lateral diffusion in the plasma membrane (206, Figure 6-III), whereas pro-PfPM2 reaches this organelle without prior trafficking to the parasite surface (Figure 6-II). Vesicles containing pro-PfPM2 are thought to fuse with the outer membrane of the cytostomal vacuole. The proenzyme is thus probably located in the space between the two vacuole membranes during transport to the DV (23). The N-terminal 70 amino acids (39 cytoplasmic, 20 transmembrane, and 11 lumenal residues) of the PfPM4 proenzyme were shown to be sufficient for targeting of a fluorescent protein version to the DV (129). Once in the DV, the proplasmepsins are anchored in the vacuolar membrane and subsequently cleaved between the residues Gly124 and Ser125 by falcipains at a low pH, resulting in their maturation and activation (23, 70, 205, 207, Figure 6). However, while this may be the prime mechanism of activation of the plasmepsins, they are capable of autoactivation when falcipain activity is inhibited. This suggests that alternate means of plasmespsin activation exists and adds a further level of redundancy between these DV proteases (70).
The cysteine proteases PfFP-2 and PfFP-3 are transported to the DV via the parasite's plasma membrane and the cytostomal system and contain large membrane-spanning prodomains with a bipartite trafficking signal consisting of a cytoplasmic and luminal portion (80, 81). It was suggested that these falcipains are processed in an intracellular compartment before reaching the DV (77, 81). Dipeptidyl aminopeptidase 1 (PfDPAP1) was shown to accumulate in the PV before being ingested through the cytostome as a soluble protein and transported to the DV (Figure 6-IV). Its signal sequence does not contain a transmembrane domain and the internal prodomain is localized between an exclusion domain and the two catalytic domains (83).
DV-trafficking via the parasite's endocytic pathway has also been demonstrated for plasmodial proteins that are not involved in hemoglobin digestion. The merozoite surface proteins 1 and 8 (PfMSP1 and PfMSP8) are both initially transported to the parasite's plasma membrane, where they are cleaved. However, whereas PfMSP1 is produced in schizonts and its proteolytic cleavage comes about upon release of free merozoites and at the time of their invasion of erythrocytes, PfMSP8 is synthesized and processed at the ring stage (30, 208). The remaining non-cleaved fragment of PfMSP1 (PfMSP119) is endocytosed together with erythrocyte cytosol into small DVs typical of early and mid-ring-stage parasites by micropinocytosis, while the PfMSP8 fragment travels to the single DV of late rings/early trophozoites (30).
Histidine-rich protein 2 (PfHRP2), which is suggested to promote the formation of hemozoin, was reported to be secreted into the host erythrocyte cytosol before it is ingested and transported to the DV via the hemoglobin ingestion pathway (45, Figure 6-V). The integral membrane protein PfCRT (see Sect. 6) was suggested to pass through the ER and the Golgi complex and then directly traffic to the DV membrane (Figure 6-I). It was postulated that phosphorylation of Thr416 in PfCRT is critical for this process (209).
The molecular mechanisms and the proteins involved in trafficking of plasmodial proteins from the ER to the cytostomal system, in membrane translocation events and in the formation of the structures of the endocytic pathway remain unclear (28). Recently, a new DV trafficking route that excludes the secretory pathway has been shown for the FVYE domain-containing protein (PfFCP), a phosphatidylinositol 3-phosphate (PI3P)-binding protein without a transmembrane domain, which is directly transported from the cytoplasm to the DV via a C-terminal 44-amino acid peptide (210). However, the precise trafficking pathway of this protein is unknown. It is clear that DV-targeted proteins take a number of different routes to reach their destination and some of these pathways are still unclear.
8. MECHANISMS OF DRUG ACTION AND RESISTANCE IN THE DV
The DV plays a central role in parasite metabolism and is an important site of action of several potent antimalarial drugs that target the specialized functions of hemoglobin digestion and/or heme detoxification. Originally, CQ offered an effective low-cost treatment of falciparum malaria. However, with the spread of CQR parasites, the search for new agents capable of reversing or circumventing CQ resistance has become a major focus of malaria research (211-213). Therefore, a better understanding of the mechanism of action of CQ as well as the pathways and physiological parameters associated with CQ resistance is essential.
CQ is a lipophilic weak base that permeates membranes as an uncharged species. It readily accumulates in the DV of the parasite by passive diffusion along the pH gradient between the pH neutral cytosol and the acidic DV. In the DV lumen, CQ is protonated and becomes trapped since the vacuolar membrane is impermeable to the diprotic base (120). The drug is suggested to bind to heme, which prevents its detoxification and leads to the accumulation of toxic byproducts of hemoglobin digestion (25, 214). The resulting heme-CQ complexes are thought to disrupt membranes and result in killing of the parasite (215, 216). As mentioned in Section 5, CQ might also cause alkalinization of the DV lumen and thus inhibit or slow down hemoglobin digestion by acid hydrolases (114-117, 120, 217).
CQ resistance is associated with a decreased accumulation of the drug in the DV and, with this, its restricted access to heme (131, 136, 218). The Ca2+ channel blocker verapamil increases CQ accumulation in CQR parasites, attenuating resistance levels (136, 219). Mutations in PfCRT are thought to affect CQ accumulation in the DV and thus the level of CQ-sensitivity of the parasite (138, 220). The substitution of the positively charged K76 residue with a neutral residue (such as Thr, Asn or Ile) in the first transmembrane domain of PfCRT might allow the protonated drug to traverse the transporter protein and travel to the parasite's cytoplasm, thereby reducing its concentration in the DV (42, 138, 185, 221). It was proposed that this CQ efflux is associated with the H+ leak from the DV and that it is sensitive to verapamil since CQ causes a verapamil-sensitive increase in the alkalinization rate of the DV in CQR strains after blocking the H+-ATPase with concanamycin A (113). Additionally, mutations in pfmdr1 (see Sect. 6) and other yet unidentified genes in addition to those in pfcrt may also be involved in the development of resistance to CQ (221).
Other quinoline drugs, such as QN, MQ and AQ, are also thought to affect DV function but it is not clear if their action involves interfering with hemozoin formation (222, 223). Furthermore, since these agents are weaker bases than CQ, they are not likely to accumulate to the same levels in the DV (224, 225). As previously mentioned, MQ resistance and treatment failure have been linked to pfmdr1 amplifications, whereas the substitutions K76T in PfCRT and N86Y in Pgh-1 were found to be associated with reduced susceptibility to both CQ and AQ (226-228). A high-throughput chemical screening of 2816 compounds of the NIH Chemical Genomics Center Pharmaceutical collection that were registered or approved for human or animal use revealed that 96% of differential responses of current P. falciparum populations to many of the analyzed compounds were associated with mutations in pfmdr1, pfcrt or pfdhfr (Plasmodium falciparum dihydrofolate reductase). These data indicate that the differences in parasite drug sensitivity are dominated by a limited number of genes (229).
ART and its derivatives, e.g. artemether, artesunate and dihydroartemisin, are the most important class of antimalarials used today. ART-based combination therapies (ACTs) are currently used as first-line treatment of uncomplicated P. falciparum malaria in most malaria-endemic areas. They comprise ART derivatives in combination with one or more longer acting antimalarial drug(s) of a distinct chemical class, for example LF, MQ or AQ (230, 231). These combinations are thought to decrease the selection pressure for the development of drug resistance (231). However, strains of P. falciparum with a decreased sensitivity to ART have recently emerged in Bangladesh, Thailand and Cambodia (232, 233), which has raised concerns about their spread and the loss of ACT efficacy. ART susceptibility was shown to be influenced by genetic changes in pfmdr1 and pfcrt (231) although studies on the exact mechanisms of decreased drug sensitivity remain to be carried out. Since point mutations in these genes and amplification of pfmdr1 are known to affect parasite responses to different antimalarials by altering drug accumulation in the DV, ART accumulation in the DV might relate to its mode of action (234).
The mechanism of action of ART and its derivatives is poorly understood (235). It was postulated that the cleavage of the endoperoxide bridge of ART by reduced heme-iron produced during hemoglobin digestion in the DV causes the generation of toxic drug metabolites that damage parasite macromolecules (236, 237). Using live-cell microscopy, it was shown that fluorescent ART trioxane derivatives are rapidly accumulated in DV-associated neutral lipid bodies of trophozoites and schizonts, where they may be activated by neutral lipid-associated heme. Oxidative damage of parasite membranes induced by ART derivatives was observed using an oxidation-sensitive BODIPY probe and by chromatographic analysis of lipid extracts from ART-exposed parasites (238). Consistent with this, ART activity can be enhanced by oxygen and oxidizing agents and decreased by reducing agents (239). However, the activation of ART derivatives by heme may not be the only manner in which these compounds operate since the drug is also effective against ring stages, which do not contain high concentrations of heme (237).
Other studies have shown that ART and its derivatives cause heme alkylation in cultured P. falciparum and in mouse malaria models, suggesting that the resulting heme-adducts are toxic to the parasite (240, 241). It was also proposed that ART activation occurs via cleavage of peroxide bonds by intracellular iron-sulfur redox centers, which are commonly found in enzymes of Plasmodium, and that alkylation of these enzymes may then cause parasite death (242). By electron microscopy, it was shown that ART derivatives cause early disruption of the membranes of the DV, the nucleus and other organelles, which is followed by the loss of organellar structures (243). Furthermore, ARTs inhibited parasite endocytosis of macromolecules from the host cell (244). Studies using radiolabelled ART identified several drug-interacting parasite proteins, suggesting that their alkylation and inactivation might result in parasite killing (245, 246). Proteins that have been proposed to be the ART target include: cysteine proteases (247), proteins of the electron transport chain (248), and PfATP6, a sarcoplasmic/endoplasmic reticulum Ca2+-ATPase (SERCA) ortholog (249).
Finally, recently identified inhibitors of the cysteine proteinases PfFP-2 and PfFP-3 directly target hemoglobin digestion and prevent the in vitro growth of P. falciparum at low nanomolar concentrations (250). Moreover, these small-molecule inhibitors cured P. falciparum infections in a human-engrafted mouse model (251, 252). HIV-1 aspartic protease inhibitors have been reported to block P. falciparum development in vitro and to reduce parasitemia in P. chabaudi-infected mice (253, 254). These findings are important for the development of antimalarial treatment strategies in regions that are endemic for both HIV and malaria. It was suggested that these antiretroviral drugs target the DV plasmepsins PfPM2 and PfPM4 (253, 254). However, since a disruption of the four DV plasmepsins is not lethal to P. falciparum (41, 255), it is unlikely that the antimalarial activity of these compounds is due to inhibition of these aspartic proteinases (256). Recently, it was shown that some HIV protease inhibitors enhance the antimalarial activity of CQ and MQ against P. falciparum in vitro (256-258), suggesting that the administration of both drug groups in combination therapies might potentiate their efficacy in patients (258).
9. DV ISOLATION AND PROTEOMIC ANALYSIS
Since the DV is hardly accessible to standard cellular experimental techniques, purification of intact and functionally active vacuoles is necessary to dissect the process of hemoglobin degradation, the regulation of ion homeostasis and the mechanisms of drug action and resistance (9, 259). A number of methods for the isolation of this organelle have been employed (Table 2). Most studies take advantage of the density of the iron-rich hemozoin crystals that allow the separation of DVs from other parasite fractions by differential centrifugation and Percoll density gradient separation (9, 259). The most commonly used procedures are those developed by Goldberg et al. (9) and Saliba et al. (259).
The Goldberg et al. method yielded 35-fold purified DVs capable of degrading hemoglobin at a pH optimum of 5.0-5.5 (9). While this preparation facilitated the study of the DV plasmepsins, it was not the method of choice for the studies of the energy-dependent accumulation of CQ in the organelle by Saliba et al. (259). The relevant differences in the protocol employed by these authors included: (a) ice-cold water at pH 4.5 instead of the osmotic laxative sorbitol was used for parasite lysis, (b) the vacuoles were triturated through a 27-G needle only 4 instead of 10 times, and (c) DNase I was used to prevent the adherence of DVs to DNA, whereas Goldberg et al. (9) added a small amount of saponin and streptomycin sulfate to avoid vacuolar aggregation to other organelles. This apparently achieved a higher yield of isolated DVs (20% instead of 2-3%) and ATP-dependent CQ accumulation in functionally intact vacuoles from different parasite strains could be studied (259). However, according to an assessment by Lamarque et al. (260), this procedure only yields an approximately 10-fold enriched DV fraction. The purity and integrity of the isolated vacuoles was assessed by light and electron microscopy, immunofluorescence microscopy, immunoblotting, enzyme assays as well as mass spectrometry (9, 73, 259, 260). Table 2 details the methods employed for DV isolation and highlights some of the protein markers of the DV as well as of other compartments used to analyze the quality of the preparations.
