ORCID Profile
0000-0001-5145-9004
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In Research Link Australia (RLA), "Research Topics" refer to ANZSRC FOR and SEO codes. These topics are either sourced from ANZSRC FOR and SEO codes listed in researchers' related grants or generated by a large language model (LLM) based on their publications.
Biochemistry and Cell Biology | Infectious Agents | Protein Trafficking | Medical Parasitology | Characterisation of Biological Macromolecules | Veterinary Sciences | Sociobiology And Behavioural Ecology | Receptors and Membrane Biology | Signal Transduction | Structural Biology (incl. Macromolecular Modelling) | Parasitology
Expanding Knowledge in the Biological Sciences | Infectious diseases | Neurodegenerative Disorders Related to Ageing | Prevention—biologicals (e.g. vaccines) | Biological sciences | Infectious Diseases |
Publisher: Springer Science and Business Media LLC
Date: 06-2009
DOI: 10.1038/NATURE08104
Publisher: American Chemical Society (ACS)
Date: 12-10-2022
Publisher: The American Association of Immunologists
Date: 15-07-2013
Abstract: The development of effective malaria vaccines and immune biomarkers of malaria is a high priority for malaria control and elimination. Ags expressed by merozoites of Plasmodium falciparum are likely to be important targets of human immunity and are promising vaccine candidates, but very few Ags have been studied. We developed an approach to assess Ab responses to a comprehensive repertoire of merozoite proteins and investigate whether they are targets of protective Abs. We expressed 91 recombinant proteins, located on the merozoite surface or within invasion organelles, and screened them for quality and reactivity to human Abs. Subsequently, Abs to 46 proteins were studied in a longitudinal cohort of 206 Papua New Guinean children to define Ab acquisition and associations with protective immunity. Ab responses were higher among older children and those with active parasitemia. High-level Ab responses to rhoptry and microneme proteins that function in erythrocyte invasion were identified as being most strongly associated with protective immunity compared with other Ags. Additionally, Abs to new or understudied Ags were more strongly associated with protection than were Abs to current vaccine candidates that have progressed to phase 1 or 2 vaccine trials. Combinations of Ab responses were identified that were more strongly associated with protective immunity than responses to their single-Ag components. This study identifies Ags that are likely to be key targets of protective human immunity and facilitates the prioritization of Ags for further evaluation as vaccine candidates and/or for use as biomarkers of immunity in malaria surveillance and control.
Publisher: Wiley
Date: 15-06-2000
Publisher: Springer Science and Business Media LLC
Date: 08-2006
DOI: 10.1038/NRMICRO1465
Abstract: Apicomplexan parasites are an ancient group of protozoan parasites that includes several significant pathogens of humans and animals. To target and invade host cells they use a unique form of actin-based motility, called gliding motility. At the centre of the molecular motor that underlies this unique mode of locomotion are short, highly dynamic actin filaments. Recent molecular work, along with the availability of completed genomes for several Apicomplexa, has highlighted unique features of parasite actin and its regulation - features that might provide new ways to block motility and, consequently, prevent infection and disease.
Publisher: Wiley
Date: 14-10-2005
DOI: 10.1111/J.1365-2958.2005.04895.X
Abstract: During the maturation of intracellular asexual stages of Plasmodium falciparum parasite-encoded proteins are exported into the erythrocyte cytosol. A number of these parasite proteins attach to the host cell cytoskeleton and facilitate transformation of a disk-shaped erythrocyte into a rounded and more rigid infected erythrocyte able to cytoadhere to the vasculature. Knob formation on the surface of infected erythrocytes is critical for this cytoadherence to the host endothelium. P. falciparum proteins have been identified that localize to the parasite-infected erythrocyte membrane: the variant cytoadherence ligand erythrocyte membrane protein 1 (PfEMP1), the knob-associated histidine-rich protein (KAHRP) and the erythrocyte membrane protein 3 (PfEMP3). In this study, we have generated parasites expressing PfEMP3-green fluorescent protein chimeras and identified domains involved in entry to the secretory pathway, export across the parasitophorous vacuolar membrane and attachment to Maurer's clefts and the erythrocyte membrane. Solubility assays, fluorescence photobleaching experiments and immunogold electron microscopy suggest that the exported chimeric proteins are trafficked in a complex rather than in vesicles. This study characterizes elements involved in the tight but transient binding of PfEMP3 to Maurer's clefts and shows that the same elements are necessary for correct assembly under the erythrocyte membrane.
Publisher: Elsevier BV
Date: 25-11-1996
DOI: 10.1016/0166-6851(96)02732-6
Abstract: A photoreactive quinolinemethanol analog, N-[4-[1-hydroxy-2-(dibutylamino)ethyl]quinolin-8yl]-4- azido-2-salicylamide (ASA-MQ) has been synthesized which closely mimics the action of mefloquine. ASA-MQ possesses potent antimalarial activity against a mefloquine-sensitive strain of Plasmodium falciparum and shows decreased activity against a mefloquine-resistant parasite strain. Radioiodinated ASA-MQ has been used in photoaffinity labeling studies to identify mefloquine-interacting proteins in serum, uninfected erythrocytes and Plasmodium falciparum-infected erythrocytes. We have shown that mefloquine interacts specifically with apo-A1, the major protein of serum high density lipoproteins. In addition, mefloquine was shown to interact specifically with the erythrocyte membrane protein, band 7.2b (stomatin). A further two high affinity mefloquine-binding proteins with apparent molecular masses of 22 and 36 kDa were identified in three different strains of Plasmodium falciparum. We suggest that these two mefloquine-binding parasite proteins may be involved in the uptake of mefloquine or may represent macromolecular targets of mefloquine action in malaria parasites.
Publisher: Elsevier BV
Date: 05-2001
DOI: 10.1016/S0166-6851(01)00266-3
Abstract: Adherence of Plasmodium falciparum-infected erythrocytes to the post-capillary endothelium is an important characteristic of malaria infection. The adhesion is mediated predominantly by P. falciparum Erythrocyte Membrane Protein-1 (PfEMP1), a clonally variant protein expressed on the surface of infected red blood cells that appears to be a target of protective immunity. A multi-membered var gene family encodes PfEMP1 and switching expression of different var genes conveys different antigenic and adhesive properties to infected red blood cells. Knowledge about transcriptional control of phenotypic expression, or the mechanisms that allow multiple binding specificities, is very limited. Here, we describe a series of phenotypic selection experiments, which resulted in the expression of different PfEMP1 and the detection of multiple full-length var gene transcripts in the mature trophozoite stage. However, a dominant form of PfEMP1 appeared to be expressed, which suggested that most var transcripts do not lead to a surface expressed PfEMP1 molecule. Parasites bound to specific receptors still expressed multiple full-length var genes and mature trophozoites selected for increased adhesion to a specific receptor retained the ability to bind to multiple receptors. Our findings suggest that a defined adhesive phenotype can be associated with expression of multiple var genes.
Publisher: Springer Science and Business Media LLC
Date: 18-03-2019
DOI: 10.1038/S41598-019-40718-Z
Abstract: Malaria remains a major burden world-wide, but the disease-causing parasites from the genus Plasmodium are difficult to study in vitro . Owing to the small size of the parasites, subcellular imaging poses a major challenge and the use of super-resolution techniques has been hindered by the parasites’ sensitivity to light. This is particularly apparent during the blood-stage of the Plasmodium life cycle, which presents an important target for drug research. The iron-rich food vacuole of the parasite undergoes disintegration when illuminated with high-power lasers such as those required for high resolution in Stimulated Emission Depletion (STED) microscopy. This causes major damage to the s le precluding the use of this super-resolution technique. Here we present guided STED, a novel adaptive illumination (AI) STED approach, which takes advantage of the highly-reflective nature of the iron deposit in the cell to identify the most light-sensitive parts of the s le. Specifically in these parts, the high-power STED laser is deactivated automatically to prevent local damage. Guided STED nanoscopy finally allows super-resolution imaging of the whole Plasmodium life cycle, enabling multicolour imaging of blood-stage malaria parasites with resolutions down to 35 nm without s le destruction.
Publisher: Rockefeller University Press
Date: 17-09-2012
Abstract: Malaria is a major disease of humans caused by protozoan parasites from the genus Plasmodium. It has a complex life cycle however, asexual parasite infection within the blood stream is responsible for all disease pathology. This stage is initiated when merozoites, the free invasive blood-stage form, invade circulating erythrocytes. Although invasion is rapid, it is the only time of the life cycle when the parasite is directly exposed to the host immune system. Significant effort has, therefore, focused on identifying the proteins involved and understanding the underlying mechanisms behind merozoite invasion into the protected niche inside the human erythrocyte.
Publisher: Elsevier BV
Date: 03-2012
DOI: 10.1016/J.VACCINE.2012.01.010
Abstract: One approach to develop a malaria blood-stage vaccine is to target proteins that play critical roles in the erythrocyte invasion of merozoites. The merozoite surface proteins (MSPs) and the erythrocyte-binding antigens (EBAs) are considered promising vaccine candidates, for they are known to play important roles in erythrocyte invasion and are exposed to host immune system. Here we focused on a Plasmodium falciparum antigen, PfMSPDBL1 (encoded by PF10_0348 gene) that is a member of the MSP3 family and has both Duffy binding-like (DBL) domain and secreted polymorphic antigen associated with merozoites (SPAM) domain. Therefore, we aimed to characterize PfMSPDBL1 as a vaccine candidate. Recombinant full-length protein (rFL) of PfMSPDBL1 was synthesized by a wheat germ cell-free system, and rabbit antiserum was raised against rFL. We show that rabbit anti-PfMSPDBL1 antibodies inhibited erythrocyte invasion of wild type parasites in vitro in a dose dependent manner, and the specificity of inhibitory activity was confirmed using PfMSPDBL1 knockout parasites. Pre-incubation of the anti-PfMSPDBL1 antibodies with the recombinant SPAM domain had no effect on the inhibitory activity suggesting that antibodies to this region were not involved. In addition, antibodies to rFL were elicited by P. falciparum infection in malaria endemic area, suggesting the PfMSLDBL1 is immunogenic to humans. Our results suggest that PfMSPDBL1 is a novel blood-stage malaria vaccine candidate.
Publisher: Public Library of Science (PLoS)
Date: 07-2014
Publisher: Elsevier BV
Date: 15-03-2000
DOI: 10.1016/S0166-6851(00)00176-6
Abstract: Approximately 50 highly erse var genes distributed throughout the haploid genome of the malaria parasite Plasmodium falciparum code for PfEMP1 variants located on the surface of infected erythrocytes. PfEMP1 is involved in cytoadherence of parasitised red blood cells and undergoes antigenic variation through differential expression of var genes. Members of the var gene family are located in chromosome-internal positions on chromosomes 4, 7, 8 and 12, and in subtelomeric regions of all chromosomes. Here we show that there are two distinct and conserved types of 5' upstream regions (var17-type and 5B1-type) of var genes, and suggest that most subtelomeric var genes are flanked by a var17-type 5' upstream sequence. In contrast, 5B1-type 5' upstream are localised to chromosomes that have been shown to contain var genes within chromosome-internal regions. Transcriptional analysis using RT-PCR revealed that var genes flanked by either type of 5' upstream sequence are transcribed in in vitro cultured trophozoite stage parasites. In addition, we have shown that the 5' flanking sequences of four different var genes are able to drive transient expression of the cat reporter gene. Our results suggest that at least the minimal regulatory sequences required for transcription of var genes are conserved among both subgroups of the var gene family. Furthermore, these sequences provide new markers for the investigation of the chromosomal organisation of var genes.
Publisher: American Society of Hematology
Date: 27-11-2014
DOI: 10.1182/BLOOD-2014-06-583054
Abstract: Maurer's clefts are P falciparum–derived membranous structures within the host erythrocyte that are essential for parasite survival. PfPTP1 functions in a large complex of proteins and is required for linking of Maurer's clefts to the host actin cytoskeleton.
Publisher: Wiley
Date: 03-2002
DOI: 10.1046/J.1365-2958.2002.02834.X
Abstract: Merozoite surface protein 3 (MSP3), an important vaccine candidate, is a soluble polymorphic antigen associated with the surface of Plasmodium falciparum merozoites. The MSP3 sequence contains three blocks of heptad repeats that are consistent with the formation of an intramolecular coiled-coil. MSP3 also contains a glutamic acid-rich region and a putative leucine zipper sequence at the C-terminus. We have disrupted the msp3 gene by homologous recombination, resulting in the expression of a truncated form of MSP3 that lacks the putative leucine zipper sequence but retains the glutamic acid-rich region and the heptad repeats. Here, we show that truncated MSP3, lacking the putative leucine zipper region, does not localize to the parasitophorous vacuole or interact with the merozoite surface. Furthermore, the acidic-basic repeat antigen (ABRA), which is present on the merozoite surface, also was not localized to the merozoite surface in parasites expressing the truncated form of MSP3. The P. falciparum merozoites lacking MSP3 and ABRA on the surface show reduced invasion into erythrocytes. These results suggest that MSP3 is not absolutely essential for blood stage growth and that the putative leucine zipper region is required for the trafficking of both MSP3 and ABRA to the parasitophorous vacuole.
Publisher: The American Association of Immunologists
Date: 15-11-2010
Abstract: Abs targeting blood-stage Ags of Plasmodium falciparum are important in acquired immunity to malaria, but major targets remain unclear. The P. falciparum reticulocyte-binding homologs (PfRh) are key ligands used by merozoites during invasion of erythrocytes. PfRh2a and PfRh2b are functionally important members of this family and may be targets of protective immunity, but their potential role in human immunity has not been examined. We expressed eight recombinant proteins covering the entire PfRh2 common region, as well as PfRh2a- and PfRh2b-specific regions. Abs were measured among a cohort of 206 Papua New Guinean children who were followed prospectively for 6 mo for reinfection and malaria. At baseline, Abs were associated with increasing age and active infection. High levels of IgG to all PfRh2 protein constructs were strongly associated with protection from symptomatic malaria and high-density parasitemia. The predominant IgG subclasses were IgG1 and IgG3, with little IgG2 and IgG4 detected. To further understand the significance of PfRh2 as an immune target, we analyzed PfRh2 sequences and found that polymorphisms are concentrated in an N-terminal region of the protein and seem to be under ersifying selection, suggesting immune pressure. Cluster analysis arranged the sequences into two main groups, suggesting that many of the haplotypes identified may be antigenically similar. These findings provide evidence suggesting that PfRh2 is an important target of protective immunity in humans and that Abs act by controlling blood-stage parasitemia and support its potential for vaccine development.
Publisher: American Association for the Advancement of Science (AAAS)
Date: 14-05-2010
Abstract: The global research community must take up the challenge to work toward the eradication of malaria. In the past, malaria research has focused on drugs and vaccines that target the blood stage of infection, and mainly on the most deadly species, Plasmodium falciparum , all of which is justified by the need to prevent and treat the disease. This work remains critically important today. However, an increased research focus is now being placed on potential interventions that aim to kill the parasite stages transmitted to and by the mosquito vector because they may represent more vulnerable targets to stop the spread of malaria. Here, we highlight some of the research into malaria parasite biology that has the potential to provide new intervention targets for antimalarial drugs and vaccines.
Publisher: Wiley
Date: 15-06-2005
DOI: 10.1111/J.1365-2958.2005.04699.X
Abstract: Plasmodium parasites are unicellular eukaryotes that undergo a series of remarkable morphological transformations during the course of a multistage life cycle spanning two hosts (mosquito and human). Relatively little is known about the dynamics of cellular organelles throughout the course of these transformations. Here we describe the morphology of three organelles (endoplasmic reticulum, apicoplast and mitochondrion) through the human blood stages of the parasite life cycle using fluorescent reporter proteins fused to organelle targeting sequences. The endoplasmic reticulum begins as a simple crescent-shaped organelle that develops into a perinuclear ring with two small protrusions, followed by transformation into an extensive reticulated network as the parasite enlarges. Similarly, the apicoplast and the mitochondrion grow from single, small, discrete organelles into highly branched structures in later-stage parasites. These branched structures undergo an ordered fission - apicoplast followed by mitochondrion - to create multiple daughter organelles that are apparently linked as pairs for packaging into daughter cells. This is the first in-depth examination of intracellular organelles in live parasites during the asexual life cycle of this important human pathogen.
Publisher: Wiley
Date: 18-08-2022
Abstract: Plasmepsin X (PMX) is an aspartyl protease that processes proteins essential for Plasmodium parasites to invade and egress from host erythrocytes during the symptomatic asexual stage of malaria. PMX substrates possess a conserved cleavage region denoted by the consensus motif, SF h E ( h =hydrophobic amino acid). Peptidomimetics reflecting the P 3 ‐P 1 positions of the consensus motif were designed and showed potent and selective inhibition of PMX. It was established that PMX prefers Phe in the P 1 position, di‐substitution at the β‐carbon of the P 2 moiety and a hydrophobic P 3 group which was supported by modelling of the peptidomimetics in complex with PMX. The peptidomimetics were shown to arrest asexual P. falciparum parasites at the schizont stage by impairing PMX substrate processing. Overall, the peptidomimetics described will assist in further understanding PMX substrate specificity and have the potential to act as a template for future antimalarial design.
Publisher: Public Library of Science (PLoS)
Date: 03-06-2010
Publisher: Oxford University Press (OUP)
Date: 14-01-2008
Abstract: Alveolates are a recently recognized group of unicellular eukaryotes that unites disparate protists including apicomplexan parasites (which cause malaria and toxoplasmosis), dinoflagellate algae (which cause red tides and are symbionts in many corals), and ciliates (which are microscopic predators and common rumen symbionts). Gene sequence trees provide robust support for the alveolate alliance, but beyond the common presence of membranous sacs (alveoli) subtending the plasma membrane, the group has no unifying morphological feature. We describe a family of proteins, alveolins, associated with these membranous sacs in apicomplexa, dinoflagellates, and ciliates. Alveolins contain numerous simple peptide repeats and are encoded by multigene families. We generated antibodies against a peptide motif common to all alveolins and identified a range of apparently abundant proteins in apicomplexa, dinoflagellates, and ciliates. Immunolocalization reveals that alveolins are associated exclusively with the cortical regions of apicomplexa, dinoflagellates, and ciliates where the alveolar sacs occur. Alveolins are the first molecular nexus between the unifying structures that defines this eukaryotic group. They provide an excellent opportunity to explore the exceptional compartment that was apparently the key to a remarkable ersification of unique protists that occupy a wide array of lifestyle niches.
Publisher: Elsevier BV
Date: 2012
Abstract: The global agenda for malaria eradication would benefit from development of a highly efficacious vaccine that protects against disease and interrupts transmission of Plasmodium falciparum. It is likely that such a vaccine will be multi-component, with antigens from different stages of the parasite life cycle. In this review, inclusion of blood stage antigens in such a vaccine is discussed. Erythrocyte binding-like (EBL) and P. falciparum reticulocyte binding-like (PfRh) proteins are reviewed with respect to their function in erythrocyte invasion, their role in eliciting antibodies contributing to protective immunity and reduction of invasion, leading subsequently to inhibition of parasite multiplication.
Publisher: Elsevier BV
Date: 04-2005
DOI: 10.1016/J.CELL.2005.01.036
Abstract: The malaria parasite Plasmodium falciparum undergoes antigenic variation to evade host immune responses through switching expression of variant surface proteins encoded by the var gene family. We demonstrate that both a subtelomeric transgene and var genes are subject to reversible gene silencing. Var gene silencing involves the SIR complex as gene disruption of PfSIR2 results in activation of this gene family. We also demonstrate that perinuclear gene activation involves chromatin alterations and repositioning into a location that may be permissive for transcription. Together, this implies that locus repositioning and heterochromatic silencing play important roles in the epigenetic regulation of virulence genes in P. falciparum.
Publisher: Elsevier BV
Date: 10-2015
Publisher: Elsevier BV
Date: 12-2019
DOI: 10.1016/J.CELREP.2019.11.073
Abstract: Plasmodium falciparum gametocytes infect mosquitoes and are responsible for malaria transmission. New interventions that block transmission could accelerate malaria elimination. Gametocytes develop within erythrocytes and activate protein export pathways that remodel the host cell. Plasmepsin V (PMV) is an aspartyl protease that is required for protein export in asexual parasites, but its function and essentiality in gametocytes has not been definitively proven, nor has PMV been assessed as a transmission-blocking drug target. Here, we show that PMV is expressed and can be inhibited specifically in P. falciparum stage I-II gametocytes. PMV inhibitors block processing and export of gametocyte effector proteins and inhibit development of stage II-V gametocytes. Gametocytogenesis in the presence of sublethal inhibitor concentrations results in stage V gametocytes that fail to infect mosquitoes. Therefore, PMV primes gametocyte effectors for export, which is essential for the development and fitness of gametocytes for transmission to mosquitoes.
Publisher: Springer Science and Business Media LLC
Date: 27-06-2018
DOI: 10.1038/S41586-018-0249-1
Abstract: Plasmodium vivax is the most widely distributed malaria parasite that infects humans
Publisher: Wiley
Date: 25-04-2022
DOI: 10.1111/MMI.14904
Abstract: Infection with Plasmodium falciparum parasites results in approximately 627,000 deaths from malaria annually. Key to the parasite’s success is their ability to invade and subsequently grow within human erythrocytes. Parasite proteins involved in parasite invasion and proliferation are therefore intrinsically of great interest, as targeting these proteins could provide novel means of therapeutic intervention. One such protein is P113 which has been reported to be both an invasion protein and an intracellular protein located within the parasitophorous vacuole (PV). The PV is delimited by a membrane (PVM) across which a plethora of parasite‐specific proteins are exported via the Plasmodium Translocon of Exported proteins (PTEX) into the erythrocyte to enact various immune evasion functions. To better understand the role of P113 we isolated its binding partners from in vitro cultures of P. falciparum . We detected interactions with the protein export machinery (PTEX and exported protein‐interacting complex) and a variety of proteins that either transit through the PV or reside on the parasite plasma membrane. Genetic knockdown or partial deletion of P113 did not significantly reduce parasite growth or protein export but did disrupt the morphology of the PVM, suggesting that P113 may play a role in maintaining normal PVM architecture.
Publisher: American Society for Microbiology
Date: 07-2007
DOI: 10.1128/IAI.00095-07
Abstract: Red blood cells infected with Plasmodium falciparum (iRBCs) have been shown to modulate maturation of human monocyte-derived dendritic cells (DCs), interfering with their ability to activate T cells. Interaction between Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) and CD36 expressed by DCs is the proposed mechanism, but we show here that DC modulation does not require CD36 binding, PfEMP1, or contact between DCs and infected RBCs and depends on the iRBC dose. iRBCs expressing a PfEMP1 variant that binds chondroitin sulfate A (CSA) but not CD36 were phagocytosed, inhibited lipopolysaccharide (LPS)-induced phenotypic maturation and cytokine secretion, and abrogated the ability of DCs to stimulate allogeneic T-cell proliferation. CD36- and CSA-binding iRBCs showed comparable inhibition. P. falciparum lines rendered deficient in PfEMP1 expression by targeted gene knockout or knockdown also inhibited LPS-induced phenotypic maturation, and separation of DCs and iRBCs in transwells showed that inhibition was not contact dependent. Inhibition was observed at an iRBC:DC ratio of 100:1 but not at a ratio of 10:1. High doses of iRBCs were associated with apoptosis of DCs, which was not activation induced. Lower doses of iRBCs stimulated DC maturation sufficient to activate autologous T-cell proliferation. In conclusion, modulation of DC maturation by P. falciparum is dose dependent and does not require interaction between PfEMP1 and CD36. Inhibition and apoptosis of DCs by high-dose iRBCs may or may not be physiological. However, our observation that low-dose iRBCs initiate functional DC maturation warrants reevaluation and further investigation of DC interactions with blood-stage P. falciparum .