Many of the DV proteins reported to date are implicated in the process of hemoglobin digestion (6). Knowledge of all DV-associated proteins (soluble and membrane-bound) would help to dissect the mechanisms involved in biogenesis, functions and interactions of the vacuole. Therefore, the proteome of DV-enriched fractions prepared with the method by Saliba et al. (259) was recently analyzed using tandem mass spectrometry by Lamarque et al. (260). Of the 116 proteins identified by these authors, many were known DV markers such as the DV plasmepsins, subunits of the vacuolar ATP synthase, V-type H+-PPase, Pgh-1 and PfCRT. Furthermore, several proteins of unknown functions that are most likely localized in the DV membrane were detected. Of note, many non-DV proteins, such as PfM17LAP, Stevor, HSP60 and other mitochondrial, cytoplasmic, ER and rhoptry proteins were also detected, which indicates that the DV preparations were contaminated with proteins of other parasitic compartments (260).
10. CONCLUSIONS AND PERSPECTIVES
The DV of the malaria parasite carries out a variety of critical and specialized functions such as hemoglobin degradation in an acidic environment, peptide and/or amino acid transport, heme biocrystallization, detoxification of oxygen radicals, as well as ion homeostasis and thus offers many opportunities for novel antimalarial drug targeting (5, 39). Debates remain concerning the purpose of hemoglobin degradation, the site of amino acid generation from hemoglobin-derived peptides and the mechanisms of antimalarial drug action and resistance. A better understanding of these processes in the malaria parasite will be important for the future development of new therapeutic strategies to reduce the devastating impact of one of the most critical public health problems worldwide.
11. ACKNOWLEDGEMENTS
JW is supported by a doctoral fellowship from the German Academic Exchange Service (DAAD). PR is supported by the Natural Sciences and Engineering Research Council of Canada (NSERC) and the Canada Foundation for Innovation (CFI). JPD is a Canadian Research Chair (Tier 1) in Infectious Diseases and supported by the Canadian Institutes of Health Research (CIHR).
12. REFERENCES
1. I Vythilingam, Y NoorAzian, T Huat, A Jiram, Y Yusri, A Azahari, I NorParina, A NoorRain, S LokmanHakim: Plasmodium knowlesi in humans, macaques and mosquitoes in peninsular Malaysia. Parasit Vectors 1(1), 26 (2008)
http://dx.doi.org/10.1186/1756-3305-1-26
2. WHO: World Malaria Report 2010. Geneva, Switzerland (2010)
3. LH Miller, MF Good, G Milon: Malaria pathogenesis. Science 264(5167), 1878-1883 (1994)
http://dx.doi.org/10.1126/science.8009217
PMid:8009217
4. J Liu, ES Istvan, IY Gluzman, J Gross, DE Goldberg: Plasmodium falciparum ensures its amino acid supply with multiple acquisition pathways and redundant proteolytic enzyme systems. Proc Natl Acad Sci USA 103(23), 8840 (2006)
http://dx.doi.org/10.1073/pnas.0601876103
PMid:16731623 PMCid:1470969
5. R Banerjee, DE Goldberg: The Plasmodium food vacuole. In: Antimalarial Chemotherapy: Mechanisms of Action, Resistance, and New Directions in Drug Discovery. Ed: PJ Rosenthal, Humana Press, Totowa, NJ (2001)
6. DE Goldberg: Hemoglobin degradation. Curr Top Microbiol Immunol 295(Part III), 275-291 (2005)
http://dx.doi.org/10.1007/3-540-29088-5_11
7. JD Fulton, TH Flewett: The relation of Plasmodium berghei and Plasmodium knowlesi to their respective red-cell hosts. Trans R Soc Trop Med Hyg 50(2), 150-156 (1956)
http://dx.doi.org/10.1016/0035-9203(56)90076-1
8. MA Rudzinska, W Trager: Intracellular phagotrophy by malaria parasites: an electron microscope study of Plasmodium lophurae. J Eukaryot Microbiol 4(3), 190-199 (1957)
http://dx.doi.org/10.1111/j.1550-7408.1957.tb02507.x
9. DE Goldberg, AF Slater, A Cerami, GB Henderson: Hemoglobin degradation in the malaria parasite Plasmodium falciparum: an ordered process in a unique organelle. Proc Natl Acad Sci USA 87(8), 2931-2935 (1990)
http://dx.doi.org/10.1073/pnas.87.8.2931
10. SE Francis, DJ Sullivan Jr, DE Goldberg: Hemoglobin metabolism in the malaria parasite Plasmodium falciparum. Annu Rev Microbiol 51(1), 97-123 (1997)
http://dx.doi.org/10.1146/annurev.micro.51.1.97
PMid:9343345
11. TJ Egan, JM Combrinck, J Egan, GR Hearne, HM Marques, S Ntenteni, BT Sewell, PJ Smith, D Taylor, DA van Schalkwyk: Fate of haem iron in the malaria parasite Plasmodium falciparum. Biochem 365(Pt 2), 343-347 (2002)
http://dx.doi.org/10.1042/BJ20020793
PMid:12033986 PMCid:1222701
12. S Kumar, M Guha, V Choubey, P Maity, U Bandyopadhyay: Antimalarial drugs inhibiting hemozoin (β-hematin) formation: A mechanistic update. Life Sci 80(9), 813-828 (2007)
http://dx.doi.org/10.1016/j.lfs.2006.11.008
PMid:17157328
13. S Luo, N Marchesini, SNJ Moreno, R Docampo: A plant-like vacuolar H⁺-pyrophosphatase in Plasmodium falciparum. FEBS Lett 460(2), 217-220 (1999)
http://dx.doi.org/10.1016/S0014-5793(99)01353-8
14. G Ostera, F Tokumasu, F Oliveira, J Sa, T Furuya, C Teixeira, J Dvorak: Plasmodium falciparum: food vacuole localization of nitric oxide-derived species in intraerythrocytic stages of the malaria parasite. Exp Parasitol 120(1), 29-38 (2008)
http://dx.doi.org/10.1016/j.exppara.2008.04.014
PMid:18504040 PMCid:2574961
15. G Ostera, F Tokumasu, C Teixeira, N Collin, J Sa, J Hume, S Kumar, J Ribeiro, GS Lukat-Rodgers, KR Rodgers: Plasmodium falciparum: Nitric oxide modulates heme speciation in isolated food vacuoles. Exp Parasitol 127(1), 1-8 (2011)
http://dx.doi.org/10.1016/j.exppara.2010.05.006
PMid:20493843
16. N Marchesini, S Luo, CO Rodrigues, SN Moreno, R Docampo: Acidocalcisomes and a vacuolar H⁺-pyrophosphatase in malaria parasites. Biochem J 347(Pt 1), 243-253 (2000)
http://dx.doi.org/10.1042/0264-6021:3470243
PMid:10727425 PMCid:1220954
17. R Docampo, W de Souza, K Miranda, P Rohloff, SNJ Moreno: Acidocalcisomes conserved from bacteria to man. Nat Rev Micro 3(3), 251-261 (2005)
http://dx.doi.org/10.1038/nrmicro1097
PMid:15738951
18. GA Biagini, PG Bray, DG Spiller, MR White, SA Ward: The digestive food vacuole of the malaria parasite is a dynamic intracellular Ca2+ store. J Biol Chem 278(30), 27910-27915 (2003)
http://dx.doi.org/10.1074/jbc.M304193200
PMid:12740366
19. AM Guimarães, JD Lima, MFB Ribeiro: Ultrastructure of Babesia equi trophozoites isolated in Minas Gerais, Brazil. Pesq Vet Bras 23(3), 101-104 (2003)
20. J Mora-Galindo, F Anaya-Velázquez, S Ramírez-Romo, A González-Robles: Entamoeba histolytica: correlation of assessment methods to measure erythrocyte digestion, and effect of cysteine proteinases inhibitors in HM-1: IMSS and HK-9: NIH strains. Exp Parasitol 108(3-4), 89-100 (2004)
http://dx.doi.org/10.1016/j.exppara.2004.08.005
PMid:15582505
21. B Naudé, JA Brzostowski, AR Kimmel, TE Wellems: Dictyostelium discoideum expresses a malaria chloroquine resistance mechanism upon transfection with mutant, but not wild-type, Plasmodium falciparum transporter PfCRT. J Biol Chem 280(27), 25596-25603 (2005)
http://dx.doi.org/10.1074/jbc.M503227200
PMid:15883156 PMCid:1779819
22. DW Fawcett, PA Conrad, JG Grootenhuis, SP Morzaria: Ultrastructure of the intra-erythrocytic stage of Theileria species from cattle and waterbuck. Tissue Cell 19(5), 643-655 (1987)
http://dx.doi.org/10.1016/0040-8166(87)90071-1
23. M Klemba, W Beatty, I Gluzman, DE Goldberg: Trafficking of plasmepsin II to the food vacuole of the malaria parasite Plasmodium falciparum. J Cell Biol 164(1), 47-56 (2004)
http://dx.doi.org/10.1083/jcb200307147
PMid:14709539 PMCid:2171955
24. M Aikawa: The fine structure of the erythrocytic stages of three avian malarial parasites, Plasmodium fallax, P. lophurae, and P. cathemerium. Am J Trop Med Hyg 15(4), 449-471 (1966)
PMid:5941169
25. DJ Sullivan, IY Gluzman, DG Russell, DE Goldberg: On the molecular mechanism of chloroquine's antimalarial action. Proc Natl Acad Sci USA 93(21), 11865-11870 (1996)
http://dx.doi.org/10.1073/pnas.93.21.11865
26. S Pagola, PW Stephens, DS Bohle, AD Kosar, SK Madsen: The structure of malaria pigment beta haematin. Nature 404(6775), 307-310 (2000)
http://dx.doi.org/10.1038/35005132
PMid:10749217
27. TJ Egan: Haemozoin formation. Mol Biochem Parasitol 157(2), 127-136 (2008)
http://dx.doi.org/10.1016/j.molbiopara.2007.11.005
PMid:18083247
28. CJ Tonkin, JA Pearce, GI McFadden, AF Cowman: Protein targeting to destinations of the secretory pathway in the malaria parasite Plasmodium falciparum. Curr Opin Microbiol 9(4), 381-387 (2006)
http://dx.doi.org/10.1016/j.mib.2006.06.015
PMid:16828333
29. LH Bannister, JM Hopkins, G Margos, AR Dluzewski, GH Mitchell: Three-Dimensional Ultrastructure of the Ring Stage of Plasmodium falciparum: Evidence for Export Pathways. Microsc Microanal 10(5), 551-562 (2004)
http://dx.doi.org/10.1017/S1431927604040917
PMid:15525429
30. AR Dluzewski, IT Ling, JM Hopkins, M Grainger, G Margos, GH Mitchell, AA Holder, LH Bannister: Formation of the food vacuole in Plasmodium falciparum: a potential role for the 19 kDa fragment of merozoite surface protein 1 (MSP119). PLoS One 3(8), e3085 (2008)
http://dx.doi.org/10.1371/journal.pone.0003085
PMid:18769730 PMCid:2518119
31. C Slomianny: Three-dimensional reconstruction of the feeding process of the malaria parasite. Blood Cells 16(2-3), 369 (1990)
PMid:2096983
32. M Aikawa, C Huff, H Spinz: Comparative feeding mechanisms of avian and primate malarial parasites. Mil Med 131(9), Suppl: 969-983 (1966)
PMid:4957848
33. MA Rudzinska, W Trager, R Bray: Pinocytotic Uptake and the Digestion of Hemoglobin in Malaria Parasites. J Eukaryot Microbiol 12(4), 563-576 (1965)
http://dx.doi.org/10.1111/j.1550-7408.1965.tb03256.x
34. DA Elliott, MT McIntosh, HD Hosgood, S Chen, G Zhang, P Baevova, KA Joiner: Four distinct pathways of hemoglobin uptake in the malaria parasite Plasmodium falciparum. Proc Natl Acad Sci USA 105(7), 2463-2468 (2008)
http://dx.doi.org/10.1073/pnas.0711067105
PMid:18263733 PMCid:2268159