Publisher: Springer Science and Business Media LLC
Date: 13-03-2017
Publisher: Elsevier BV
Date: 11-2006
Publisher: Rockefeller University Press
Date: 21-11-2005
Abstract: Intracellular parasites from the genus Plasmodium reside and multiply in a variety of cells during their development. After invasion of human erythrocytes, asexual stages from the most virulent malaria parasite, P. falciparum, drastically change their host cell and export remodelling and virulence proteins. Recent data demonstrate that a specific NH2-terminal signal conserved across the genus Plasmodium plays a central role in this export process.
Publisher: Hindawi Limited
Date: 15-05-2012
DOI: 10.1111/J.1462-5822.2012.01803.X
Abstract: Centromeres are essential for the faithful transmission of chromosomes to the next generation, therefore being essential in all eukaryotic organisms. The centromeres of Plasmodium falciparum, the causative agent of the most severe form of malaria, have been broadly mapped on most chromosomes, but their epigenetic composition remained undefined. Here, we reveal that the centromeric histone variant PfCENH3 occupies a 4-4.5 kb region on each P. falciparum chromosome, which is devoid of pericentric heterochromatin but harbours another histone variant, PfH2A.Z. These CENH3 covered regions pinpoint the exact position of the centromere on all chromosomes and revealed that all centromeric regions have similar size and sequence composition. Immunofluorescence assay of PfCENH3 strongly suggests that P. falciparum centromeres cluster to a single nuclear location prior to and during mitosis and cytokinesis but dissociate soon after invasion. In summary, we reveal a dynamic association of Plasmodium centromeres, which bear a unique epigenetic signature and conform to a strict structure. These findings suggest that DNA-associated and epigenetic elements play an important role in centromere establishment in this important human pathogen.
Publisher: Research Square Platform LLC
Date: 03-2023
DOI: 10.21203/RS.3.RS-2632356/V1
Abstract: Plasmodium falciparum assembles a protein translocon (PTEX) at the parasitophorous vacuole membrane (PVM) of infected erythrocytes, through which several hundred proteins are exported. The preceding Plasmodium liver stage develops in hepatocytes within a PVM however, the importance of PTEX and identification of exported proteins in P. falciparum liver stages remains unexplored. Here, we apply the FlpL/ FRT system to P. falciparum NF54 to conditionally excise genes in sporozoites, enabling studies at the liver stage. Conditional disruption of PTEX components PTEX150 and EXP2 in sporozoites does not affect their development or infectivity but attenuates liver stage growth. While PTEX150-deficiency significantly reduces liver load in humanized mice, EXP2-deficiency conferred a severe fitness cost, demonstrating that PTEX is essential for P. falciparum liver stage development. We show that liver specific protein 2 (LISP2) and circumsporozoite protein (CSP) contain putative PEXEL sequences cleaved by plasmepsin V, yet they localize to the PVM of infected hepatocytes. The abundance of LISP2 is reduced in PTEX-deficient liver stages, suggesting this protein is degraded in the absence of a functioning PTEX complex. This study employs the FlpL/ FRT system for functional analysis of P. falciparum pre-erythrocytic biology, revealing that the protein export translocon required for growth in erythrocytes is essential for P. falciparum development in hepatocytes and normal LISP2 expression. It also describes two P. falciparum proteins that contain putative PEXEL motifs that are targeted to the PVM.
Publisher: Public Library of Science (PLoS)
Date: 13-06-2012
Publisher: Elsevier BV
Date: 08-2005
Publisher: American Society for Microbiology
Date: 10-2002
DOI: 10.1128/IAI.70.10.5751-5758.2002
Abstract: Apical membrane antigen 1 of Plasmodium falciparum (PfAMA1) contains an N-terminal propeptide that is removed prior to the translocation of the mature protein onto the merozoite surface. We localized unprocessed PfAMA1 to the microneme organelles of the intraerythrocytic schizont. The results have suggested that the processed form of PfAMA1 translocates from the microneme compartment independently of another microneme protein, EBA175, which is also involved in the invasion of human erythrocytes.
Publisher: Elsevier BV
Date: 08-2010
Publisher: Springer Science and Business Media LLC
Date: 28-12-2005
DOI: 10.1038/NATURE04407
Abstract: Mono-allelic expression of gene families is used by many organisms to mediate phenotypic variation of surface proteins. In the apicomplexan parasite Plasmodium falciparum, responsible for the severe form of malaria in humans, this is exemplified by antigenic variation of the highly polymorphic P. falciparum erythrocyte membrane protein 1 (PfEMP1). PfEMP1, encoded by the 60-member var gene family, represents a major virulence factor due to its central role in immune evasion and intravascular parasite sequestration. Mutually exclusive expression of PfEMP1 is controlled by epigenetic mechanisms involving chromatin modification and perinuclear var locus repositioning. Here we show that a var promoter mediates the nucleation and spreading of stably inherited silenced chromatin. Transcriptional activation of this promoter occurs at the nuclear periphery in association with chromosome-end clusters. Additionally, the var promoter sequence is sufficient to infiltrate a transgene into the allelic exclusion programme of var gene expression, as transcriptional activation of this transgene results in silencing of endogenous var gene transcription. These results show that a var promoter is sufficient for epigenetic silencing and mono-allelic transcription of this virulence gene family, and are fundamental for our understanding of antigenic variation in P. falciparum. Furthermore, the PfEMP1 knockdown parasites obtained in this study will be important tools to increase our understanding of P. falciparum-mediated virulence and immune evasion.
Publisher: Elsevier BV
Date: 07-2003
DOI: 10.1016/S0166-6851(03)00118-X
Abstract: Plasmodium falciparum merozoite surface protein (MSP)-2, is a polymorphic protein whose variable regions define two allelic families, the 3D7/IC-1 and FC27/D10 families. The gene encoding MSP-2 is located on chromosome 2 immediately 3' of the gene encoding merozoite surface protein-5 (MSP-5) with a 1096 bp intergenic region that presumably contains the MSP-2 promoter. Here we present characterization of the MSP-2 promoter using transient and stable transfection of P. falciparum. The mRNA transcription initiation site was mapped to a position 256 bp upstream of the MSP-2 translation start site. The ability of the intergenic region between MSP-5 and MSP-2 to promote the expression of chlor henicol acetyl transferase (CAT) has been tested using a series of nested deletions in transient transfection experiments. The minimal region required for CAT expression has been defined and putative regulatory elements delineated. These nested deletions were used for heterologous expression of an FC27 family MSP-2 allele in the 3D7 allelic background in transfected 3D7 lines. In each case, the transgenic P. falciparum lines generated co-express both 3D7 and FC27 allelic forms of MSP-2 at the merozoite surface. These results have identified the functional promoter for MSP-2.
Publisher: Proceedings of the National Academy of Sciences
Date: 27-04-1999
Abstract: Chondroitin sulfate A (CSA) is an important receptor for the sequestration of Plasmodium falciparum in the placenta, but the parasite ligand involved in adhesion has not previously been identified. Here we report the identification of a var gene transcribed in association with binding to CSA and present evidence that the P. falciparum erythrocyte membrane protein 1 product of the gene is the parasite ligand mediating CSA binding. Description of this gene and the implication of P. falciparum erythrocyte membrane protein 1 as the parasite ligand paves the way to a more detailed understanding of the pathogenesis of placental infection and potential therapeutic strategies targeting the interaction.
Publisher: Elsevier BV
Date: 04-2020
Publisher: The American Association of Immunologists
Date: 15-07-2013
Abstract: Abs that inhibit Plasmodium falciparum invasion of erythrocytes form an important component of human immunity against malaria, but key target Ags are largely unknown. Phenotypic variation by P. falciparum mediates the evasion of inhibitory Abs, contributing to the capacity of P. falciparum to cause repeat and chronic infections. However, Ags involved in mediating immune evasion have not been defined, and studies of the function of human Abs are limited. In this study, we used novel approaches to determine the importance of P. falciparum erythrocyte-binding Ags (EBAs), which are important invasion ligands, as targets of human invasion-inhibitory Abs and define their role in contributing to immune evasion through variation in function. We evaluated the invasion-inhibitory activity of acquired Abs from malaria-exposed children and adults from Kenya, using P. falciparum with disruption of genes encoding EBA140, EBA175, and EBA181, either in idually or combined as EBA140/EBA175 or EBA175/EBA181 double knockouts. Our findings provide important new evidence that variation in the expression and function of the EBAs plays an important role in evasion of acquired Abs and that a substantial amount of phenotypic ersity results from variation in expression of different EBAs that contributes to immune evasion by P. falciparum. All three EBAs were identified as important targets of naturally acquired inhibitory Abs demonstrated by differential inhibition of parental parasites greater than EBA knockout lines. This knowledge will help to advance malaria vaccine development and suggests that multiple invasion ligands need to be targeted to overcome the capacity of P. falciparum for immune evasion.
Publisher: EMBO
Date: 04-2022
Publisher: Oxford University Press (OUP)
Date: 02-02-2015
Abstract: Increasing evidence suggests that antibodies against merozoite surface proteins (MSPs) play an important role in clinical immunity to malaria. Two unusual members of the MSP-3 family, merozoite surface protein duffy binding-like (MSPDBL)1 and MSPDBL2, have been shown to be extrinsically associated to MSP-1 on the parasite surface. In addition to a secreted polymorphic antigen associated with merozoite (SPAM) domain characteristic of MSP-3 family members, they also contain Duffy binding-like (DBL) domain and were found to bind to erythrocytes, suggesting that they play a role in parasite invasion. Antibody responses to these proteins were investigated in a treatment-reinfection study conducted in an endemic area of Papua New Guinea to determine their contribution to naturally acquired immunity. Antibodies to the SPAM domains of MSPDBL1 and MSPDBL2 as well as the DBL domain of MSPDBL1 were found to be associated with protection from Plasmodium falciparum clinical episodes. Moreover, affinity-purified anti-MSPDBL1 and MSPDBL2 were found to inhibit in vitro parasite growth and had strong merozoite opsonizing capacity, suggesting that protection targeting these antigens results from ≥2 distinct effector mechanisms. Together these results indicate that MSPDBL1 and MSPDBL2 are important targets of naturally acquired immunity and might constitute potential vaccine candidates.
Publisher: Elsevier BV
Date: 05-2016
DOI: 10.1016/J.BMC.2016.03.027
Abstract: The use of arginine isosteres is a known strategy to overcome poor membrane permeability commonly associated with peptides or peptidomimetics that possess this highly polar amino acid. Here, we apply this strategy to peptidomimetics that are potent inhibitors of the malarial protease, plasmepsin V, with the aim of enhancing their activity against Plasmodium parasites, and exploring the structure-activity relationship of the P3 arginine within the S3 pocket of plasmepsin V. Of the arginine isosteres trialled in the P3 position, we discovered that canavanine was the ideal and that this peptidomimetic potently inhibits plasmepsin V, efficiently blocks protein export and inhibits parasite growth. Structure studies of the peptidomimetics bound to plasmepsin V provided insight into the structural basis for the enzyme activity observed in vitro and provides further evidence why plasmepsin V is highly sensitive to substrate modification.
Publisher: Elsevier BV
Date: 09-2012
Publisher: Hindawi Limited
Date: 11-07-2205
DOI: 10.1111/J.1462-5822.2010.01477.X
Abstract: Plasmodium falciparum is predicted to transport over 300 proteins to the cytosol of its chosen host cell, the mature human erythrocyte, including 19 members of the Hsp40 family. Here, we have generated transfectant lines expressing GFP- or HA-Strep-tagged versions of these proteins, and used these to investigate both localization and other properties of these Hsp40 co-chaperones. These fusion proteins labelled punctate structures within the infected erythrocyte, initially suggestive of a Maurer's clefts localization. Further experiments demonstrated that these structures were distinct from the Maurer's clefts in protein composition. Transmission electron microscopy verifies a non-cleft localization for HA-Strep-tagged versions of these proteins. We were not able to label these structures with BODIPY-ceramide, suggesting a lower size and/or different lipid composition compared with the Maurer's clefts. Solubility studies revealed that the Hsp40-GFP fusion proteins appear to be tightly associated with membranes, but could be released from the bilayer under conditions affecting membrane cholesterol content or organization, suggesting interaction with a binding partner localized to cholesterol-rich domains. These novel structures are highly mobile in the infected erythrocyte, but based on velocity calculations, can be distinguished from the 'highly mobile vesicles' previously described. Our study identifies a further extra-parasitic structure in the P. falciparum-infected erythrocyte, which we name 'J-dots' (as their defining characteristic so far is the content of J-proteins). We suggest that these J-dots are involved in trafficking of parasite-encoded proteins through the cytosol of the infected erythrocyte.