35. I Coppens, DJ Sullivan, ST Prigge: An update on the rapid advances in malaria parasite cell biology. Trends Parasitol 26(6), 305-310 (2010)
http://dx.doi.org/10.1016/j.pt.2010.03.007
PMid:20382563 PMCid:2883679
36. NA Bakar, N Klonis, E Hanssen, C Chan, L Tilley: Digestive-vacuole genesis and endocytic processes in the early intraerythrocytic stages of Plasmodium falciparum. J Cell Sci 123(3), 441-450 (2010)
http://dx.doi.org/10.1242/jcs.061499
PMid:20067995
37. MD Lazarus, TG Schneider, TF Taraschi: A new model for hemoglobin ingestion and transport by the human malaria parasite Plasmodium falciparum. J Cell Sci 121(Pt 11), 1937-1949 (2008)
http://dx.doi.org/10.1242/jcs.023150
PMid:18477610
38. A Yayon, R Timberg, S Friedman, H Ginsburg: Effects of Chloroquine on the Feeding Mechanism of the Intraerythrocytic Human Malarial Parasite Plasmodium falciparum. J Eukaryot Microbiol 31(3), 367-372 (1984)
http://dx.doi.org/10.1111/j.1550-7408.1984.tb02981.x
39. PL Olliaro, DE Goldberg: The Plasmodium digestive vacuole: metabolic headquarters and choice drug target. Parasitol Today 11(8), 294-297 (1995)
http://dx.doi.org/10.1016/0169-4758(95)80042-5
40. A Waters, D Higgins, T McCutchan: Plasmodium falciparum appears to have arisen as a result of lateral transfer between avian and human hosts. Proc Natl Acad Sci USA 88(8), 3140-3144 (1991)
http://dx.doi.org/10.1073/pnas.88.8.3140
41. AL Omara-Opyene, PA Moura, CR Sulsona, JA Bonilla, CA Yowell, H Fujioka, DA Fidock, JB Dame: Genetic disruption of the Plasmodium falciparum digestive vacuole plasmepsins demonstrates their functional redundancy. J Biol Chem 279(52), 54088-54096 (2004)
http://dx.doi.org/10.1074/jbc.M409605200
PMid:15491999
42. RA Cooper, MT Ferdig, XZ Su, L Ursos, J Mu, T Nomura, H Fujioka, DA Fidock, PD Roepe, TE Wellems: Alternative mutations at position 76 of the vacuolar transmembrane protein PfCRT are associated with chloroquine resistance and unique stereospecific quinine and quinidine responses in Plasmodium falciparum. Mol Pharmacol 61(1), 35-42 (2002)
http://dx.doi.org/10.1124/mol.61.1.35
PMid:11752204
43. KL Waller, RA Muhle, LM Ursos, P Horrocks, D Verdier-Pinard, ABS Sidhu, H Fujioka, PD Roepe, DA Fidock: Chloroquine resistance modulated in vitro by expression levels of the Plasmodium falciparum chloroquine resistance transporter. J Biol Chem 278(35), 33593-33601 (2003)
http://dx.doi.org/10.1074/jbc.M302215200
PMid:12813054
44. KE Jackson, N Klonis, DJP Ferguson, A Adisa, C Dogovski, L Tilley: Food vacuole associated lipid bodies and heterogeneous lipid environments in the malaria parasite, Plasmodium falciparum. Mol Microbiol 54(1), 109-122 (2004)
http://dx.doi.org/10.1111/j.1365-2958.2004.04284.x
PMid:15458409
45. DJ Sullivan, IY Gluzman, DE Goldberg: Plasmodium hemozoin formation mediated by histidine-rich proteins. Science 271(5246), 219-222 (1996)
http://dx.doi.org/10.1126/science.271.5246.219
PMid:8539625
46. P Parroche, FN Lauw, N Goutagny, E Latz, BG Monks, A Visintin, KA Halmen, M Lamphier, M Olivier, DC Bartholomeu: Malaria hemozoin is immunologically inert but radically enhances innate responses by presenting malaria DNA to Toll-like receptor 9. Proc Natl Acad Sci USA 104(6), 1919-1924 (2007)
http://dx.doi.org/10.1073/pnas.0608745104
PMid:17261807 PMCid:1794278
47. MT Shio, SC Eisenbarth, M Savaria, AF Vinet, M-J Bellemare, KW Harder, FS Sutterwala, DS Bohle, A Descoteaux, RA Flavell, M Olivier: Malarial Hemozoin Activates the NLRP3 Inflammasome through Lyn and Syk Kinases. PLoS Pathog 5(8), e1000559 (2009)
http://dx.doi.org/10.1371/journal.ppat.1000559
PMid:19696895 PMCid:2722371
48. C Slomianny, G Prensier, P Charet: Ingestion of erythrocytic stroma by Plasmodium chabaudi trophozoites: ultrastructural study by serial sectioning and 3-dimensional reconstruction. Parasitology 90(3), 579-587 (1985)
http://dx.doi.org/10.1017/S0031182000055578
49. P Conrad, D Denham, C Brown: Intraerythrocytic multiplication of Theileria parva in vitro: an ultrastructural study. Int J Parasitol 16(3), 223-229 (1986)
http://dx.doi.org/10.1016/0020-7519(86)90047-0
50. S Hellerstein, W Spees, LO Surapathana: Hemoglobin concentration and erythrocyte cation content. J Lab Clin Med 76(1), 10-24 (1970)
PMid:5425358
51. EG Ball, RW McKee, CB Anfinsen, WO Cruz, QM Geiman: Studies on malarial parasites. IX. Chemical and metabolic changes during growth and multiplication in vivo and in vitro. J Biol Chem 175, 547-571 (1948)
PMid:18880753
52. M Krugliak, J Zhang, H Ginsburg: Intraerythrocytic Plasmodium falciparum utilizes only a fraction of the amino acids derived from the digestion of host cell cytosol for the biosynthesis of its proteins. Mol Biochem Parasitol 119(2), 249-256 (2002)
http://dx.doi.org/10.1016/S0166-6851(01)00427-3
53. P Loria, S Miller, M Foley, L Tilley: Inhibition of the peroxidative degradation of haem as the basis of action of chloroquine and other quinoline antimalarials. Biochem J 339(Pt 2), 363-370 (1999)
http://dx.doi.org/10.1042/0264-6021:3390363
PMid:10191268 PMCid:1220166
54. IW Sherman, L Tanigoshi: Incorporation of 14C-amino-acids by malaria (Plasmodium lophurae) IV. In vivo utilization of host cell haemoglobin. Int J Biochem 1(5), 635-637 (1970)
http://dx.doi.org/10.1016/0020-711X(70)90033-9
55. R Theakston, K Fletcher, B Maegraith: The use of electron microscope autoradiography for examining the uptake and degradation of haemoglobin by Plasmodium berghei. Ann Trop Med Parasit 64(1), 63-71 (1970)
PMid:5485712
56. AA Divo, TG Geary, NL Davis, JB Jensen: Nutritional requirements of Plasmodium falciparum in culture. I. Exogenously supplied dialyzable components necessary for continuous growth. J Protozool 32(1), 59-64 (1985)
PMid:3886898
57. H Ginsburg: Some reflections concerning host erythrocyte-malarial parasite interrelationships. Blood Cells 16(2-3), 225-235 (1990)
58. S Zarchin, M Krugliak, H Ginsburg: Digestion of the host erythrocyte by malaria parasites is the primary target for quinoline-containing antimalarials. Biochem Pharmacol 35(14), 2435-2442 (1986)
http://dx.doi.org/10.1016/0006-2952(86)90473-9
59. SA Desai, DJ Krogstad, EW McCleskey: A nutrient-permeable channel on the intraerythrocytic malaria parasite. Nature 362(6421), 643-646 (1993)
http://dx.doi.org/10.1038/362643a0
PMid:7681937
60. VL Lew, L Macdonald, H Ginsburg, M Krugliak, T Tiffert: Excess haemoglobin digestion by malaria parasites: a strategy to prevent premature host cell lysis. Blood Cell Mol Dis 32(3), 353-359 (2004)
http://dx.doi.org/10.1016/j.bcmd.2004.01.006
PMid:15121091
61. HM Staines, JC Ellory, K Kirk: Perturbation of the pump-leak balance for Na+ and K+ in malaria-infected erythrocytes. Am J Physiol Cell Physiol 280(6), C1576-C1587 (2001)
PMid:11350753
62. PJ Rosenthal: Antimalarial Chemotherapy: Mechanisms of Action, Resistance, and New Directions in Drug Discovery. Humana Press, Totowa, NJ (2001)
63. W Trager, J Williams, GS Gill: Extracellular development of erythrocytic stages of Plasmodium falciparum. Methods Cell Sci 18(3), 219-227 (1996)
http://dx.doi.org/10.1007/BF00132887
64. W Trager, J Williams: Extracellular (axenic) development in vitro of the erythrocytic cycle of Plasmodium falciparum. Proc Natl Acad Sci USA 89(12), 5351-5355 (1992)
http://dx.doi.org/10.1073/pnas.89.12.5351
65. W Trager: Loose ends: axenic culture, parasitophorous vacuoles, bird malaria. Parassitologia 44(1-2), 117-121 (2002)
PMid:12404819
66. W Trager, J Williams, GS Gill: Extracellular Development, in vitro, of the Erythrocytic Cycle of Plasmodium falciparum. Parasitol Today 8(11), 384-387 (1992)
http://dx.doi.org/10.1016/0169-4758(92)90177-4
67. GH Coombs, DE Goldberg, M Klemba, C Berry, J Kay, JC Mottram: Aspartic proteases of Plasmodium falciparum and other parasitic protozoa as drug targets. Trends Parasitol 17(11), 532-537 (2001)
http://dx.doi.org/10.1016/S1471-4922(01)02037-2
68. R Banerjee, J Liu, W Beatty, L Pelosof, M Klemba, DE Goldberg: Four plasmepsins are active in the Plasmodium falciparum food vacuole, including a protease with an active-site histidine. Proc Natl Acad Sci USA 99(2), 990-995 (2002)
http://dx.doi.org/10.1073/pnas.022630099
PMid:11782538 PMCid:117418
69. I Gluzman, S Francis, A Oksman, C Smith, K Duffin, D Goldberg: Order and specificity of the Plasmodium falciparum hemoglobin degradation pathway. J Clin Invest 93(4), 1602-1608 (1994)
http://dx.doi.org/10.1172/JCI117140
PMid:8163662 PMCid:294190
70. ME Drew, R Banerjee, EW Uffman, S Gilbertson, PJ Rosenthal, DE Goldberg: Plasmodium Food Vacuole Plasmepsins Are Activated by Falcipains. J Biol Chem 283(19), 12870-12876 (2008)
http://dx.doi.org/10.1074/jbc.M708949200
PMid:18308731 PMCid:2442342
71. BR Shenai, PS Sijwali, A Singh, PJ Rosenthal: Characterization of Native and Recombinant Falcipain-2, a Principal Trophozoite Cysteine Protease and Essential Hemoglobinase of Plasmodium falciparum. J Biol Chem 275(37), 29000-29010 (2000)
http://dx.doi.org/10.1074/jbc.M004459200
PMid:10887194
72. PS Sijwali, BR Shenai, J Gut, A Singh, PJ Rosenthal: Expression and characterization of the Plasmodium falciparum haemoglobinase falcipain-3. Biochem J 360(2), 481-489 (2001)
http://dx.doi.org/10.1042/0264-6021:3600481
73. JB Dame, CA Yowell, L Omara-Opyene, JM Carlton, RA Cooper, T Li: Plasmepsin 4, the food vacuole aspartic proteinase found in all Plasmodium spp. infecting man. Mol Biochem Parasitol 130(1), 1-12 (2003)
http://dx.doi.org/10.1016/S0166-6851(03)00137-3
74. AA Escalante, FJ Ayala: Phylogeny of the malarial genus Plasmodium, derived from rRNA gene sequences. Proc Natl Acad Sci USA 91(24), 11373-11377 (1994)
http://dx.doi.org/10.1073/pnas.91.24.11373
75. MC Leclerc, JP Hugot, P Durand, F Renaud: Evolutionary relationships between 15 Plasmodium species from New and Old World primates (including humans): a 18S rDNA cladistic analysis. Parasitology 129(6), 677-684 (2004)
http://dx.doi.org/10.1017/S0031182004006146
PMid:15648690