Publisher: Elsevier BV
Date: 11-1996
DOI: 10.1016/S0006-2952(96)00556-4
Abstract: MDR1 P-glycoprotein in membranes of human tumor cells of the CEM/VBL100 line was selectively labelled using photoreactive analogs of verapamil, N-(p-azido-3-[125I]salicyl)amino-verapamil ([125I]ASA-V) and prazosin, 2-[4-(4-azido-3-[125I]iodobenzoyl)piperazin-1-yl]4 -amino-6,7-dimethoxyyquinazoline ([125I]ASA-P). Mefloquine, a quinolinemethanol antimalarial drug, was shown to inhibit the labelling of P-glycoprotein with an efficiency similar to that for verapamil, a known chemosensitizer. By contrast, chloroquine competed poorly for the binding site on P-glycoprotein. Mefloquine also inhibited the functional activity of P-glycoprotein. It decreased the rates of extrusion of [3H]vinblastine and the fluorescent dyes, fluo-3 acetomethoxy ester and rhodamine 123, from drug-resistant cells and decreased the level of resistance of these cells to vinblastine. The ability of mefloquine to inhibit P-glycoprotein function may be involved in the neurotoxic side-effects occasionally associated with the use of mefloquine as an antimalarial drug.
Publisher: Springer Science and Business Media LLC
Date: 02-2016
DOI: 10.1038/NCOMMS10470
Abstract: Plasmodium falciparum exports proteins into erythrocytes using the Plasmodium export element (PEXEL) motif, which is cleaved in the endoplasmic reticulum (ER) by plasmepsin V (PMV). A recent study reported that phosphatidylinositol-3-phosphate (PI(3)P) concentrated in the ER binds to PEXEL motifs and is required for export independent of PMV, and that PEXEL motifs are functionally interchangeable with RxLR motifs of oomycete effectors. Here we show that the PEXEL does not bind PI(3)P, and that this lipid is not concentrated in the ER. We find that RxLR motifs cannot mediate export in P. falciparum . Parasites expressing a mutated version of KAHRP, with the PEXEL motif repositioned near the signal sequence, prevented PMV cleavage. This mutant possessed the putative PI(3)P-binding residues but is not exported. Reinstatement of PEXEL to its original location restores processing by PMV and export. These results challenge the PI(3)P hypothesis and provide evidence that PEXEL position is conserved for co-translational processing and export.
Publisher: Public Library of Science (PLoS)
Date: 29-09-2011
Publisher: Elsevier BV
Date: 08-2006
DOI: 10.1016/J.MOLBIOPARA.2006.03.006
Abstract: Malaria during pregnancy causes serious disease that is associated with sequestration in the placenta of Plasmodium falciparum infected erythrocytes that adhere to several host receptors, including chondroitin sulfate A (CSA). The principal CSA binding ligand associated with placental sequestration is the P. falciparum erythrocyte membrane protein 1 (PfEMP1), encoded by the var2csa gene. We disrupted the var2csa gene in two allogeneic parasites and ablated CSA binding. However, in one parasite line we were able to re-select for adhesion to bovine trachea CSA associated with transcription of two var genes, var-CS2 and varP. Parasites transcribing parts of var-CS2 and varP were present in the placentae of some infected women but the mutant parasites that transcribed var-CS2 and varP were recognized by sera from men and pregnant women independent of parity. This work raises the possibility that the PfEMP1 molecules encoded by var-CS2 and varP may be minor contributors to placental malaria but also confirms the importance of the immunodominant, conserved var2csa PfEMP1s in pregnancy associated malaria and strengthens the case for var2csa as a pregnancy-specific malaria vaccine.
Publisher: Springer Science and Business Media LLC
Date: 16-07-2019
DOI: 10.1038/S41598-019-46500-5
Abstract: We developed a novel series of antimalarial compounds based on a 4-cyano-3-methylisoquinoline. Our lead compound MB14 achieved modest inhibition of the growth in vitro of the human malaria parasite, Plasmodium falciparum . To identify its biological target we selected for parasites resistant to MB14. Genome sequencing revealed that all resistant parasites bore a single point S374R mutation in the sodium (Na + ) efflux transporter PfATP4. There are many compounds known to inhibit PfATP4 and some are under preclinical development. MB14 was shown to inhibit Na + dependent ATPase activity in parasite membranes, consistent with the compound targeting PfATP4 directly. PfATP4 inhibitors cause swelling and lysis of infected erythrocytes, attributed to the accumulation of Na + inside the intracellular parasites and the resultant parasite swelling. We show here that inhibitor-induced lysis of infected erythrocytes is dependent upon the parasite protein RhopH2, a component of the new permeability pathways that are induced by the parasite in the erythrocyte membrane. These pathways mediate the influx of Na + into the infected erythrocyte and their suppression via RhopH2 knockdown limits the accumulation of Na + within the parasite hence protecting the infected erythrocyte from lysis. This study reveals a role for the parasite-induced new permeability pathways in the mechanism of action of PfATP4 inhibitors.
Publisher: Springer Science and Business Media LLC
Date: 17-11-2022
DOI: 10.1038/S41564-022-01261-2
Abstract: The most severe form of malaria is caused by Plasmodium falciparum . These parasites invade human erythrocytes, and an essential step in this process involves the ligand PfRh5, which forms a complex with cysteine-rich protective antigen (CyRPA) and PfRh5-interacting protein (PfRipr) (RCR complex) and binds basigin on the host cell. We identified a heteromeric disulfide-linked complex consisting of P. falciparum Plasmodium thrombospondin-related apical merozoite protein (PfPTRAMP) and P. falciparum cysteine-rich small secreted protein (PfCSS) and have shown that it binds RCR to form a pentameric complex, PCRCR. Using P. falciparum lines with conditional knockouts, invasion inhibitory nanobodies to both PfPTRAMP and PfCSS, and lattice light-sheet microscopy, we show that they are essential for merozoite invasion. The PCRCR complex functions to anchor the contact between merozoite and erythrocyte membranes brought together by strong parasite deformations. We solved the structure of nanobody–PfCSS complexes to identify an inhibitory epitope. Our results define the function of the PCRCR complex and identify invasion neutralizing epitopes providing a roadmap for structure-guided development of these proteins for a blood stage malaria vaccine.
Publisher: Elsevier BV
Date: 04-1987
DOI: 10.1016/0014-4894(87)90160-3
Abstract: An in vivo limiting dilution technique was used to produce several Babesia bovis cloned lines with which to study the basis of virulence and immunogenicity in this parasite. DNA hybridization using a cloned DNA fragment from the BabR locus demonstrated that the cloned lines were a more restricted genetic population than the parent strain. Biosynthetic labeling and immunoprecipitation studies indicated that the cloned lines differed from each other and from the parentals in the expression of a small number of polypeptides and antigens. Animal trials with three of the lines demonstrated that the parental line contains both virulent and avirulent parasite populations, at least three of which are not tick transmissible, and that while the lines do provide significant protection against heterologous challenge, they may not give as effective protection as the parental line. These experiments demonstrated the existence of subpopulations with distinctive molecular and biological properties, providing evidence that the attenuation process is based on the selection of preexisting parasite subpopulations combined with the ability of these parasites to vary genetically.
Publisher: Springer Science and Business Media LLC
Date: 20-08-2018
DOI: 10.1038/S41564-018-0219-2
Abstract: Plasmodium falciparum exports hundreds of virulence proteins within infected erythrocytes, a process that requires cleavage of a pentameric motif called Plasmodium export element or vacuolar transport signal by the endoplasmic reticulum (ER)-resident protease plasmepsin V. We identified plasmepsin V-binding proteins that form a unique interactome required for the translocation of effector cargo into the parasite ER. These interactions are functionally distinct from the Sec61-signal peptidase complex required for the translocation of proteins destined for the classical secretory pathway. This interactome does not involve the signal peptidase (SPC21) and consists of PfSec61, PfSPC25, plasmepsin V and PfSec62, which is an essential component of the post-translational ER translocon. Together, they form a distinct portal for the recognition and translocation of a large subset of Plasmodium export element effector proteins into the ER, thereby remodelling the infected erythrocyte that is required for parasite survival and pathogenesis.
Publisher: Wiley
Date: 29-12-2008
Publisher: Elsevier BV
Date: 25-08-2006
Publisher: American Society for Clinical Investigation
Date: 04-09-2012
DOI: 10.1172/JCI62182
Publisher: Public Library of Science (PLoS)
Date: 27-02-2015
Publisher: Elsevier BV
Date: 08-1992
Publisher: Elsevier BV
Date: 12-2004
Publisher: Springer Science and Business Media LLC
Date: 15-06-2021
DOI: 10.1038/S41467-021-23626-7
Abstract: Host membrane remodeling is indispensable for viruses, bacteria, and parasites, to subvert the membrane barrier and obtain entry into cells. The malaria parasite Plasmodium spp . induces biophysical and molecular changes to the erythrocyte membrane through the ordered secretion of its apical organelles. To understand this process and address the debate regarding how the parasitophorous vacuole membrane (PVM) is formed, we developed an approach using lattice light-sheet microscopy, which enables the parasite interaction with the host cell membrane to be tracked and characterized during invasion. Our results show that the PVM is predominantly formed from the erythrocyte membrane, which undergoes biophysical changes as it is remodeled across all stages of invasion, from pre-invasion through to PVM sealing. This approach enables a functional interrogation of parasite-derived lipids and proteins in PVM biogenesis and echinocytosis during Plasmodium falciparum invasion and promises to yield mechanistic insights regarding how this is more generally orchestrated by other intracellular pathogens.
Publisher: Public Library of Science (PLoS)
Date: 26-12-2012
Publisher: Elsevier BV
Date: 12-2021
DOI: 10.1016/J.BIOORG.2021.105359
Abstract: Malaria is a devastating disease caused by Plasmodium parasites. Emerging resistance against current antimalarial therapeutics has engendered the need to develop antimalarials with novel structural classes. We recently described the identification and initial optimization of the 2-anilino quinazoline antimalarial class. Here, we refine the physicochemical properties of this antimalarial class with the aim to improve aqueous solubility and metabolism and to reduce adverse promiscuity. We show the physicochemical properties of this class are intricately balanced with asexual parasite activity and human cell cytotoxicity. Structural modifications we have implemented improved LipE, aqueous solubility and in vitro metabolism while preserving fast acting P. falciparum asexual stage activity. The lead compounds demonstrated equipotent activity against P. knowlesi parasites and were not predisposed to resistance mechanisms of clinically used antimalarials. The optimized compounds exhibited modest activity against early-stage gametocytes, but no activity against pre-erythrocytic liver parasites. Confoundingly, the refined physicochemical properties installed in the compounds did not engender improved oral efficacy in a P. berghei mouse model of malaria compared to earlier studies on the 2-anilino quinazoline class. This study provides the framework for further development of this antimalarial class.
Publisher: Elsevier BV
Date: 08-2007
DOI: 10.1016/J.CHOM.2007.06.012
Abstract: Plasmodium falciparum erythrocyte membrane protein-1 (PfEMP-1) is a variable antigen expressed by P. falciparum, the malarial parasite. PfEMP-1, present on the surface of infected host erythrocytes, mediates erythrocyte binding to vascular endothelium, enabling the parasite to avoid splenic clearance. In addition, PfEMP-1 is proposed to regulate host immune responses via interactions with the CD36 receptor on antigen-presenting cells. We investigated the immunoregulatory function of PfEMP-1 by comparing host cell responses to erythrocytes infected with either wild-type parasites or transgenic parasites lacking PfEMP-1. We showed that PfEMP-1 suppresses the production of the cytokine interferon-gamma by human peripheral blood mononuclear cells early after exposure to P. falciparum. Suppression of this rapid proinflammatory response was CD36 independent and specific to interferon-gamma production by gammadelta-T, NK, and alphabeta-T cells. These data demonstrate a parasite strategy for downregulating the proinflammatory interferon-gamma response and further establish transgenic parasites lacking PfEMP-1 as powerful tools for elucidating PfEMP-1 functions.