76. KG Le Roch, Y Zhou, PL Blair, M Grainger, JK Moch, JD Haynes, P De la Vega, AA Holder, S Batalov, DJ Carucci, EA Winzeler: Discovery of Gene Function by Expression Profiling of the Malaria Parasite Life Cycle. Science 301(5639), 1503-1508 (2003)
http://dx.doi.org/10.1126/science.1087025
PMid:12893887
77. EL Dahl, PJ Rosenthal: Biosynthesis, localization, and processing of falcipain cysteine proteases of Plasmodium falciparum. Mol Biochem Parasitol 139(2), 205-212 (2005)
http://dx.doi.org/10.1016/j.molbiopara.2004.11.009
PMid:15664655
78. Z Bozdech, M Llinás, BL Pulliam, ED Wong, J Zhu, JL DeRisi: The Transcriptome of the Intraerythrocytic Developmental Cycle of Plasmodium falciparum. PLoS Biol 1(1), e5 (2003)
http://dx.doi.org/10.1371/journal.pbio.0000005
PMid:12929205 PMCid:176545
79. KK Eggleson, KL Duffin, DE Goldberg: Identification and characterization of falcilysin, a metallopeptidase involved in hemoglobin catabolism within the malaria parasite Plasmodium falciparum. J Biol Chem 274(45), 32411-32417 (1999)
http://dx.doi.org/10.1074/jbc.274.45.32411
PMid:10542284
80. S Subramanian, PS Sijwali, PJ Rosenthal: Falcipain cysteine proteases require bipartite motifs for trafficking to the Plasmodium falciparum food vacuole. J Biol Chem 282(34), 24961-24969 (2007)
http://dx.doi.org/10.1074/jbc.M703316200
PMid:17565983
81. PVN Dasaradhi, R Korde, JK Thompson, C Tanwar, TC Nag, VS Chauhan, AF Cowman, A Mohmmed, P Malhotra: Food vacuole targeting and trafficking of falcipain-2, an important cysteine protease of human malaria parasite Plasmodium falciparum. Mol Biochem Parasitol 156(1), 12-23 (2007)
http://dx.doi.org/10.1016/j.molbiopara.2007.06.008
PMid:17698213
82. M Llinás, Z Bozdech, ED Wong, AT Adai, JL DeRisi: Comparative whole genome transcriptome analysis of three Plasmodium falciparum strains. Nucleic Acids Res 34(4), 1166-1173 (2006)
http://dx.doi.org/10.1093/nar/gkj517
PMid:16493140 PMCid:1380255
83. M Klemba, I Gluzman, DE Goldberg: A Plasmodium falciparum dipeptidyl aminopeptidase I participates in vacuolar hemoglobin degradation. J Biol Chem 279(41), 43000-43007 (2004)
http://dx.doi.org/10.1074/jbc.M408123200
PMid:15304495
84. CS Gavigan, JP Dalton, A Bell: The role of aminopeptidases in haemoglobin degradation in Plasmodium falciparum-infected erythrocytes. Mol Biochem Parasitol 117(1), 37-48 (2001)
http://dx.doi.org/10.1016/S0166-6851(01)00327-9
85. KA Kolakovich, IY Gluzman, KL Duffin, DE Goldberg: Generation of hemoglobin peptides in the acidic digestive vacuole of Plasmodium falciparum implicates peptide transport in amino acid production. Mol Biochem Parasitol 87(2), 123-135 (1997)
http://dx.doi.org/10.1016/S0166-6851(97)00062-5
86. MF Nankya-Kitaka, GP Curley, CS Gavigan, A Bell, JP Dalton: Plasmodium chabaudi chabaudi and P. falciparum: inhibition of aminopeptidase and parasite growth by bestatin and nitrobestatin. Parasitol Res 84(7), 552-558 (1998)
http://dx.doi.org/10.1007/s004360050447
PMid:9694371
87. GP Curley, SM O'Donovan, J McNally, M Mullally, H O'Hara, A Troy, S-A O'Callaghan, JP Dalton: Aminopeptidases from Plasmodium falciparum, Plasmodium chabaudi chabaudi and Plasmodium berghei. J Eukaryot Microbiol 41(2), 119-123 (1994)
http://dx.doi.org/10.1111/j.1550-7408.1994.tb01483.x
PMid:8167617
88. TS Skinner-Adams, CM Stack, KR Trenholme, CL Brown, J Grembecka, J Lowther, A Mucha, M Drag, P Kafarski, S McGowan, JC Whisstock, DL Gardiner, JP Dalton: Plasmodium falciparum neutral aminopeptidases: new targets for anti-malarials. Trends Biochem Sci 35(1), 53-61 (2010)
http://dx.doi.org/10.1016/j.tibs.2009.08.004
PMid:19796954
89. M Thamotharan, YB Lombardo, SZ Bawani, SA Adibi: An Active Mechanism for Completion of the Final Stage of Protein Degradation in the Liver, Lysosomal Transport of Dipeptides. J Biol Chem 272(18), 11786-11790 (1997)
http://dx.doi.org/10.1074/jbc.272.18.11786
PMid:9115234
90. S Dalal, M Klemba: Roles for Two Aminopeptidases in Vacuolar Hemoglobin Catabolism in Plasmodium falciparum. J Biol Chem 282(49), 35978-35987 (2007)
http://dx.doi.org/10.1074/jbc.M703643200
PMid:17895246
91. D Ragheb, K Bompiani, S Dalal, M Klemba: Evidence for Catalytic Roles for Plasmodium falciparum Aminopeptidase P in the Food Vacuole and Cytosol. J Biol Chem 284(37), 24806-24815 (2009)
http://dx.doi.org/10.1074/jbc.M109.018424
PMid:19574214 PMCid:2757184
92. MB Harbut, G Velmourougane, S Dalal, G Reiss, JC Whisstock, O Onder, D Brisson, S McGowan, M Klemba, DC Greenbaum: Bestatin-based chemical biology strategy reveals distinct roles for malaria M1- and M17-family aminopeptidases. Proc Natl Acad Sci USA 108(34), E526-E534 (2011)
http://dx.doi.org/10.1073/pnas.1105601108
PMid:21844374
93. DL Gardiner, TS Skinner-Adams, CL Brown, KT Andrews, CM Stack, JS McCarthy, JP Dalton, KR Trenholme: Plasmodium falciparum: new molecular targets with potential for antimalarial drug development. Expert Rev Anti Infect Ther 7(9), 1087-1098 (2009)
http://dx.doi.org/10.1586/eri.09.93
PMid:19883329
94. F Teuscher, J Lowther, TS Skinner-Adams, T Spielmann, MWA Dixon, CM Stack, S Donnelly, A Mucha, P Kafarski, S Vassiliou, DL Gardiner, JP Dalton, KR Trenholme: The M18 Aspartyl Aminopeptidase of the Human Malaria Parasite Plasmodium falciparum. J Biol Chem 282(42), 30817-30826 (2007)
http://dx.doi.org/10.1074/jbc.M704938200
PMid:17720817
95. KR Trenholme, CL Brown, TS Skinner-Adams, C Stack, J Lowther, J To, MW Robinson, SM Donnelly, JP Dalton, DL Gardiner: Aminopeptidases of malaria parasites: new targets for chemotherapy. Infect Disord Drug Targets 10(3), 217-225 (2010)
PMid:20334618
96. SK Volkman, AF Cowman, DF Wirth: Functional complementation of the ste6 gene of Saccharomyces cerevisiae with the pfmdr1 gene of Plasmodium falciparum. Proc Natl Acad Sci USA 92(19), 8921-8925 (1995)
http://dx.doi.org/10.1073/pnas.92.19.8921
97. P Rohrbach, CP Sanchez, K Hayton, O Friedrich, J Patel, AB Sidhu, MT Ferdig, DA Fidock, M Lanzer: Genetic linkage of pfmdr1 with food vacuolar solute import in Plasmodium falciparum. EMBO J 25, 3000-3011 (2006)
http://dx.doi.org/10.1038/sj.emboj.7601203
PMid:16794577 PMCid:1500988
98. PJ Rosenthal, SR Meshnick: Hemoglobin catabolism and iron utilization by malaria parasites. Mol Biochem Parasitol 83(2), 131-139 (1996)
http://dx.doi.org/10.1016/S0166-6851(96)02763-6
99. JA Bonilla, TD Bonilla, CA Yowell, H Fujioka, JB Dame: Critical roles for the digestive vacuole plasmepsins of Plasmodium falciparum in vacuolar function. Mol Microbiol 65(1), 64-75 (2007)
http://dx.doi.org/10.1111/j.1365-2958.2007.05768.x
PMid:17581121
100. P Rosenthal, J McKerrow, M Aikawa, H Nagasawa, J Leech: A malarial cysteine proteinase is necessary for hemoglobin degradation by Plasmodium falciparum. J Clin Invest 82(5), 1560-1566 (1988)
http://dx.doi.org/10.1172/JCI113766
PMid:3053784 PMCid:442723
101. PS Sijwali, PJ Rosenthal: Gene disruption confirms a critical role for the cysteine protease falcipain-2 in hemoglobin hydrolysis by Plasmodium falciparum. Proc Natl Acad Sci USA 101(13), 4384-4389 (2004)
http://dx.doi.org/10.1073/pnas.0307720101
PMid:15070727 PMCid:384756
102. PS Sijwali, J Koo, N Singh, PJ Rosenthal: Gene disruptions demonstrate independent roles for the four falcipain cysteine proteases of Plasmodium falciparum. Mol Biochem Parasitol 150(1), 96-106 (2006)
http://dx.doi.org/10.1016/j.molbiopara.2006.06.013
PMid:16890302
103. TS Skinner-Adams, J Lowther, F Teuscher, CM Stack, J Grembecka, A Mucha, P Kafarski, KR Trenholme, JP Dalton, DL Gardiner: Identification of Phosphinate Dipeptide Analog Inhibitors Directed against the Plasmodium falciparum M17 Leucine Aminopeptidase as Lead Antimalarial Compounds. J Med Chem 50(24), 6024-6031 (2007)
http://dx.doi.org/10.1021/jm070733v
PMid:17960925
104. CS Gavigan, SG Machado, JP Dalton, A Bell: Analysis of antimalarial synergy between bestatin and endoprotease inhibitors using statistical response-surface modelling. Antimicrob Agents Chemother 45(11), 3175-3181 (2001)
http://dx.doi.org/10.1128/AAC.45.11.3175-3181.2001
PMid:11600374 PMCid:90800
105. N Campanale, C Nickel, CA Daubenberger, DA Wehlan, JJ Gorman, N Klonis, K Becker, L Tilley: Identification and characterization of heme-interacting proteins in the malaria parasite, Plasmodium falciparum. J Biol Chem 278(30), 27354-27361 (2003)
http://dx.doi.org/10.1074/jbc.M303634200
PMid:12748176
106. H Ginsburg, O Famin, J Zhang, M Krugliak: Inhibition of gluthathione-dependent degradation of heme by chloroquine and amadiaquine as a possible basis for their antimalarial mode of action. Biochem Pharmacol 56(10), 1305-1313 (1998)
http://dx.doi.org/10.1016/S0006-2952(98)00184-1
107. DE Goldberg, A Slater, R Beavis, B Chait, A Cerami, G Henderson: Hemoglobin degradation in the human malaria pathogen Plasmodium falciparum: a catabolic pathway initiated by a specific aspartic protease. J Exp Med 173(4), 961-969 (1991)
http://dx.doi.org/10.1084/jem.173.4.961
PMid:2007860
108. A Dorn, SR Vippagunta, H Matile, C Jaquet, JL Vennerstrom, RG Ridley: An assessment of drug-haematin binding as a mechanism for inhibition of haematin polymerisation by quinoline antimalarials. Biochem Pharmacol 55(6), 727-736 (1998)
http://dx.doi.org/10.1016/S0006-2952(97)00510-8
109. AV Pandey, N Singh, BL Tekwani, SK Puri, VS Chauhan: Assay of beta-hematin formation by malaria parasite. J Pharm Biomed Anal 20(1-2), 203-207 (1999)
http://dx.doi.org/10.1016/S0731-7085(99)00021-7
110. I Choi, JL Mego: Purification of Plasmodium falciparum digestive vacuoles and partial characterization of the vacuolar membrane ATPase. Mol Biochem Parasitol 31(1), 71-78 (1988)
http://dx.doi.org/10.1016/0166-6851(88)90146-6
111. KJ Saliba, RJW Allen, S Zissis, PG Bray, SA Ward, K Kirk: Acidification of the Malaria Parasite's Digestive Vacuole by a H+-ATPase and a H+-pyrophosphatase. J Biol Chem 278(8), 5605-5612 (2003)
http://dx.doi.org/10.1074/jbc.M208648200
PMid:12427765
112. Y Moriyama, M Hayashi, S Yatsushiro, A Yamamoto: Vacuolar Proton Pumps in Malaria Parasite Cells. J Bioenerg Biomembr 35(4), 367-375 (2003)
http://dx.doi.org/10.1023/A:1025785000544