Publisher: American Society for Microbiology
Date: 05-2006
DOI: 10.1128/EC.5.5.849-860.2006
Abstract: The particular virulence of the human malaria parasite Plasmodium falciparum derives from export of parasite-encoded proteins to the surface of the mature erythrocytes in which it resides. The mechanisms and machinery for the export of proteins to the erythrocyte membrane are largely unknown. In other eukaryotic cells, cholesterol-rich membrane microdomains or “rafts” have been shown to play an important role in the export of proteins to the cell surface. Our data suggest that depletion of cholesterol from the erythrocyte membrane with methyl-β-cyclodextrin significantly inhibits the delivery of the major virulence factor P. falciparum erythrocyte membrane protein 1 (PfEMP1). The trafficking defect appears to lie at the level of transfer of PfEMP1 from parasite-derived membranous structures within the infected erythrocyte cytoplasm, known as the Maurer's clefts, to the erythrocyte membrane. Thus our data suggest that delivery of this key cytoadherence-mediating protein to the host erythrocyte membrane involves insertion of PfEMP1 at cholesterol-rich microdomains. GTP-dependent vesicle budding and fusion events are also involved in many trafficking processes. To determine whether GTP-dependent events are involved in PfEMP1 trafficking, we have incorporated non-membrane-permeating GTP analogs inside resealed erythrocytes. Although these nonhydrolyzable GTP analogs reduced erythrocyte invasion efficiency and partially retarded growth of the intracellular parasite, they appeared to have little direct effect on PfEMP1 trafficking.
Publisher: Elsevier BV
Date: 08-1990
DOI: 10.1016/0166-6851(90)90109-Y
Abstract: Selection of the rodent malaria Plasmodium chabaudi with low levels of the antifolate drug pyrimethamine has previously been shown by us to result in duplication of the dihydrofolate reductase-thymidylate synthase (DHFR-TS) gene by a duplication of chromosome 7 and subsequent rearrangements. We have selected this resultant parasite line with large doses of pyrimethamine and analysed the DHFR-TS gene and chromosomes for any changes. Increased drug pressure has resulted in reappearance of a chromosome with the same structure as chromosome 7 from DS the parent line. Sequencing of the DHFR gene from each of the chromosomes has identified a single point mutation that results in a serine to asparagine change at position 106. This is the equivalent mutation that has been identified as the key residue in the mechanism of resistance to pyrimethamine in Plasmodium falciparum. There is no apparent increase in transcription of the DHFR-TS gene and the large increase in resistance is most likely a result of the mutation in the DHFR gene.
Publisher: Elsevier BV
Date: 10-2021
Publisher: American Society for Microbiology
Date: 08-2016
DOI: 10.1128/IAI.00145-16
Abstract: It is unclear whether naturally acquired immunity to Plasmodium falciparum results from the acquisition of antibodies to multiple, erse antigens or to fewer, highly conserved antigens. Moreover, the specific antibody functions required for malaria immunity are unknown, and hence informative immunological assays are urgently needed to address these knowledge gaps and guide vaccine development. In this study, we investigated whether merozoite-opsonizing antibodies are associated with protection from malaria in a strain-specific or strain-transcending manner by using a novel field isolate and an immune plasma-matched cohort from Papua New Guinea with our validated assay of merozoite phagocytosis. Highly correlated opsonization responses were observed across the 15 parasite strains tested, as were strong associations with protection (composite phagocytosis score across all strains in children uninfected at baseline: hazard ratio of 0.15, 95% confidence interval of 0.04 to 0.63). Opsonizing antibodies had a strong strain-transcending component, and the opsonization of transgenic parasites deficient for MSP3, MSP6, MSPDBL1, or P. falciparum MSP1-19 (PfMSP1-19) was similar to that of wild-type parasites. We have provided the first evidence that merozoite opsonization is predominantly strain transcending, and the highly consistent associations with protection against erse parasite strains strongly supports the use of merozoite opsonization as a correlate of immunity for field studies and vaccine trials. These results demonstrate that conserved domains within merozoite antigens targeted by opsonization generate strain-transcending immune responses and represent promising vaccine candidates.
Publisher: Rockefeller University Press
Date: 18-06-2001
Abstract: Antibodies that bind to antigens expressed on the merozoite form of the malaria parasite can inhibit parasite growth by preventing merozoite invasion of red blood cells. Inhibitory antibodies are found in the sera of malaria-immune in iduals, however, the specificity of those that are important to this process is not known. In this paper, we have used allelic replacement to construct a Plasmodium falciparum parasite line that expresses the complete COOH-terminal fragment of merozoite surface protein (MSP)-119 from the ergent rodent malaria P. chabaudi. By comparing this transfected line with parental parasites that differ only in MSP-119, we show that antibodies specific for this domain are a major component of the inhibitory response in P. falciparum–immune humans and P. chabaudi–immune mice. In some in idual human sera, MSP-119 antibodies dominated the inhibitory activity. The finding that antibodies to a small region of a single protein play a major role in this process has important implications for malaria immunity and is strongly supportive of further understanding and development of MSP-119–based vaccines.
Publisher: Elsevier BV
Date: 07-2008
Publisher: Public Library of Science (PLoS)
Date: 14-04-2009
Publisher: American Society for Microbiology
Date: 11-2011
DOI: 10.1128/IAI.05412-11
Abstract: One of the solutions for reducing the global mortality and morbidity due to malaria is multivalent vaccines comprising antigens of several life cycle stages of the malarial parasite. Hence, there is a need for supplementing the current set of malaria vaccine candidate antigens. Here, we aimed to characterize glycosylphosphatidylinositol (GPI)-anchored micronemal antigen (GAMA) encoded by the PF08_0008 gene in Plasmodium falciparum . Antibodies were raised against recombinant GAMA synthesized by using a wheat germ cell-free system. Immunoelectron microscopy demonstrated for the first time that GAMA is a microneme protein of the merozoite. Erythrocyte binding assays revealed that GAMA possesses an erythrocyte binding epitope in the C-terminal region and it binds a nonsialylated protein receptor on human erythrocytes. Growth inhibition assays revealed that anti-GAMA antibodies can inhibit P. falciparum invasion in a dose-dependent manner and GAMA plays a role in the sialic acid (SA)-independent invasion pathway. Anti-GAMA antibodies in combination with anti-erythrocyte binding antigen 175 exhibited a significantly higher level of invasion inhibition, supporting the rationale that targeting of both SA-dependent and SA-independent ligands athways is better than targeting either of them alone. Human sera collected from areas of malaria endemicity in Mali and Thailand recognized GAMA. Since GAMA in P. falciparum is refractory to gene knockout attempts, it is essential to parasite invasion. Overall, our study indicates that GAMA is a novel blood-stage vaccine candidate antigen.
Publisher: Elsevier BV
Date: 09-1997
DOI: 10.1016/S0166-6851(97)00095-9
Abstract: We have previously shown that a radioiodinated photoreactive analogue of chloroquine, [125I]N-(4-(4-diethylamino-1-methylbutylamino)quinolin-6-yl) -4-azido-2-hydroxybenzamide ([125I]ASA-Q), specifically labels two proteins in Plasmodium falciparum with apparent molecular weights (Mr) of 42 and 33 kDa (Foley M, Deady LW, Ng K, Cowman AF, Tilley L. J Biol Chem 1994:269:6955-6961). We now report the identification of the 33 kDa protein. The 33 kDa protein was purified from Plasmodium falciparum using photoaffinity labeling with [125I]ASA-Q to monitor the enrichment process. N-terminal sequence analysis of the purified protein revealed exact identity of the first 35 amino acids with P. falciparum lactate dehydrogenase (PfLDH). The plasmodial enzyme was cloned and expressed in E. coli and the recombinant protein used to produce a rabbit antiserum. Immunoprecipitation using affinity-purified anti-PfLDH antibodies confirmed the identity of the 33 kDa CQ-binding protein. The enzyme activity of purified PfLDH was not significantly affected by chloroquine indicating that PfLDH is not a direct target of CQ. PfLDH was, however, shown to be exquisitely sensitive to inhibition by free heme and chloroquine protected against this inhibitory effect.
Publisher: Elsevier BV
Date: 04-1984
DOI: 10.1016/0166-6851(84)90057-4
Abstract: Independent isolates of Babesia bovis differ by only a limited number of polypeptides, some of which may be important as host protective antigens. Avirulent derivatives of these parasites also differ from their virulent counterparts in only a few polypeptides. To identify genes encoding such polypeptides we have isolated cDNA clones corresponding to poly(A)+ RNAs that are expressed only in certain isolates. For this purpose a cDNA clone library was constructed from poly(A)+ RNA of the K-avirulent isolate (KA). These clones were screened by colony hybridization using [32P]cDNA complementary to poly(A)+ RNA from KA and from virulent isolates, in order to identify clones that selectively hybridize to one cDNA probe. Hybridization of DNA from three clones, designated pK4, pK5 and pK6 to poly(A)+ RNA from various isolates revealed different and complex patterns. The gene represented by clone pK5 appeared to be transcribed predominantly in avirulent parasites. Analysis of genomic DNA by the Southern procedure enabled each isolate to be distinguished and suggested that most isolates are comprised of a heterogeneous mixture of subpopulations. Analysis of genomic DNA from parasites obtained after passage of KA through the tick vector (Boophilus microplus) suggested that a subpopulation was being selected that more closely resembled KV than KA.
Publisher: eLife Sciences Publications, Ltd
Date: 22-02-2017
DOI: 10.7554/ELIFE.21083
Abstract: The most lethal form of malaria in humans is caused by Plasmodium falciparum. These parasites invade erythrocytes, a complex process involving multiple ligand-receptor interactions. The parasite makes initial contact with the erythrocyte followed by dramatic deformations linked to the function of the Erythrocyte binding antigen family and P. falciparum reticulocyte binding-like families. We show EBA-175 mediates substantial changes in the deformability of erythrocytes by binding to glycophorin A and activating a phosphorylation cascade that includes erythrocyte cytoskeletal proteins resulting in changes in the viscoelastic properties of the host cell. TRPM7 kinase inhibitors FTY720 and waixenicin A block the changes in the deformability of erythrocytes and inhibit merozoite invasion by directly inhibiting the phosphorylation cascade. Therefore, binding of P. falciparum parasites to the erythrocyte directly activate a signaling pathway through a phosphorylation cascade and this alters the viscoelastic properties of the host membrane conditioning it for successful invasion.
Publisher: Elsevier BV
Date: 08-2210
DOI: 10.1016/J.CHOM.2017.07.003
Abstract: Plasmodium species cause malaria by proliferating in human erythrocytes. Invasion of immunologically privileged erythrocytes provides a relatively protective niche as well as access to a rich source of nutrients. Plasmodium spp. target erythrocytes of different ages, but share a common mechanism of invasion. Specific engagement of erythrocyte receptors defines target cell tropism, activating downstream events and resulting in the physical penetration of the erythrocyte, powered by the parasite's actinomyosin-based motor. Here we review the latest in our understanding of the molecular composition of this highly complex and fascinating biological process.
Publisher: American Society for Microbiology
Date: 29-10-2019
Abstract: Malaria continues to present a major global health burden, particularly in low-resource countries. Plasmodium falciparum , the parasite responsible for the most severe form of malaria, causes disease through rapid and repeated rounds of invasion and replication within red blood cells. Invasion into red blood cells is essential for P. falciparum survival, and the molecular events mediating this process have gained much attention as potential therapeutic targets. With no effective vaccine available, and with the emergence of resistance to antimalarials, there is an urgent need for the development of new therapeutics. Our research has used genetic techniques to provide evidence of an essential protein kinase involved in P. falciparum invasion. Our work adds to the current understanding of parasite signaling processes required for invasion, highlighting PKA as a potential drug target to inhibit invasion for the treatment of malaria.