PMid:14635782
113. AM Lehane, R Hayward, KJ Saliba, K Kirk: A verapamil-sensitive chloroquine-associated H+ leak from the digestive vacuole in chloroquine-resistant malaria parasites. J Cell Sci 121(10), 1624-1632 (2008)
http://dx.doi.org/10.1242/jcs.016758
PMid:18445688
114. CA Homewood, DC Warhurst, W Peters, VC Baggaley: Lysosomes, pH and the anti-malarial action of chloroquine. Nature 235, 50-52 (1972)
http://dx.doi.org/10.1038/235050a0
PMid:4550396
115. DL Vander Jagt, LA Hunsaker, NM Campos: Comparison of proteases from chloroquine-sensitive and chloroquine-resistant strains of Plasmodium falciparum. Biochem Pharmacol 36(19), 3285-3291 (1987)
http://dx.doi.org/10.1016/0006-2952(87)90646-0
116. DJ Krogstad, PH Schlesinger: The basis of antimalarial action: non-weak base effects of chloroquine on acid vesicle pH. Am J Trop Med Hyg 36(2), 213-220 (1987)
PMid:2435182
117. A Yayon, ZI Cabantchik, H Ginsburg: Susceptibility of human malaria parasites to chloroquine is pH dependent. Proc Natl Acad Sci USA 82(9), 2784-2788 (1985)
http://dx.doi.org/10.1073/pnas.82.9.2784
118. R Hayward, KJ Saliba, K Kirk: The pH of the digestive vacuole of Plasmodium falciparum is not associated with chloroquine resistance. J Cell Sci 119, 1016-1025 (2006)
http://dx.doi.org/10.1242/jcs.02795
PMid:16492710
119. C de Duve, T de Barsy, B Poole, A Trouet, P Tulkens, F Van Hoof: Lysosomotropic agents. Biochem Pharmacol 23(18), 2495-2531 (1974)
http://dx.doi.org/10.1016/0006-2952(74)90174-9
120. A Yayon, ZI Cabantchik, H Ginsburg: Identification of the acidic compartment of Plasmodium falciparum-infected human erythrocytes as the target of the antimalarial drug chloroquine. EMBO J 3(11), 2695-2700 (1984)
PMid:6391917 PMCid:557751
121. H Ginsburg, E Nissani, M Krugliak: Alkalinization of the food vacuole of malaria parasites by quinoline drugs and alkylamines is not correlated with their antimalarial activity. Biochem Pharmacol 38(16), 2645-2654 (1989)
http://dx.doi.org/10.1016/0006-2952(89)90550-9
122. LMB Ursos, SM Dzekunov, PD Roepe: The effects of chloroquine and verapamil on digestive vacuolar pH of P. falciparum either sensitive or resistant to chloroquine. Mol Biochem Parasitol 110(1), 125-134 (2000)
http://dx.doi.org/10.1016/S0166-6851(00)00262-0
123. SM Dzekunov, LMB Ursos, PD Roepe: Digestive vacuolar pH of intact intraerythrocytic P. falciparum either sensitive or resistant to chloroquine. Mol Biochem Parasitol 110(1), 107-124 (2000)
http://dx.doi.org/10.1016/S0166-6851(00)00261-9
124. F Wissing, CP Sanchez, P Rohrbach, S Ricken, M Lanzer: Illumination of the malaria parasite Plasmodium falciparum alters intracellular pH. Implications for live cell imaging. J Biol Chem 277(40), 37747-37755 (2002)
http://dx.doi.org/10.1074/jbc.M204845200
PMid:12140286
125. P Rohrbach: Imaging ion flux and ion homeostasis in blood stage malaria parasites. Biotechnol J 4(6), 812-825 (2009)
http://dx.doi.org/10.1002/biot.200900084
PMid:19507147
126. TN Bennett, AD Kosar, LMB Ursos, S Dzekunov, AB Singh Sidhu, DA Fidock, PD Roepe: Drug resistance-associated pfCRT mutations confer decreased Plasmodium falciparum digestive vacuolar pH. Mol Biochem Parasitol 133(1), 99-114 (2004)
http://dx.doi.org/10.1016/j.molbiopara.2003.09.008
PMid:14668017
127. PG Bray, RE Howells, SA Ward: Vacuolar acidification and chloroquine sensitivity in Plasmodium falciparum. Biochem Pharmacol 43(6), 1219-1227 (1992)
http://dx.doi.org/10.1016/0006-2952(92)90495-5
128. N Klonis, O Tan, K Jackson, D Goldberg, M Klemba, L Tilley: Evaluation of pH during cytostomal endocytosis and vacuolar catabolism of haemoglobin in Plasmodium falciparum. Biochem J 407(3), 343-354 (2007)
http://dx.doi.org/10.1042/BJ20070934
PMid:17696875 PMCid:2275073
129. Y Kuhn, P Rohrbach, M Lanzer: Quantitative pH measurements in Plasmodium falciparum-infected erythrocytes using pHluorin. Cell Microbiol 9(4), 1004-1013 (2007)
http://dx.doi.org/10.1111/j.1462-5822.2006.00847.x
PMid:17381432
130. G Miesenböck, DA De Angelis, JE Rothman: Visualizing secretion and synaptic transmission with pH-sensitive green fluorescent proteins. Nature 394(6689), 192-195 (1998)
http://dx.doi.org/10.1038/28190
PMid:9671304
131. CD Fitch: Plasmodium falciparum in owl monkeys: drug resistance and chloroquine binding capacity. Science 169(3942), 289-290 (1970)
http://dx.doi.org/10.1126/science.169.3942.289
PMid:4988896
132. PG Bray, M Mungthin, RG Ridley, SA Ward: Access to hematin: the basis of chloroquine resistance. Mol Pharmacol 54(1), 170-179 (1998)
PMid:9658203
133. CP Sanchez, W Stein, M Lanzer: Trans stimulation provides evidence for a drug efflux carrier as the mechanism of chloroquine resistance in Plasmodium falciparum. Biochemistry 42(31), 9383-9394 (2003)
http://dx.doi.org/10.1021/bi034269h
PMid:12899625
134. CP Sanchez, JE McLean, P Rohrbach, DA Fidock, WD Stein, M Lanzer: Evidence for a pfcrt-associated chloroquine efflux system in the human malarial parasite Plasmodium falciparum. Biochemistry 44(29), 9862-9870 (2005)
http://dx.doi.org/10.1021/bi050061f
PMid:16026158
135. CP Sanchez, JE McLean, W Stein, M Lanzer: Evidence for a substrate specific and inhibitable drug efflux system in chloroquine resistant Plasmodium falciparum strains. Biochemistry 43(51), 16365-16373 (2004)
http://dx.doi.org/10.1021/bi048241x
PMid:15610031
136. DJ Krogstad, IY Gluzman, DE Kyle, AM Oduola, SK Martin, WK Milhous, PH Schlesinger: Efflux of chloroquine from Plasmodium falciparum: mechanism of chloroquine resistance. Science 238(4831), 1283-1285 (1987)
http://dx.doi.org/10.1126/science.3317830
PMid:3317830
137. JMR Carlton, DA Fidock, A Djimdé, CV Plowe, TE Wellems: Conservation of a novel vacuolar transporter in Plasmodium species and its central role in chloroquine resistance of P. falciparum. Curr Opin Microbiol 4(4), 415-420 (2001)
http://dx.doi.org/10.1016/S1369-5274(00)00228-9
138. DA Fidock, T Nomura, AK Talley, RA Cooper, SM Dzekunov, MT Ferdig, L Ursos, AbS Sidhu, B Naudé, KW Deitsch, X-Z Su, JC Wootton, PD Roepe, TE Wellems: Mutations in the P. falciparum digestive vacuole transmembrane protein PfCRT and evidence for their role in chloroquine resistance. Mol Cell 6(4), 861-871 (2000)
http://dx.doi.org/10.1016/S1097-2765(05)00077-8
139. MB Reed, KJ Saliba, SR Caruana, K Kirk, AF Cowman: Pgh1 modulates sensitivity and resistance to multiple antimalarials in Plasmodium falciparum. Nature 403(6772), 906-909 (2000)
http://dx.doi.org/10.1038/35002615
PMid:10706290
140. CT Hotta, ML Gazarini, FH Beraldo, FP Varotti, C Lopes, RP Markus, T Pozzan, CR Garcia: Calcium-dependent modulation by melatonin of the circadian rhythm in malarial parasites. Nat Cell Biol 2(7), 466-468 (2000)
http://dx.doi.org/10.1038/35017112
PMid:10878815
141. Y Matsumoto, G Perry, L Scheibel, M Aikawa: Role of calmodulin in Plasmodium falciparum: implications for erythrocyte invasion by the merozoite. Eur J Cell Biol 45(1), 36-43 (1987)
PMid:3127214
142. M Wasserman, JP Vernot, PM Mendoza: Role of calcium and erythrocyte cytoskeleton phosphorylation in the invasion of Plasmodium falciparum. Parasitol Res 76(8), 681-688 (1990)
http://dx.doi.org/10.1007/BF00931087
PMid:2251243
143. JG Johnson, N Epstein, T Shiroishi, LH Miller: Factors affecting the ability of isolated Plasmodium knowlesi merozoites to attach to and invade erythrocytes. Parasitology 80(3), 539-550 (1980)
http://dx.doi.org/10.1017/S0031182000000998
144. J Adovelande, B Bastide, J Deleze, J Schrevel: Cytosolic Free Calcium in Plasmodium falciparum-Infected Erythrocytes and the Effect of Verapamil: A Cytofluorometric Study. Exp Parasitol 76(3), 247-258 (1993)
http://dx.doi.org/10.1006/expr.1993.1030
PMid:8500585
145. F Kawamoto, R Alejo-Blanco, SL Fleck, Y Kawamoto, RE Sinden: Possible roles of Ca2+ and cGMP as mediators of the exflagellation of Plasmodium berghei and Plasmodium falciparum. Mol Biochem Parasitol 42(1), 101-108 (1990)
http://dx.doi.org/10.1016/0166-6851(90)90117-5
146. F Kawamoto, H Fujioka, R Murakami, Syafruddin, M Hagiwara, T Ishikawa, H Hidaka: The roles of Ca2+/calmodulin- and cGMP-dependent pathways in gametogenesis of a rodent malaria parasite, Plasmodium berghei. Eur J Cell Biol 60(1), 101-107 (1993)
PMid:8385016
147. ML Gazarini, CR Garcia: The malaria parasite senses cytosolic Ca2+ fluctuations. Biochem Biophys Res Commun 321(1), 138-144 (2004)
http://dx.doi.org/10.1016/j.bbrc.2004.06.141
PMid:15358226
148. ML Gazarini, AP Thomas, T Pozzan, CR Garcia: Calcium signaling in a low calcium environment: how the intracellular malaria parasite solves the problem. J Cell Biol 161(1), 103-110 (2003)
http://dx.doi.org/10.1083/jcb.200212130
PMid:12682086 PMCid:2172890
149. FP Varotti, FH Beraldo, ML Gazarini, CR Garcia: Plasmodium falciparum malaria parasites display a THG-sensitive Ca2+ pool. Cell Calcium 33(2), 137-144 (2003)
http://dx.doi.org/10.1016/S0143-4160(02)00224-5
150. P Rohrbach, O Friedrich, J Hentschel, H Plattner, RH Fink, M Lanzer: Quantitative Calcium Measurements in Subcellular Compartments of Plasmodium falciparum-infected Erythrocytes. J Biol Chem 280(30), 27960-27969 (2005)
http://dx.doi.org/10.1074/jbc.M500777200
PMid:15927958
151. CR Garcia, SE Ann, ES Tavares, AR Dluzewski, WT Mason, FB Paiva: Acidic calcium pools in intraerythrocytic malaria parasites. Eur J Cell Biol 76(2), 133-138 (1998)
PMid:9696353
152. A Rotmann, C Sanchez, A Guiguemde, P Rohrbach, A Dave, N Bakouh, G Planelles, M Lanzer: PfCHA is a mitochondrial divalent cation/H+ antiporter in Plasmodium falciparum. Mol Microbiol 76(6), 1591-1606 (2010)
http://dx.doi.org/10.1111/j.1365-2958.2010.07187.x
PMid:20487273
153. C Garcia, A Dluzewski, L Catalani, R Burting, J Hoyland, W Mason: Calcium homeostasis in intraerythrocytic malaria parasites. Eur J Cell Biol 71(4), 409-413 (1996)
PMid:8980913
154. LM Alleva, K Kirk: Calcium regulation in the intraerythrocytic malaria parasite Plasmodium falciparum. Mol Biochem Parasitol 117(2), 121-128 (2001)
http://dx.doi.org/10.1016/S0166-6851(01)00338-3