Publisher: American Society for Clinical Investigation
Date: 02-01-2008
DOI: 10.1172/JCI32138
Publisher: American Chemical Society (ACS)
Date: 11-09-2014
DOI: 10.1021/JM500797G
Abstract: Following erythrocyte invasion, malaria parasites export a catalogue of remodeling proteins into the infected cell that enable parasite development in the human host. Export is dependent on the activity of the aspartyl protease, plasmepsin V (PMV), which cleaves proteins within the Plasmodium export element (PEXEL RxL↓xE/Q/D) in the parasite's endoplasmic reticulum. Here, we generated transition state mimetics of the native PEXEL substrate that potently inhibit PMV isolated from Plasmodium falciparum and Plasmodium vivax. Through optimization, we identified that the activity of the mimetics was completely dependent on the presence of P1 Leu and P3 Arg. Treatment of P. falciparum-infected erythrocytes with a set of optimized mimetics impaired PEXEL processing and killed the parasites. The striking effect of the compounds provides a clearer understanding of the accessibility of the PMV active site and reaffirms the enzyme as an attractive target for the design of future antimalarials.
Publisher: American Chemical Society (ACS)
Date: 26-01-2017
DOI: 10.1021/ACS.JMEDCHEM.6B01673
Abstract: Novel antimalarial therapeutics that target multiple stages of the parasite lifecycle are urgently required to tackle the emerging problem of resistance with current drugs. Here, we describe the optimization of the 2-anilino quinazoline class as antimalarial agents. The class, identified from publicly available antimalarial screening data, was optimized to generate lead compounds that possess potent antimalarial activity against P. falciparum parasites comparable to the known antimalarials, chloroquine and mefloquine. During the optimization process, we defined the functionality necessary for activity and improved in vitro metabolism and solubility. The resultant lead compounds possess potent activity against a multidrug resistant strain of P. falciparum and arrest parasites at the ring phase of the asexual stage and also gametocytogensis. Finally, we show that the lead compounds are orally efficacious in a 4 day murine model of malaria disease burden.
Publisher: Elsevier BV
Date: 07-2017
Publisher: Elsevier BV
Date: 2010
DOI: 10.1016/J.IJPARA.2009.09.002
Abstract: The life cycle of the malaria parasite Plasmodium falciparum involves dramatic morphological and molecular changes required for infection of insect and mammalian hosts. Stage-specific gene expression is crucial, yet few nuclear factors, including potential epigenetic regulators, have been identified. Epigenetic mechanisms play an important role in the switched expression of members of species-specific gene families, which encode proteins exported into the cytoplasm and onto the surface of infected erythrocytes. This includes the large virulence-associated var gene family, in which monoallelic transcription of a single member and switching to other var genes leads to a display of different surface ligands with distinct antigenic and adhesive properties. Using a bio-informatic approach we identified 24 putative nuclear proteins. Tagging with sequences encoding GFP or haemagglutinin (HA) epitopes allowed for identification and localisation analysis of 12 nuclear proteins that are potential regulators of P. falciparum gene expression. These proteins specifically localise to distinct areas of the nucleus, reaching from the centre towards the nuclear envelope, giving new insights into the apicomplexan nuclear architecture. Proteins presenting a punctate distribution in the perinuclear sub-compartments are potential virulence gene regulators as silenced and active var genes reside at the nuclear periphery either clustered or in small expression sites, respectively. These analyses demonstrated an ordered compartmentalisation, indicating a complex sub-nuclear organisation that contributes to the complexity of transcriptional regulation in P. falciparum.
Publisher: Springer Science and Business Media LLC
Date: 06-08-2018
Publisher: Elsevier BV
Date: 2014
Publisher: Wiley
Date: 30-08-2006
DOI: 10.1111/J.1365-2958.2006.05364.X
Abstract: The malaria parasite Plasmodium falciparum undergoes antigenic variation through allelic exclusion and variant expression of surface proteins encoded by the var gene family. Regulation of var genes is under epigenetic control and involves reversible silencing and activation that requires the physical repositioning of a var locus into a transcriptionally permissive zone of the nuclear periphery. P. falciparum chromosome ends appear to aggregate into large perinuclear clusters which house both subtelomeric and chromosome central var genes. In this study we further define the composition of telomeric clusters using fluorescent in situ hybridization, and provide evidence that chromosome end clusters are formed by cross-linking protein. In addition, we demonstrate that a subtelomeric reporter gene and a var gene remain within clusters regardless of their transcriptional status. Our findings support a model whereby a highly localized structure dedicated to the activation of a single var gene can be housed within a gene dense chromosome end cluster that is otherwise transcriptionally silent.
Publisher: Wiley
Date: 12-02-2008
DOI: 10.1111/J.1365-2958.2008.06125.X
Abstract: The Golgi apparatus forms the heart of the secretory pathway in eukaryotic cells where proteins are modified, processed and sorted. The transport of proteins from the endoplasmic reticulum (ER) to the cis-side of the Golgi complex takes place at specialized ER sub-domains known as transitional ER (tER). We used the Plasmodium falciparum orthologue of Sec13p to analyse tER organization. We show that the distribution of PfSec13p is restricted to defined areas of the ER membrane. These foci are juxtaposed to the Golgi apparatus and might represent tER sites. To further analyse cis- to trans-Golgi architecture, we generated a double transfectant parasite line that expresses the Golgi marker Golgi reassembly stacking protein (GRASP) as a green fluorescent protein fusion and the trans-Golgi marker Rab6 as a DsRed fusion protein. Our data demonstrate that Golgi multiplication is closely linked to tER multiplication, and that parasite maturation is accompanied by the spatial separation of the cis- and trans- face of this organelle.
Publisher: Informa UK Limited
Date: 11-1989
Abstract: We selected lines of Plasmodium chabaudi that are resistant to high levels of the antifolate drug pyrimethamine and have shown that rearrangement and duplication of a portion of chromosome 7 has occurred in the resistant lines. This chromosomal duplication results in an increase in the chromosome number from 14 to 15: two derived chromosomes (450 kilobases and 1.1 megabases) were smaller than the original chromosome 7 (1.3 megabases), so that essentially only a 200-kilobase region was duplicated. This region contained the DHFR-TS gene and the closely linked Hsp70 gene. We have macrorestriction mapped chromosome 7 from the pyrimethamine-susceptible line (DS) and also the duplicated chromosome 7s in the resistant line. From these maps, we have proposed a process for the karyotype changes. Sequencing of the DHFR gene from the parent and derived chromosomes showed that there were no mutations in the coding sequence. As a result of the duplication of the DHFR-TS gene, there is at least a twofold increase in expression of the DHFR-TS gene, and this may explain the ability of the pyrimethamine-resistant lines to grow in increased amounts of the drug.
Publisher: American Geophysical Union (AGU)
Date: 02-1998
DOI: 10.1021/JS970279Q
Publisher: Elsevier BV
Date: 02-2006
Publisher: Elsevier BV
Date: 04-1997
DOI: 10.1016/S0166-6851(96)02822-8
Abstract: The pfmdr 1 gene encodes a Plasmodium falciparum homologue of the human P-glycoprotein expressed on the surface of the parasite food vacuole. Variation in copy number and specific codon mutations of pfmdr 1 have been implicated in the development of parasite resistance to antimalarial drugs. We describe here the technique of Tandem-Competitive Polymerase Chain Reaction (TC-PCR), which allows accurate measurement of pfmdr 1 copy number in parasite DNA obtained directly from small quantities (100 microliters) of red blood cells. We reliably quantified pfmdr1 in previously well characterised strains of Plasmodium falciparum with differing pfmdr1 gene copy numbers using starting amounts of between 3,000 and 40,000 gene copies. We then used TC-PCR to determine pfmdr1 gene copy number in field specimens of venous blood taken from 10 patients with malaria contracted along the Thai-Burmese border. In this region of high grade parasite resistance to mefloquine greater than 70% of s les had a copy number greater than 1 of pfmdr1 determined with a repeatability coefficient of 0.58.
Publisher: Royal Society of Chemistry (RSC)
Date: 2015
DOI: 10.1039/C4MD00409D
Abstract: An N -methylation strategy has been applied to transition state mimetics that are potent inhibitors of plasmepsin V to improve their physical characteristics and their ability to reduce the viability of Plasmodium parasites in culture.
Publisher: Research Square Platform LLC
Date: 07-10-2021
DOI: 10.21203/RS.3.RS-944781/V2
Abstract: Plasmepsin IX (PMIX) and X (PMX) are aspartyl proteases of Plasmodium spp. that play essential roles in parasite egress, invasion and development. Consequently, they are important drug targets for Plasmodium falciparum and P. vivax. WM4 and WM382 are potent inhibitors of PMIX and PMX that block invasion of liver and blood stages and transmission to mosquitoes. WM4 specifically inhibits PMX whilst WM382 is a dual inhibitor of PMIX and PMX. To understand the function of PMIX and PMX proteases we identified new protein substrates in P. falciparum and together with detailed kinetic analyses and structural analyses identified key molecular interactions in the active site responsible for the specificity of WM4 and WM382 inhibition. The crystal structures of PMX apo enzyme and the protease/drug complexes of PMX/WM382 and PMX/WM4 for P. falciparum and P. vivax have been solved. We show PMIX and PMX have similar substrate selectivity, however, there are distinct differences for both peptide and full-length protein substrates through differences in localised 3-dimensional structures for the enzyme substrate-binding cleft and substrate interface. The differences in affinities of WM4 and WM382 binding for PMIX and PMX map to variations in surface interactions with each protease in the S' region of the active sites. Crystal structures of PMX reveal interactions and mechanistic detail on the selectivity of drug binding which will be important for further development of clinical candidates against these important molecular targets.
Publisher: Elsevier BV
Date: 09-2004
Publisher: American Society of Hematology
Date: 18-08-2011
DOI: 10.1182/BLOOD-2011-03-341305
Abstract: The Plasmodium falciparum adhesin PfRh4 binds to complement receptor type-1 (CR1) on human erythrocytes and mediates a glycophorin-independent invasion pathway. CR1 is a complement regulator and immune-adherence receptor on erythrocytes required for shuttling of C3b/C4b-opsonized particles to liver and spleen for phagocytosis. Using recombinant CR1 constructs, we mapped the recognition site for PfRh4 to complement control protein modules 1 to 3 (CCP1-3) at the membrane-distal amino terminus of CR1. This region of CR1 binds to C4b and C3b and accelerates decay of both classic pathway and alternative pathway C3 and C5 convertases. CCP1-3 competed for PfRh4 binding to erythroid CR1 and inhibited the PfRh4-CR1 invasion pathways across a wide range of P falciparum strains. PfRh4 did not bind significantly to other CR1 constructs, including CCP15-17, which is 85% identical to CCP1-3. PfRh4 binding to CR1 did not affect its C3b/C4b binding capability, and we show evidence for a ternary complex between CCP1-3, C4b, and PfRh4. PfRh4 binding specifically inhibited CR1's convertase decay-accelerating activity, whereas there was no effect on factor H-mediated decay-accelerating activity. These results increase our understanding of the functional implications of CR1 engagement with PfRh4 and highlight the interplay between complement regulation and infection.