155. K Kirk: Channels and transporters as drug targets in the Plasmodium-infected erythrocyte. Acta Trop 89(3), 285-298 (2004)
http://dx.doi.org/10.1016/j.actatropica.2003.10.002
PMid:14744555
156. RE Martin, RI Henry, JL Abbey, JD Clements, K Kirk: The 'permeome' of the malaria parasite: an overview of the membrane transport proteins of Plasmodium falciparum. Genome Biol 6(3), R26 (2005)
http://dx.doi.org/10.1186/gb-2005-6-3-r26
PMid:15774027 PMCid:1088945
157. MI Borges-Walmsley, KS McKeegan, AR Walmsley: Structure and function of efflux pumps that confer resistance to drugs. Biochem J 376(Pt 2), 313-338 (2003)
http://dx.doi.org/10.1042/BJ20020957
PMid:13678421 PMCid:1223791
158. SJ Foote, JK Thompson, AF Cowman, DJ Kemp: Amplification of the multidrug resistance gene in some chloroquine-resistant isolates of P. falciparum. Cell 57(6), 921-930 (1989)
http://dx.doi.org/10.1016/0092-8674(89)90330-9
159. C Wilson, A Serrano, A Wasley, M Bogenschutz, A Shankar, D Wirth: Amplification of a gene related to mammalian mdr genes in drug-resistant Plasmodium falciparum. Science 244(4909), 1184-1186 (1989)
http://dx.doi.org/10.1126/science.2658061
PMid:2658061
160. AF Cowman, S Karcz, D Galatis, JG Culvenor: A P-glycoprotein homologue of Plasmodium falciparum is localized on the digestive vacuole. J Cell Biol 113(5), 1033-1042 (1991)
http://dx.doi.org/10.1083/jcb.113.5.1033
PMid:1674943
161. G Cremer, LK Basco, J Lebras, D Camus, C Slomianny: Plasmodium falciparum: Detection of P-Glycoprotein in Chloroquine-Susceptible and Chloroquine-Resistant Clones and Isolates. Exp Parasitol 81(1), 1-8 (1995)
http://dx.doi.org/10.1006/expr.1995.1086
PMid:7628557
162. SR Karcz, D Galatis, AF Cowman: Nucleotide binding properties of a P-glycoprotein homologue from Plasmodium falciparum. Mol Biochem Parasitol 58(2), 269-276 (1993)
http://dx.doi.org/10.1016/0166-6851(93)90048-3
163. MI Veiga, PE Ferreira, BA Schmidt, U Ribacke, A Björkman, A Tichopad, JP Gil: Antimalarial Exposure Delays Plasmodium falciparum Intra-Erythrocytic Cycle and Drives Drug Transporter Genes Expression. PLoS One 5(8), e12408 (2010)
http://dx.doi.org/10.1371/journal.pone.0012408
PMid:20811640 PMCid:2928296
164. RN Price, AC Uhlemann, A Brockman, R McGready, E Ashley, L Phaipun, R Patel, K Laing, S Looareesuwan, NJ White, F Nosten, S Krishna: Mefloquine resistance in Plasmodium falciparum and increased pfmdr1 gene copy number. Lancet 364, 438-447 (2004)
http://dx.doi.org/10.1016/S0140-6736(04)16767-6
165. ABS Sidhu, A-C Uhlemann, SG Valderramos, J-C Valderramos, S Krishna, DA Fidock: Decreasing pfmdr1 Copy Number in Plasmodium falciparum Malaria Heightens Susceptibility to Mefloquine, Lumefantrine, Halofantrine, Quinine, and Artemisinin. J Infect Dis 194(4), 528-535 (2006)
http://dx.doi.org/10.1086/507115
PMid:16845638 PMCid:2978021
166. AF Cowman, D Galatis, JK Thompson: Selection for mefloquine resistance in Plasmodium falciparum is linked to amplification of the pfmdr1 gene and cross-resistance to halofantrine and quinine. Proc Natl Acad Sci USA 91(3), 1143-1147 (1994)
http://dx.doi.org/10.1073/pnas.91.3.1143
167. CM Wilson, SK Volkman, S Thaithong, RK Martin, DE Kyle, WK Milhous, DF Wirth: Amplification of pfmdr1 associated with mefloquine and halofantrine resistance in Plasmodium falciparum from Thailand. Mol Biochem Parasitol 57(1), 151-160 (1993)
http://dx.doi.org/10.1016/0166-6851(93)90252-S
168. RN Price, C Cassar, A Brockman, M Duraisingh, M van Vugt, NJ White, F Nosten, S Krishna: The pfmdr1 Gene Is Associated with a Multidrug-Resistant Phenotype in Plasmodium falciparum from the Western Border of Thailand. Antimicrob Agents Chemother 43(12), 2943-2949 (1999)
PMid:10582887 PMCid:89592
169. AMJ Oduola, WK Milhous, NF Weatherly, JH Bowdre, RE Desjardins: Plasmodium falciparum: Induction of resistance to mefloquine in cloned strains by continuous drug exposure in vitro. Exp Parasitol 67(2), 354-360 (1988)
http://dx.doi.org/10.1016/0014-4894(88)90082-3
170. A Djimdé, OK Doumbo, JF Cortese, K Kayentao, S Doumbo, Y Diourté, A Dicko, X-z Su, T Nomura, DA Fidock, TE Wellems, CV Plowe, D Coulibaly: A Molecular Marker for Chloroquine-Resistant Falciparum Malaria. N Eng J Med 344(4), 257-263 (2001)
http://dx.doi.org/10.1056/NEJM200101253440403
PMid:11172152
171. IS Adagut, DC Warhust: Plasmodium falciparum: linkage disequilibrium between loci in chromosomes 7 and 5 and chloroquine selective pressure in Northern Nigeria. Parasitology 123(3), 219-224 (2001)
PMid:11578085
172. HA Babiker, SJ Pringle, A Abdel-Muhsin, M Mackinnon, P Hunt, D Walliker: High-Level Chloroquine Resistance in Sudanese Isolates of Plasmodium falciparum Is Associated with Mutations in the Chloroquine Resistance Transporter Gene pfcrt and the Multidrug Resistance Gene pfmdr1. J Infect Dis 183(10), 1535-1538 (2001)
http://dx.doi.org/10.1086/320195
PMid:11319692
173. S Picot, P Olliaro, F de Monbrison, A-L Bienvenu, R Price, P Ringwald: A systematic review and meta-analysis of evidence for correlation between molecular markers of parasite resistance and treatment outcome in falciparum malaria. Malar J 8(1), 89 (2009)
http://dx.doi.org/10.1186/1475-2875-8-89
PMid:19413906 PMCid:2681474
174. T Ngo, M Duraisingh, M Reed, DB Hipgrave, B-A Biggs, AF Cowman: Analysis of pfcrt, pfmdr1, dhfr, and dhps mutations and drug sensitivities in Plasmodium falciparum isolates from patients in Vietnam before and after treatment with artemisinin. Am J Trop Med Hyg 68(3), 350-356 (2003)
PMid:12685644
175. MT Duraisingh, P Refour: Multiple drug resistance genes in malaria - from epistasis to epidemiology. Mol Microbiol 57(4), 874-877 (2005)
http://dx.doi.org/10.1111/j.1365-2958.2005.04748.x
PMid:16091030
176. HH van Es, S Karcz, F Chu, AF Cowman, S Vidal, P Gros, E Schurr: Expression of the plasmodial pfmdr1 gene in mammalian cells is associated with increased susceptibility to chloroquine. Mol Cell Biol 14(4), 2419-2428 (1994)
PMid:7511206 PMCid:358609
177. RE Martin, K Kirk: The malaria parasite's chloroquine resistance transporter is a member of the drug/metabolite transporter superfamily. Mol Biol Evol 21(10), 1938-1949 (2004)
http://dx.doi.org/10.1093/molbev/msh205
PMid:15240840
178. CV Tran, MH Saier Jr: The principal chloroquine resistance protein of Plasmodium falciparum is a member of the drug/metabolite transporter superfamily. Microbiology 150(1), 1-3 (2004)
http://dx.doi.org/10.1099/mic.0.26818-0
PMid:14702390
179. PG Bray, M Mungthin, IM Hastings, GA Biagini, DK Saidu, V Lakshmanan, DJ Johnson, RH Hughes, PA Stocks, PM O'Neill, DA Fidock, DC Warhurst, SA Ward: PfCRT and the trans-vacuolar proton electrochemical gradient: regulating the access of chloroquine to ferriprotoporphyrin IX. Mol Microbiol 62(1), 238-251 (2006)
http://dx.doi.org/10.1111/j.1365-2958.2006.05368.x
PMid:16956382 PMCid:2943415
180. RE Martin, RV Marchetti, AI Cowan, SM Howitt, S Bröer, K Kirk: Chloroquine Transport via the Malaria Parasite's Chloroquine Resistance Transporter. Science 325(5948), 1680-1682 (2009)
http://dx.doi.org/10.1126/science.1175667
PMid:19779197
181. CP Sanchez, P Rohrbach, JE McLean, DA Fidock, WD Stein, M Lanzer: Differences in trans-stimulated chloroquine efflux kinetics are linked to PfCRT in Plasmodium falciparum. Mol Microbiol 64(2), 407-420 (2007)
http://dx.doi.org/10.1111/j.1365-2958.2007.05664.x
PMid:17493125 PMCid:2944662
182. JE Hyde: Drug-resistant malaria. Trends Parasitol 21(11), 494-498 (2005)
http://dx.doi.org/10.1016/j.pt.2005.08.020
PMid:16140578 PMCid:2722032
183. CP Sanchez, A Dave, WD Stein, M Lanzer: Transporters as mediators of drug resistance in Plasmodium falciparum. Int J Parasitol 40(10), 1109-1118 (2010)
http://dx.doi.org/10.1016/j.ijpara.2010.04.001
PMid:20399785
184. ABS Sidhu, D Verdier-Pinard, DA Fidock: Chloroquine resistance in Plasmodium falciparum malaria parasites conferred by pfcrt mutations. Science 298(5591), 210-213 (2002)
http://dx.doi.org/10.1126/science.1074045
PMid:12364805 PMCid:2954758
185. V Lakshmanan, PG Bray, D Verdier-Pinard, DJ Johnson, P Horrocks, RA Muhle, GE Alakpa, RH Hughes, SA Ward, DJ Krogstad, ABS Sidhu, DA Fidock: A critical role for PfCRT K76T in Plasmodium falciparum verapamil-reversible chloroquine resistance. EMBO J 24(13), 2294-2305 (2005)
http://dx.doi.org/10.1038/sj.emboj.7600681
PMid:15944738 PMCid:1173140
186. JM Sá, O Twu, K Hayton, S Reyes, MP Fay, P Ringwald, TE Wellems: Geographic patterns of Plasmodium falciparum drug resistance distinguished by differential responses to amodiaquine and chloroquine. Proc Natl Acad Sci USA 106(45), 18883-18889 (2009)
http://dx.doi.org/10.1073/pnas.0911317106
PMid:19884511 PMCid:2771746
187. S Yatsushiro, S Taniguchi, T Mitamura, H Omote, Y Moriyama: Proteolipid of vacuolar H+-ATPase of Plasmodium falciparum: cDNA cloning, gene organization and complementation of a yeast null mutant. Biochim Biophys Acta 1717(2), 89-96 (2005)
http://dx.doi.org/10.1016/j.bbamem.2005.08.011
PMid:16293223
188. SR Karcz, VR Herrmann, AF Cowman: Cloning and characterization of a vacuolar ATPase A subunit homologue from Plasmodium falciparum. Mol Biochem Parasitol 58(2), 333-344 (1993)
http://dx.doi.org/10.1016/0166-6851(93)90056-4
189. SR Karcz, VR Herrmann, F Trottein, AF Cowman: Cloning and characterization of the vacuolar ATPase B subunit from Plasmodium falciparum. Mol Biochem Parasitol 65(1), 123-133 (1994)
http://dx.doi.org/10.1016/0166-6851(94)90121-X
190. BL Herwaldt, PH Schlesinger, DJ Krogstad: Accumulation of chloroquine by membrane preparations from Plasmodium falciparum. Mol Biochem Parasitol 42(2), 257-267 (1990)
http://dx.doi.org/10.1016/0166-6851(90)90169-M
191. DJ Krogstad, IY Gluzman, BL Herwaldt, PH Schlesinger, TE Wellems: Energy dependence of chloroquine accumulation and chloroquine efflux in Plasmodium falciparum. Biochem Pharmacol 43(1), 57-62 (1992)
http://dx.doi.org/10.1016/0006-2952(92)90661-2
192. PA Rea, RJ Poole: Vacuolar H+-Translocating Pyrophosphatase. Annu Rev Plant Physiol Plant Mol Biol 44(1), 157-180 (1993)