Publisher: The American Association of Immunologists
Date: 15-06-2017
Abstract: The complement system is a front-line defense system that opsonizes and lyses invading pathogens. To survive, microbes exposed to serum must evade the complement response. To achieve this, many pathogens recruit soluble human complement regulators to their surfaces and hijack their regulatory function for protection from complement activation. C1 esterase inhibitor (C1-INH) is a soluble regulator of complement activation that negatively regulates the classical and lectin pathways of complement to protect human tissue from aberrant activation. In this article, we show that Plasmodium falciparum merozoites, the invasive form of blood stage malaria parasites, actively recruit C1-INH to their surfaces when exposed to human serum. We identified PfMSP3.1, a member of the merozoite surface protein 3 family of merozoite surface proteins, as the direct interaction partner. When bound to the merozoite surface, C1-INH retains its ability to complex with and inhibit C1s, MASP1, and MASP2, the activating proteases of the complement cascade. P. falciparum merozoites that lack PfMSP3.1 showed a marked reduction in C1-INH recruitment and increased C3b deposition on their surfaces. However, these ΔPfMSP3.1 merozoites exhibit enhanced invasion of RBCs in the presence of active complement. This study characterizes an immune-evasion strategy used by malaria parasites and highlights the complex relationship between merozoites and the complement system.
Publisher: American Association for the Advancement of Science (AAAS)
Date: 28-01-2011
Abstract: A protein that controls the entry of Toxoplasma gondii into a host cell also plays a role in controlling the parasite's replication.
Publisher: American Society for Microbiology
Date: 02-2016
DOI: 10.1128/CVI.00452-15
Abstract: IgG antibodies to Plasmodium falciparum are transferred from the maternal to fetal circulation during pregnancy, wane after birth, and are subsequently acquired in response to natural infection. We examined the dynamics of malaria antibody responses of 84 Kenyan infants from birth to 36 months of age by (i) serology, (ii) variant surface antigen (VSA) assay, (iii) growth inhibitory activity (GIA), and (iv) invasion inhibition assays (IIA) specific for merozoite surface protein 1 (MSP1) and sialic acid-dependent invasion pathway. Maternal antibodies in each of these four categories were detected in cord blood and decreased to their lowest level by approximately 6 months of age. Serologic antibodies to 3 preerythrocytic and 10 blood-stage antigens subsequently increased, reaching peak prevalence by 36 months. In contrast, antibodies measured by VSA, GIA, and IIA remained low even up to 36 months. Infants sensitized to P. falciparum in utero , defined by cord blood lymphocyte recall responses to malaria antigens, acquired antimalarial antibodies at the same rate as those who were not sensitized in utero , indicating that fetal exposure to malaria antigens did not affect subsequent infant antimalarial responses. Infants with detectable serologic antibodies at 12 months of age had an increased risk of P. falciparum infection during the subsequent 24 months. We conclude that serologic measures of antimalarial antibodies in children 36 months of age or younger represent biomarkers of malaria exposure rather than protection and that functional antibodies develop after 36 months of age in this population.
Publisher: American Society of Hematology
Date: 15-05-2005
DOI: 10.1182/BLOOD-2004-12-4666
Abstract: After invading human red blood cells (RBCs) the malaria parasite Plasmodium falciparum remodels the host cell by trafficking proteins to the RBC compartment. The virulence protein P falciparum erythrocyte membrane protein 1 (PfEMP1) is responsible for cytoadherence of infected cells to host endothelial receptors. This protein is exported across the parasite plasma membrane and parasitophorous vacuole membrane and inserted into the RBC membrane. We have used green fluorescent protein chimeras and fluorescence photobleaching experiments to follow PfEMP1 export through the infected RBC. Our data show that a knob-associated histidine-rich protein (KAHRP) N-terminal protein export element appended to the PfEMP1 transmembrane and C-terminal domains was sufficient for efficient trafficking of protein domains to the outside of the P falciparum–infected RBC. The physical state of the exported proteins suggests trafficking as a complex rather than in vesicles and supports the hypothesis that endogenous PfEMP1 is trafficked in a similar manner. This study identifies the sequences required for expression of proteins to the outside of the P falciparum–infected RBC membrane.
Publisher: Elsevier BV
Date: 07-2016
DOI: 10.1016/J.CHOM.2016.06.004
Abstract: Plasmodium falciparum parasites in the merozoite stage invade human erythrocytes and cause malaria. Invasion requires multiple interactions between merozoite ligands and erythrocyte receptors. P. falciparum reticulocyte binding homolog 5 (PfRh5) forms a complex with the PfRh5-interacting protein (PfRipr) and Cysteine-rich protective antigen (CyRPA) and binds erythrocytes via the host receptor basigin. However, the specific role that PfRipr and CyRPA play during invasion is unclear. Using P. falciparum lines conditionally expressing PfRipr and CyRPA, we show that loss of PfRipr or CyRPA function blocks growth due to the inability of merozoites to invade erythrocytes. Super-resolution microscopy revealed that PfRipr, CyRPA, and PfRh5 colocalize at the junction between merozoites and erythrocytes during invasion. PfRipr, CyRPA, and PfRipr/CyRPA/PfRh5-basigin complex is required for triggering the Ca(2+) release and establishing the tight junction. Together, these results establish that the PfRh5/PfRipr/CyRPA complex is essential in the sequential molecular events leading to parasite invasion of human erythrocytes.
Publisher: eLife Sciences Publications, Ltd
Date: 08-10-2014
DOI: 10.7554/ELIFE.04187
Abstract: Plasmodium falciparum causes the most severe form of malaria in humans and is responsible for over 700,000 deaths annually. It is an obligate intracellular parasite and invades erythrocytes where it grows in a relatively protected niche. Invasion of erythrocytes is essential for parasite survival and this involves interplay of multiple protein–protein interactions. One of the most important interactions is binding of parasite invasion ligand families EBLs and PfRhs to host receptors on the surface of erythrocytes. PfRh5 is the only essential invasion ligand within the PfRh family and is an important vaccine candidate. PfRh5 binds the host receptor basigin. In this study, we have determined the crystal structure of PfRh5 using diffraction data to 2.18 Å resolution. PfRh5 exhibits a novel fold, comprising nine mostly anti-parallel α-helices encasing an N-terminal β-hairpin, with the overall shape being an elliptical disk. This is the first three-dimensional structure determined for the PfRh family of proteins.
Publisher: Public Library of Science (PLoS)
Date: 17-02-2011
Publisher: American Association for the Advancement of Science (AAAS)
Date: 10-12-2004
Abstract: To establish infection in the host, malaria parasites export remodeling and virulence proteins into the erythrocyte. These proteins can traverse a series of membranes, including the parasite membrane, the parasitophorous vacuole membrane, and the erythrocyte membrane. We show that a conserved pentameric sequence plays a central role in protein export into the host cell and predict the exported proteome in Plasmodium falciparum . We identified 400 putative erythrocyte-targeted proteins corresponding to ∼8% of all predicted genes, with 225 virulence proteins and a further 160 proteins likely to be involved in remodeling of the host erythrocyte. The conservation of this signal across Plasmodium species has implications for the development of new antimalarials.
Publisher: American Society for Microbiology
Date: 04-2009
DOI: 10.1128/IAI.01331-08
Abstract: Invasion of human erythrocytes by the malaria parasite Plasmodium falciparum utilizes multiple ligand-receptor interactions involving erythrocyte receptors and parasite erythrocyte binding proteins of the Duffy binding-like family. Erythrocyte binding antigen 175 (EBA-175) binds to glycophorin A, the most abundant protein on the human erythrocyte surface and EBA-140 (also known as BAEBL) binds to glycophorin C, while the receptor for EBA-181 (also known as JESEBL) remains unknown. EBA binding is mediated via region II, a highly structured extracellular domain that shows a degree of sequence variability between different laboratory strains/isolates. Here, we determined the influence of region II polymorphisms on host cell receptor binding and overall function during invasion of EBA-140, EBA-175, and EBA-181. Polymorphisms in the binding domains of EBA-140 and EBA-181 have been suggested previously to alter their respective receptor specificities. In our hands, these polymorphisms affected the levels of EBA-140 and EBA-181 binding to receptors but, critically, not the receptor specificities of these proteins. The degree of EBA-140 binding to glycophorin C correlates with the level of function for this ligand-receptor interaction in merozoite invasion. In contrast, EBA-175, which is highly polymorphic in region II, shows no variability in its ability to bind to its receptor, glycophorin A. Combined, these data highlight the importance of sequence variability in EBAs as driven by immune selection but not by receptor specificity.
Publisher: Proceedings of the National Academy of Sciences
Date: 09-12-1997
Abstract: Plasmodium falciparum causes the most severe form of malaria in humans. An important class of drugs in malaria treatment is the sulfone/sulfonamide group, of which sulfadoxine is the most commonly used. The target of sulfadoxine is the enzyme dihydropteroate synthase (DHPS), and sequencing of the DHPS gene has identified amino acid differences that may be involved in the mechanism of resistance to this drug. In this study we have sequenced the DHPS gene in 10 isolates from Thailand and identified a new allele of DHPS that has a previously unidentified amino acid difference. We have expressed eight alleles of P. falciparum PPPK-DHPS in Escherichia coli and purified the functional enzymes to homogeneity. Strikingly, the K i for sulfadoxine varies by almost three orders of magnitude from 0.14 μM for the DHPS allele from sensitive isolates to 112 μM for an enzyme expressed in a highly resistant isolate. Comparison of the K i of different sulfonamides and the sulfone dapsone has suggested that the amino acid differences in DHPS would confer cross-resistance to these compounds. These results show that the amino acid differences in the DHPS enzyme of sulfadoxine-resistant isolates of P. falciparum are central to the mechanism of resistance to sulfones and sulfonamides.
Publisher: Elsevier BV
Date: 04-2005
DOI: 10.1016/J.IJPARA.2005.02.010
Abstract: This study is motivated by two data sets which employ a custom Plasmodium falciparum version of the Affymetrix GeneChip, containing only perfect match (PM) oligonucleotides. A PM-only chip cannot be analysed using the standard Affymetrix-supplied software. We compared the performance of three match-only algorithms on these data: the Match Only Integral Distribution (MOID) algorithm, Robust Multichip Analysis (RMA), and the Model Based Expression Index (MBEI). We validated the differential expression of several genes using quantitative reverse transcriptase-PCR. We also performed a comparison using two publicly available 'benchmarking' data sets: the Latin Square spike-in data set generated by Affymetrix, and the Gene Logic dilution series. Since we know what the true fold changes are in these special data sets, they are helpful for assessment of expression algorithms.
Publisher: Wiley
Date: 2008
Abstract: Key issues relating to glycomics research were discussed after the workshop entitled "Frontiers in Glycomics: Bioinformatics and Biomarkers in Disease" by two focus groups nominated by the organizers. The groups focused on two themes: (i) glycomics as the new frontier for the discovery of biomarkers of disease and (ii) requirements for the development of informatics for glycomics and glycobiology. The mandate of the focus groups was to build consensus on these issues and develop a summary of findings and recommendations for presentation to the NIH and the greater scientific community. A list of scientific priorities was developed, presented, and discussed at the workshops. Additional suggestions were solicited from workshop participants and collected using the workshop mailing list. The results are summarized in this White Paper, authored by the co-chairs of the focus groups.
Publisher: Hindawi Limited
Date: 11-2009
Publisher: Elsevier BV
Date: 03-2008
DOI: 10.1016/J.CHOM.2008.02.006
Abstract: Malaria parasites invade host cells using actin-based motility, a process requiring parasite actin filament nucleation and polymerization. Malaria and other apicomplexan parasites lack Arp2/3 complex, an actin nucleator widely conserved across eukaryotes, but do express formins, another type of actin nucleator. Here, we demonstrate that one of two malaria parasite formins, Plasmodium falciparum formin 1 (PfFormin 1), and its ortholog in the related parasite Toxoplasma gondii, follows the moving tight junction between the invading parasite and the host cell, which is the predicted site of the actomyosin motor that powers motility. Furthermore, in vitro, the PfFormin1 actin-binding formin homology 2 domain is a potent nucleator, stimulating actin polymerization and, like other formins, localizing to the barbed end during filament elongation. These findings support a conserved molecular mechanism underlying apicomplexan parasite motility and, given the essential role that actin plays in cell invasion, highlight formins as important determinants of malaria parasite pathogenicity.