http://dx.doi.org/10.1146/annurev.pp.44.060193.001105
193. MT McIntosh, AB Vaidya: Vacuolar type H+ pumping pyrophosphatases of parasitic protozoa. Int J Parasitol 32(1), 1-14 (2002)
http://dx.doi.org/10.1016/S0020-7519(01)00325-3
194. DA Scott, W de Souza, M Benchimol, L Zhong, H-G Lu, SNJ Moreno, R Docampo: Presence of a Plant-like Proton-pumping Pyrophosphatase in Acidocalcisomes of Trypanosoma cruzi. J Biol Chem 273(34), 22151-22158 (1998)
http://dx.doi.org/10.1074/jbc.273.34.22151
PMid:9705361
195. CO Rodrigues, DA Scott, R Docampo: Characterization of a Vacuolar Pyrophosphatase in Trypanosoma brucei and Its Localization to Acidocalcisomes. Mol Cell Biol 19(11), 7712-7723 (1999)
PMid:10523660 PMCid:84816
196. CO Rodrigues, DA Scott, R Docampo: Presence of a vacuolar H+-pyrophosphatase in promastigotes of Leishmania donovani and its localization to a different compartment from the vacuolar H+-ATPase. Biochem J 340(3), 759-766 (1999)
http://dx.doi.org/10.1042/0264-6021:3400759
PMid:10359662 PMCid:1220309
197. MT McIntosh, YM Drozdowicz, K Laroiya, PA Rea, AB Vaidya: Two classes of plant-like vacuolar-type H+-pyrophosphatases in malaria parasites. Mol Biochem Parasitol 114(2), 183-195 (2001)
http://dx.doi.org/10.1016/S0166-6851(01)00251-1
198. S Krishna, A-C Uhlemann, RK Haynes: Artemisinins: mechanisms of action and potential for resistance. Drug Resist Updat 7(4-5), 233-244 (2004)
http://dx.doi.org/10.1016/j.drup.2004.07.001
PMid:15533761
199. K Simons, J Gruenberg: Jamming the endosomal system: lipid rafts and lysosomal storage diseases. Trends Cell Biol 10(11), 459-462 (2000)
http://dx.doi.org/10.1016/S0962-8924(00)01847-X
200. K Malathi, K Higaki, AH Tinkelenberg, DA Balderes, D Almanzar-Paramio, LJ Wilcox, N Erdeniz, F Redican, M Padamsee, Y Liu, S Khan, F Alcantara, ED Carstea, JA Morris, SL Sturley: Mutagenesis of the putative sterol-sensing domain of yeast Niemann Pick C-related protein reveals a primordial role in subcellular sphingolipid distribution. J Cell Biol 164(4), 547-556 (2004)
http://dx.doi.org/10.1083/jcb.200310046
PMid:14970192 PMCid:2171978
201. M Tabuchi, T Yoshimori, K Yamaguchi, T Yoshida, F Kishi: Human NRAMP2/DMT1, Which Mediates Iron Transport across Endosomal Membranes, Is Localized to Late Endosomes and Lysosomes in HEp-2 Cells. J Biol Chem 275(29), 22220-22228 (2000)
http://dx.doi.org/10.1074/jbc.M001478200
PMid:10751401
202. RF Waller, MB Reed, AF Cowman, GI McFadden: Protein trafficking to the plastid of Plasmodium falciparum is via the secretory pathway. EMBO J 19(8), 1794-1802 (2000)
http://dx.doi.org/10.1093/emboj/19.8.1794
PMid:10775264 PMCid:302007
203. ME Wickham, M Rug, SA Ralph, N Klonis, GI McFadden, L Tilley, AF Cowman: Trafficking and assembly of the cytoadherence complex in Plasmodium falciparum-infected human erythrocytes. EMBO J 20(20), 5636-5649 (2001)
http://dx.doi.org/10.1093/emboj/20.20.5636
PMid:11598007 PMCid:125667
204. GG van Dooren, M Marti, CJ Tonkin, LM Stimmler, AF Cowman, GI McFadden: Development of the endoplasmic reticulum, mitochondrion and apicoplast during the asexual life cycle of Plasmodium falciparum. Mol Microbiol 57(2), 405-419 (2005)
http://dx.doi.org/10.1111/j.1365-2958.2005.04699.x
PMid:15978074
205. SE Francis, R Banerjee, DE Goldberg: Biosynthesis and maturation of the malaria aspartic hemoglobinases plasmepsins I and II. J Biol Chem 272(23), 14961-14968 (1997)
http://dx.doi.org/10.1074/jbc.272.23.14961
PMid:9169469
206. SE Francis, I Gluzman, A Oksman, A Knickerbocker, R Mueller, M Bryant, D Sherman, D Russell, D Goldberg: Molecular characterization and inhibition of a Plasmodium falciparum aspartic hemoglobinase. EMBO J 13(2), 306-317 (1994)
PMid:8313875 PMCid:394809
207. R Banerjee, SE Francis, DE Goldberg: Food vacuole plasmepsins are processed at a conserved site by an acidic convertase activity in Plasmodium falciparum. Mol Biochem Parasitol 129(2), 157-165 (2003)
http://dx.doi.org/10.1016/S0166-6851(03)00119-1
208. DR Drew, PR Sanders, BS Crabb: Plasmodium falciparum merozoite surface protein 8 is a ring-stage membrane protein that localizes to the parasitophorous vacuole of infected erythrocytes. Infect Immun 73(7), 3912-3922 (2005)
http://dx.doi.org/10.1128/IAI.73.7.3912-3922.2005
PMid:15972477 PMCid:1168550
209. Y Kuhn, CP Sanchez, D Ayoub, T Saridaki, A Van Dorsselaer, M Lanzer: Trafficking of the Phosphoprotein PfCRT to the Digestive Vacuolar Membrane in Plasmodium falciparum. Traffic 11(2), 236-249 (2010)
http://dx.doi.org/10.1111/j.1600-0854.2009.01018.x
PMid:20015114
210. MT McIntosh, A Vaid, HD Hosgood, J Vijay, A Bhattacharya, MH Sahani, P Baevova, KA Joiner, P Sharma: Traffic to the Malaria Parasite Food Vacuole. J Biol Chem 282(15), 11499-11508 (2007)
http://dx.doi.org/10.1074/jbc.M610974200
PMid:17289673
211. M Henry, S Alibert, E Orlandi-Pradines, H Bogreau, T Fusai, C Rogier, J Barbe, B Pardines: Chloroquine resistance reversal agents as promising antimalarial drugs. Curr Drug Targets 7(8), 935-948 (2006)
http://dx.doi.org/10.2174/138945006778019372
PMid:16918322
212. GD Shanks: Treatment of falciparum malaria in the age of drug resistance. J Postgrad Med 52(4), 277-280 (2006)
PMid:17102546
213. P Winstanley, S Ward: Malaria chemotherapy. Adv Parasitol 61, 47-76 (2006)
http://dx.doi.org/10.1016/S0065-308X(05)61002-0
214. A Leed, K DuBay, LM Ursos, D Sears, AC de Dios, PD Roepe: Solution structures of antimalarial drug-heme complexes. Biochemistry 41(32), 10245-10255 (2002)
http://dx.doi.org/10.1021/bi020195i
PMid:12162739
215. Y Zhang, E Hempelmann: Lysis of malarial parasites and erythrocytes by ferriprotoporphyrin IX-chloroquine and the inhibition of this effect by proteins. Biochem Pharmacol 36(8), 1267-1273 (1987)
http://dx.doi.org/10.1016/0006-2952(87)90080-3
216. CD Fitch, R Chevli, HS Banyal, G Philipps: Lysis of Plasmodium falciparum by ferriprotoporphyrin IX and a chloroquine-ferriprotoporphyrin IX complex. Antimicrob Agents Chemother 21, 819-822 (1982)
PMid:7049079 PMCid:182018
217. DJ Krogstad, PH Schlesinger, IY Gluzman: Antimalarials increase vesicle pH in Plasmodium falciparum. J Cell Biol 101(6), 2302-2309 (1985)
http://dx.doi.org/10.1083/jcb.101.6.2302
PMid:3905824
218. TG Geary, JB Jensen, H Ginsburg: Uptake of (3H)chloroquine by drug-sensitive and -resistant strains of the human malaria parasite Plasmodium falciparum. Biochem Pharmacol 35(21), 3805-3812 (1986)
http://dx.doi.org/10.1016/0006-2952(86)90668-4
219. SK Martin, AM Oduola, WK Milhous: Reversal of chloroquine resistance in Plasmodium falciparum by verapamil. Science 235(4791), 899-901 (1987)
http://dx.doi.org/10.1126/science.3544220
PMid:3544220
220. W Chaijaroenkul, SA Ward, M Mungthin, D Johnson, A Owen, PG Bray, K Na-Bangchang: Sequence and gene expression of chloroquine resistance transporter (pfcrt) in the association of in vitro drugs resistance of Plasmodium falciparum. Malar J 10(42) (2011)
221. DA Fidock, RT Eastman, SA Ward, SR Meshnick: Recent highlights in antimalarial drug resistance and chemotherapy research. Trends Parasitol 24(12), 537-544 (2008)
http://dx.doi.org/10.1016/j.pt.2008.09.005
PMid:18938106 PMCid:2718548
222. SR Hawley, PG Bray, M Mungthin, JD Atkinson, PM O'Neill, SA Ward: Relationship between Antimalarial Drug Activity, Accumulation, and Inhibition of Heme Polymerization in Plasmodium falciparum In Vitro. Antimicrob Agents Chemother 42(3), 682-686 (1998)
PMid:9517951 PMCid:105517
223. M Mungthin, PG Bray, RG Ridley, SA Ward: Central Role of Hemoglobin Degradation in Mechanisms of Action of 4-Aminoquinolines, Quinoline Methanols, and Phenanthrene Methanols. Antimicrob Agents Chemother 42(11), 2973-2977 (1998)
PMid:9797235 PMCid:105975
224. ER Watkins, SR Meshnick: Drugs for malaria. Sem Pediatr Infect Dis 11(3), 202-212 (2000)
http://dx.doi.org/10.1053/pi.2000.6232
225. DJ Krogstad, PH Schlesinger: A perspective on antimalarial action: Effects of weak bases on Plasmodium falciparum. Biochem Pharmacol 35(4), 547-552 (1986)
http://dx.doi.org/10.1016/0006-2952(86)90345-X
226. PG Bray, SR Hawley, M Mungthin, SA Ward: Physicochemical properties correlated with drug resistance and the reversal of drug resistance in Plasmodium falciparum. Mol Pharmacol 50(6), 1559-1566 (1996)
PMid:8967978
227. E Ochong, I van den Broek, K Keus, A Nzila: Short Report: Association Between Chloroquine and Amodiaquine Resistance and Allelic Variation in the Plasmodium Falciparum Multiple Drug Resistance 1 Gene and the Chloroquine Resistance Transporter Gene in Isolates from the Upper Nile in Southern Sudan. Am J Trop Med Hyg 69(2), 184-187 (2003)
PMid:13677373
228. G Holmgren, JP Gil, PM Ferreira, MI Veiga, CO Obonyo, A Björkman: Amodiaquine resistant Plasmodium falciparum malaria in vivo is associated with selection of pfcrt 76T and pfmdr1 86Y. Infect Genet Evol 6(4), 309-314 (2006)
http://dx.doi.org/10.1016/j.meegid.2005.09.001
PMid:16271310
229. J Yuan, KC-C Cheng, RL Johnson, R Huang, S Pattaradilokrat, A Liu, R Guha, DA Fidock, J Inglese, TE Wellems, CP Austin, X-z Su: Chemical Genomic Profiling for Antimalarial Therapies, Response Signatures, and Molecular Targets. Science 333(6043), 724-729 (2011)
http://dx.doi.org/10.1126/science.1205216
PMid:21817045
230. AM Dondorp, S Yeung, L White, C Nguon, NPJ Day, D Socheat, L von Seidlein: Artemisinin resistance: current status and scenarios for containment. Nat Rev Micro 8(4), 272-280 (2010)
PMid:20208550
231. RT Eastman, DA Fidock: Artemisinin-based combination therapies: a vital tool in efforts to eliminate malaria. Nat Rev Micro 7(12), 864-874 (2009)
PMid:19881520 PMCid:2901398
232. H Noedl, D Socheat, W Satimai: Artemisinin-Resistant Malaria in Asia. N Eng J Med 361(5), 540-541 (2009)
http://dx.doi.org/10.1056/NEJMc0900231
PMid:19641219
233. AM Dondorp, F Nosten, P Yi, D Das, AP Phyo, J Tarning, KM Lwin, F Ariey, W Hanpithakpong, SJ Lee, P Ringwald, K Silamut, M Imwong, K Chotivanich, P Lim, T Herdman, SS An, S Yeung, P Singhasivanon, NPJ Day, N Lindegardh, D Socheat, NJ White: Artemisinin Resistance in Plasmodium falciparum Malaria. N Eng J Med 361(5), 455-467 (2009)
http://dx.doi.org/10.1056/NEJMoa0808859
PMid:19641202
234. SG Valderramos, DA Fidock: Transporters involved in resistance to antimalarial drugs. Trends Pharmacol Sci 27(11), 594-601 (2006)
http://dx.doi.org/10.1016/j.tips.2006.09.005
PMid:16996622 PMCid:2944664
235. PM O'Neill, VE Barton, SA Ward: The Molecular Mechanism of Action of Artemisinin - The Debate Continues. Molecules 15(3), 1705-1721 (2010)
http://dx.doi.org/10.3390/molecules15031705
PMid:20336009
236. R Kannan, K Kumar, D Sahal, S Kukreti, VS Chauhan: Reaction of artemisinin with haemoglobin: implications for antimalarial activity. Biochem J 385(Pt 2), 409 (2005)
PMid:15361062 PMCid:1134711
237. PL Olliaro, RK Haynes, B Meunier, Y Yuthavong: Possible modes of action of the artemisinin-type compounds. Trends Parasitol 17(3), 122-126 (2001)
http://dx.doi.org/10.1016/S1471-4922(00)01838-9
238. CL Hartwig, AS Rosenthal, J D'Angelo, CE Griffin, GH Posner, RA Cooper: Accumulation of artemisinin trioxane derivatives within neutral lipids of Plasmodium falciparum malaria parasites is endoperoxide-dependent. Biochem Pharmacol 77(3), 322-336 (2009)
http://dx.doi.org/10.1016/j.bcp.2008.10.015
PMid:19022224 PMCid:2659783
239. SR Krungkrai, Y Yuthavong: The antimalarial action on Plasmodium falciparum of qinghaosu and artesunate in combination with agents which modulate oxidant stress. Trans R Soc Trop Med Hyg 81(5), 710-714 (1987)
http://dx.doi.org/10.1016/0035-9203(87)90003-4
240. A Robert, F Benoit-Vical, C Claparols, B Meunier: The antimalarial drug artemisinin alkylates heme in infected mice. Proc Natl Acad Sci USA 102(38), 13676-13680 (2005)
http://dx.doi.org/10.1073/pnas.0500972102
PMid:16155128 PMCid:1224611
241. DJ Creek, WN Charman, FCK Chiu, RJ Prankerd, Y Dong, JL Vennerstrom, SA Charman: Relationship between Antimalarial Activity and Heme Alkylation for Spiro- and Dispiro-1,2,4-Trioxolane Antimalarials. Antimicrob Agents Chemother 52(4), 1291-1296 (2008)
http://dx.doi.org/10.1128/AAC.01033-07
PMid:18268087 PMCid:2292547
242. Y Wu: How Might Qinghaosu (Artemisinin) and Related Compounds Kill the Intraerythrocytic Malaria Parasite? A Chemist's View. Acc Chem Res 35(5), 255-259 (2002)
http://dx.doi.org/10.1021/ar000080b
PMid:12020162
243. M del Pilar Crespo, TD Avery, E Hanssen, E Fox, TV Robinson, P Valente, DK Taylor, L Tilley: Artemisinin and a Series of Novel Endoperoxide Antimalarials Exert Early Effects on Digestive Vacuole Morphology. Antimicrob Agents Chemother 52(1), 98-109 (2008)
http://dx.doi.org/10.1128/AAC.00609-07
PMid:17938190 PMCid:2223901
244. HC Hoppe, DA van Schalkwyk, UIM Wiehart, SA Meredith, J Egan, BW Weber: Antimalarial Quinolines and Artemisinin Inhibit Endocytosis in Plasmodium falciparum. Antimicrob Agents Chemother 48(7), 2370-2378 (2004)
http://dx.doi.org/10.1128/AAC.48.7.2370-2378.2004
PMid:15215083 PMCid:434207
245. J Bhisutthibhan, SR Meshnick: Immunoprecipitation of (3H) dihydroartemisinin translationally controlled tumor protein (TCTP) adducts from Plasmodium falciparum-infected erythrocytes by using anti-TCTP antibodies. Antimicrob Agents Chemother 45(8), 2397 (2001)
http://dx.doi.org/10.1128/AAC.45.8.2397-2399.2001
PMid:11451708 PMCid:90665
246. J Bhisutthibhan, X-Q Pan, PA Hossler, DJ Walker, CA Yowell, J Carlton, JB Dame, SR Meshnick: The Plasmodium falciparum Translationally Controlled Tumor Protein Homolog and Its Reaction with the Antimalarial Drug Artemisinin. J Biol Chem 273(26), 16192-16198 (1998)
http://dx.doi.org/10.1074/jbc.273.26.16192
PMid:9632675
247. AV Pandey, BL Tekwani, RL Singh, VS Chauhan: Artemisinin, an Endoperoxide Antimalarial, Disrupts the Hemoglobin Catabolism and Heme Detoxification Systems in Malarial Parasite. J Biol Chem 274(27), 19383-19388 (1999)
http://dx.doi.org/10.1074/jbc.274.27.19383
PMid:10383451
248. W Li, W Mo, D Shen, L Sun, J Wang, S Lu, JM Gitschier, B Zhou: Yeast Model Uncovers Dual Roles of Mitochondria in the Action of Artemisinin. PLoS Genet 1(3), e36 (2005)
http://dx.doi.org/10.1371/journal.pgen.0010036
PMid:16170412 PMCid:1201371
249. U Eckstein-Ludwig, RJ Webb, IDA van Goethem, JM East, AG Lee, M Kimura, PM O'Neill, PG Bray, SA Ward, S Krishna: Artemisinins target the SERCA of Plasmodium falciparum. Nature 424(6951), 957-961 (2003)
http://dx.doi.org/10.1038/nature01813
PMid:12931192
250. JM Coterón, D Catterick, J Castro, MJ Chaparro, B Díaz, E Fernández, S Ferrer, FJ Gamo, M Gordo, J Gut, L de las Heras, J Legac, M Marco, J Miguel, V Muñoz, E Porras, JC de la Rosa, JR Ruiz, E Sandoval, P Ventosa, PJ Rosenthal, JM Fiandor: Falcipain Inhibitors: Optimization Studies of the 2-Pyrimidinecarbonitrile Lead Series. J Med Chem 53(16), 6129-6152 (2010)
PMid:20672841
251. I Angulo-Barturen, MB Jiménez-Díaz, T Mulet, J Rullas, E Herreros, S Ferrer, E Jiménez, A Mendoza, J Regadera, PJ Rosenthal, I Bathurst, DL Pompliano, F Gómez de las Heras, D Gargallo-Viola: A Murine Model of falciparum-Malaria by In Vivo Selection of Competent Strains in Non-Myelodepleted Mice Engrafted with Human Erythrocytes. PLoS One 3(5), e2252 (2008)
http://dx.doi.org/10.1371/journal.pone.0002252
PMid:18493601 PMCid:2375113
252. PJ Rosenthal: Characterization of falcipain cysteine proteases and their inhibitors. AntiMal Conference: Antimalarial Drugs - Chemistry, Development & Future Challenges, London, U.K. (2011)
253. S Parikh, J Gut, E Istvan, DE Goldberg, DV Havlir, PJ Rosenthal: Antimalarial Activity of Human Immunodeficiency Virus Type 1 Protease Inhibitors. Antimicrob Agents Chemother 49(7), 2983-2985 (2005)
http://dx.doi.org/10.1128/AAC.49.7.2983-2985.2005
PMid:15980379 PMCid:1168637
254. KT Andrews, DP Fairlie, PK Madala, J Ray, DM Wyatt, PM Hilton, LA Melville, L Beattie, DL Gardiner, RC Reid, MJ Stoermer, T Skinner-Adams, C Berry, JS McCarthy: Potencies of Human Immunodeficiency Virus Protease Inhibitors In Vitro against Plasmodium falciparum and In Vivo against Murine Malaria. Antimicrob Agents Chemother 50(2), 639-648 (2006)
http://dx.doi.org/10.1128/AAC.50.2.639-648.2006
PMid:16436721 PMCid:1366900
255. J Liu, IY Gluzman, ME Drew, DE Goldberg: The Role of Plasmodium falciparum Food Vacuole Plasmepsins. J Biol Chem 280(2), 1432-1437 (2005)
http://dx.doi.org/10.1074/jbc.M409740200
PMid:15513918
256. TS Skinner-Adams, KT Andrews, L Melville, J McCarthy, DL Gardiner: Synergistic interactions of antiretroviral protease inhibitors saquinavir and ritonavir with chloroquine and mefloquine against Plasmodium falciparum in vitro. Antimicrob Agents Chemother 51(2), 759-762 (2007)
http://dx.doi.org/10.1128/AAC.00840-06
PMid:17088482 PMCid:1797772
257. Z He, L Chen, J You, L Qin, X Chen: Antiretroviral protease inhibitors potentiate chloroquine antimalarial activity in malaria parasites by regulating intracellular glutathione metabolism. Exp Parasitol 123(2), 122-127 (2009)
http://dx.doi.org/10.1016/j.exppara.2009.06.008
PMid:19538959
258. Z He, L Qin, L Chen, N Peng, J You, X Chen: Synergy of Human Immunodeficiency Virus Protease Inhibitors with Chloroquine against Plasmodium falciparum in vitro and Plasmodium chabaudi in vivo. Antimicrob Agents Chemother, AAC.01329-07 (2008)
259. KJ Saliba, PI Folb, PJ Smith: Role for the Plasmodium falciparum digestive vacuole in chloroquine resistance. Biochem Pharmacol 56(3), 313-320 (1998)
http://dx.doi.org/10.1016/S0006-2952(98)00140-3
260. M Lamarque, C Tastet, J Poncet, E Demettre, P Jouin, H Vial, J-F Dubremetz: Food vacuole proteome of the malarial parasite Plasmodium falciparum. Proteomics Clin Appl 2(9), 1361-1374 (2008)
http://dx.doi.org/10.1002/prca.200700112
PMid:21136929
261. H Wickert, W Göttler, G Krohne, M Lanzer: Maurer's cleft organization in the cytoplasm of Plasmodium falciparum-infected erythrocytes: new insights from three-dimensional reconstruction of serial ultrathin sections. Eur J Cell Biol 83(10), 567-582 (2004)
http://dx.doi.org/10.1078/0171-9335-00432
PMid:15679102
262. H Wickert, F Wissing, KT Andrews, A Stich, G Krohne, M Lanzer: Evidence for trafficking of PfEMP1 to the surface of P. falciparum-infected erythrocytes via a complex membrane network. Eur J Cell Biol 82(6), 271-284 (2003)
http://dx.doi.org/10.1078/0171-9335-00319
PMid:12868595
263. MT Duraisingh, AF Cowman: Contribution of the pfmdr1 gene to antimalarial drug-resistance. Acta Trop 94(3), 181-190 (2005)
http://dx.doi.org/10.1016/j.actatropica.2005.04.008
PMid:15876420
264. SA Peel: The ABC transporter genes of Plasmodium falciparum and drug resistance. Drug Resist Updat 4(1), 66-74 (2001)
http://dx.doi.org/10.1054/drup.2001.0183
PMid:11512154
265. S Koncarevic, P Rohrbach, M Deponte, G Krohne, JH Prieto, J Yates, S Rahlfs, K Becker: The malarial parasite Plasmodium falciparum imports the human protein peroxiredoxin 2 for peroxide detoxification. Proc Natl Acad Sci USA 106(32), 13323-13328 (2009)
http://dx.doi.org/10.1073/pnas.0905387106
PMid:19666612 PMCid:2726359
Abbreviations: ACT: artemisinin-based combination therapy, AO: acridine orange, AQ: amodiaquine, ART: artemisinin, ATP: adenosine triphosphate, CQ: chloroquine, CQR: chloroquine-resistant, CQS: chloroquine-sensitive, DV: digestive vacuole, ER: endoplasmic reticulum, HF: halofantrine, LF: lumefantrine, MF: mefloquine, NOD: nucleotide-binding oligomerization domain, PPase: pyrophosphatase, PQ: primaquine, PV: parasitophorous vacuole, PVM: parasitophorous vacuole membrane, QC: quinacrine, QD: quinidine, QN: quinine
Key Words: Plasmodium falciparum, Digestive vacuole, Feeding mechanism, Hemoglobin degradation, pH and calcium homeostasis, Transport proteins, Antimalarial drug action, Resistance mechanisms, Isolation procedures, Proteomics, Review
Send correspondence to: Petra Rohrbach, Institute of Parasitology, McGill University, 21 111 Lakeshore Road, Ste. Anne de Bellevue, Quebec, H9X 3V9, Canada, Tel: 514-398-7726, Fax: 514-398-7857, E-mail:petra.rohrbach@mcgill.ca