Publisher: Springer Science and Business Media LLC
Date: 04-12-2017
DOI: 10.1038/S41598-017-17084-9
Abstract: We sought to identify a subset of Plasmodium falciparum antibody targets that would inform monitoring efforts needed to eliminate malaria in high transmission settings. IgG antibodies to 28 recombinant Pf antigens were measured in residents of two communities in western Kenya examined in 2003 and 2013, when the respective prevalence of asymptomatic parasitemia among children was 81 and 15 percent by microscopy. Annual seroconversion rates based on a sero-catalytic model that dichotomised antibody values to negative versus positive showed that rates were higher in 2003 than 2013 for 1 pre-erythrocytic and 7 blood-stage antigens. Antibody acquisition models that considered antibody levels as continuous variables showed that age-related antibody levels to Circumsporozoite Protein and 10 merozoite proteins increased at different rates with age in 2003 versus 2013. Both models found that antibodies to 5 proteins of the Merozoite Surface Protein 1 complex were differentially acquired between the cohorts, and that changes in antibody levels to Apical Membrane Antigen 1 suggested a decrease in transmission that occurred ~10 years before 2013. Further studies evaluating antibodies to this subset of Pf antigens as biomarkers of malaria exposure and naturally acquired immunity are warranted in endemic settings where transmission has been reduced but persists.
Publisher: Proceedings of the National Academy of Sciences
Date: 13-10-1998
Abstract: A vestigial, nonphotosynthetic plastid has been identified recently in protozoan parasites of the phylum Apicomplexa. The apicomplexan plastid, or “apicoplast,” is indispensable, but the complete sequence of both the Plasmodium falciparum and Toxoplasma gondii apicoplast genomes has offered no clue as to what essential metabolic function(s) this organelle might perform in parasites. To investigate possible functions of the apicoplast, we sought to identify nuclear-encoded genes whose products are targeted to the apicoplast in Plasmodium and Toxoplasma . We describe here nuclear genes encoding ribosomal proteins S9 and L28 and the fatty acid biosynthetic enzymes acyl carrier protein (ACP), β-ketoacyl-ACP synthase III (FabH), and β-hydroxyacyl-ACP dehydratase (FabZ). These genes show high similarity to plastid homologues, and immunolocalization of S9 and ACP verifies that the proteins accumulate in the plastid. All the putatively apicoplast-targeted proteins bear N-terminal presequences consistent with plastid targeting, and the ACP presequence is shown to be sufficient to target a recombinant green fluorescent protein reporter to the apicoplast in transgenic T. gondii . Localization of ACP, and very probably FabH and FabZ, in the apicoplast implicates fatty acid biosynthesis as a likely function of the apicoplast. Moreover, inhibition of P. falciparum growth by thiolactomycin, an inhibitor of FabH, indicates a vital role for apicoplast fatty acid biosynthesis. Because the fatty acid biosynthesis genes identified here are of a plastid/bacterial type, and distinct from those of the equivalent pathway in animals, fatty acid biosynthesis is potentially an excellent target for therapeutics directed against malaria, toxoplasmosis, and other apicomplexan-mediated diseases.
Publisher: Springer Science and Business Media LLC
Date: 14-04-2020
DOI: 10.1038/S41467-020-15440-4
Abstract: A promising new compound class for treating human malaria is the imidazolopiperazines (IZP) class. IZP compounds KAF156 (Ganaplacide) and GNF179 are effective against Plasmodium symptomatic asexual blood-stage infections, and are able to prevent transmission and block infection in animal models. But despite the identification of resistance mechanisms in P. falciparum , the mode of action of IZPs remains unknown. To investigate, we here combine in vitro evolution and genome analysis in Saccharomyces cerevisiae with molecular, metabolomic, and chemogenomic methods in P. falciparum . Our findings reveal that IZP-resistant S. cerevisiae clones carry mutations in genes involved in Endoplasmic Reticulum (ER)-based lipid homeostasis and autophagy. In Plasmodium , IZPs inhibit protein trafficking, block the establishment of new permeation pathways, and cause ER expansion. Our data highlight a mechanism for blocking parasite development that is distinct from those of standard compounds used to treat malaria, and demonstrate the potential of IZPs for studying ER-dependent protein processing.
Publisher: Springer Science and Business Media LLC
Date: 08-11-2021
DOI: 10.1186/S12916-021-02150-X
Abstract: There is a clear need for novel approaches to malaria vaccine development. We aimed to develop a genetically attenuated blood-stage vaccine and test its safety, infectivity, and immunogenicity in healthy volunteers. Our approach was to target the gene encoding the knob-associated histidine-rich protein (KAHRP), which is responsible for the assembly of knob structures at the infected erythrocyte surface. Knobs are required for correct display of the polymorphic adhesion ligand P. falciparum erythrocyte membrane protein 1 (PfEMP1), a key virulence determinant encoded by a repertoire of var genes. The gene encoding KAHRP was deleted from P. falciparum 3D7 and a master cell bank was produced in accordance with Good Manufacturing Practice. Eight malaria naïve males were intravenously inoculated (day 0) with 1800 (2 subjects), 1.8 × 10 5 (2 subjects), or 3 × 10 6 viable parasites (4 subjects). Parasitemia was measured using qPCR immunogenicity was determined using standard assays. Parasites were rescued into culture for in vitro analyses (genome sequencing, cytoadhesion assays, scanning electron microscopy, var gene expression). None of the subjects who were administered with 1800 or 1.8 × 10 5 parasites developed parasitemia 3/4 subjects administered 3× 10 6 parasites developed significant parasitemia, first detected on days 13, 18, and 22. One of these three subjects developed symptoms of malaria simultaneously with influenza B (day 17 14,022 parasites/mL) one subject developed mild symptoms on day 28 (19,956 parasites/mL) and one subject remained asymptomatic up to day 35 (5046 parasites/mL). Parasitemia rapidly cleared with artemether/lumefantrine. Parasitemia induced a parasite-specific antibody and cell-mediated immune response. Parasites cultured ex vivo exhibited genotypic and phenotypic properties similar to inoculated parasites, although the var gene expression profile changed during growth in vivo. This study represents the first clinical investigation of a genetically attenuated blood-stage human malaria vaccine. A P. falciparum 3D7 kahrp – strain was tested in vivo and found to be immunogenic but can lead to patent parasitemia at high doses. Australian New Zealand Clinical Trials Registry (number: ACTRN12617000824369 date: 06 June 2017).
Publisher: The Company of Biologists
Date: 07-2008
DOI: 10.1242/JCS.021154
Abstract: Plasmodium falciparum, the causative agent of malaria, relies on a complex protein-secretion system for protein targeting into numerous subcellular destinations. Recently, a homologue of the Golgi re-assembly stacking protein (GRASP) was identified and used to characterise the Golgi organisation in this parasite. Here, we report on the presence of a splice variant that leads to the expression of a GRASP isoform. Although the first GRASP protein (GRASP1) relies on a well-conserved myristoylation motif, the variant (GRASP2) displays a different N-terminus, similar to GRASPs found in fungi. Phylogenetic analyses between GRASP proteins of numerous taxa point to an independent evolution of the unusual N-terminus that could reflect unique requirements for Golgi-dependent protein sorting and organelle biogenesis in P. falciparum. Golgi association of GRASP2 depends on the hydrophobic N-terminus that resembles a signal anchor, leading to a unique mode of Golgi targeting and membrane attachment.
Publisher: Springer Science and Business Media LLC
Date: 02-2000
DOI: 10.1038/35002615
Abstract: Throughout the latter half of this century, the development and spread of resistance to most front-line antimalarial compounds used in the prevention and treatment of the most severe form of human malaria has given cause for grave clinical concern. Polymorphisms in pfmdr1, the gene encoding the P-glycoprotein homologue 1 (Pgh1) protein of Plasmodium falciparum, have been linked to chloroquine resistance Pgh1 has also been implicated in resistance to mefloquine and halofantrine. However, conclusive evidence of a direct causal association between pfmdr1 and resistance to these antimalarials has remained elusive, and a single genetic cross has suggested that Pgh1 is not involved in resistance to chloroquine and mefloquine. Here we provide direct proof that mutations in Pgh1 can confer resistance to mefloquine, quinine and halofantrine. The same mutations influence parasite resistance towards chloroquine in a strain-specific manner and the level of sensitivity to the structurally unrelated compound, artemisinin. This has important implications for the development and efficacy of future antimalarial agents.
Publisher: American Society for Microbiology
Date: 03-2008
DOI: 10.1128/IAI.00144-07
Abstract: One of the key processes in the pathobiology of the malaria parasite is the invasion and subsequent modification of the human erythrocyte. In this complex process, an unknown number of parasite proteins are involved, some of which are leading vaccine candidates. The majority of the proteins that play pivotal roles in invasion are either stored in the apical secretory organelles or located on the surface of the merozoite, the invasive stage of the parasite. Using transcriptional and structural features of these known proteins, we performed a genomewide search that identified 49 hypothetical proteins with a high probability of being located on the surface of the merozoite or in the secretory organelles. Of these candidates, we characterized a novel leucine zipper-like protein in Plasmodium falciparum that is conserved in Plasmodium spp. This protein is expressed in late blood stages and localizes to the rhoptries of the parasite. We demonstrate that this Plasmodium sp.-specific protein has a high degree of conservation within field isolates and that it is refractory to gene knockout attempts and thus might play an important role in invasion.
Publisher: American Association for the Advancement of Science (AAAS)
Date: 22-07-2011
Abstract: A study of an important parasite reveals the core structure of the apicomplexan moving junction.
Publisher: Elsevier BV
Date: 12-2002
Publisher: Springer Science and Business Media LLC
Date: 19-04-0018
DOI: 10.1038/S41467-023-37890-2
Abstract: Plasmodium falciparum causes the most severe form of malaria in humans. The protozoan parasite develops within erythrocytes to mature schizonts, that contain more than 16 merozoites, which egress and invade fresh erythrocytes. The aspartic protease plasmepsin X (PMX), processes proteins and proteases essential for merozoite egress from the schizont and invasion of the host erythrocyte, including the leading vaccine candidate PfRh5. PfRh5 is anchored to the merozoite surface through a 5-membered complex (PCRCR), consisting of Plasmodium thrombospondin-related apical merozoite protein, cysteine-rich small secreted protein, Rh5-interacting protein and cysteine-rich protective antigen. Here, we show that PCRCR is processed by PMX in micronemes to remove the N-terminal prodomain of PhRh5 and this activates the function of the complex unmasking a form that can bind basigin on the erythrocyte membrane and mediate merozoite invasion. The ability to activate PCRCR at a specific time in merozoite invasion most likely masks potential deleterious effects of its function until they are required. These results provide an important understanding of the essential role of PMX and the fine regulation of PCRCR function in P. falciparum biology.
Publisher: Public Library of Science (PLoS)
Date: 20-09-2012
Publisher: Oxford University Press (OUP)
Date: 15-10-2010
DOI: 10.1086/656413
Abstract: Antibodies targeting blood stage antigens are important in protection against malaria, but the principle targets remain unclear. Erythrocyte-binding antigens (EBAs) are important erythrocyte invasion ligands used by merozoites and may be targets of protective immunity, but there are limited data examining their potential importance. We examined antibodies among 206 Papua New Guinean children who were treated with antimalarials at enrollment and observed prospectively for 6 months for reinfection and malaria. Immunoglobulin (Ig) G, IgG subclasses, and IgM to different regions of EBA175, EBA140, and EBA181 expressed as recombinant proteins were assessed in comparison with several other merozoite antigens. High levels of IgG to each of the EBAs were strongly associated with protection from symptomatic malaria and high density parasitemia, but not with risk of reinfection per se. The predominant IgG subclasses were either IgG1 or IgG3, depending on the antigen. The predominance of IgG1 versus IgG3 reflected structural features of specific regions of the proteins. IgG3 was most strongly associated with protection, even for those antigens that had an IgG1 predominant response. The EBAs appear important targets of acquired protective immunity. These findings support their further development as vaccine candidates.
Publisher: eLife Sciences Publications, Ltd
Date: 30-08-2017
Publisher: Public Library of Science (PLoS)
Date: 05-12-2012
Publisher: Springer Science and Business Media LLC
Date: 10-2008
DOI: 10.1038/NATURE07306
Publisher: Proceedings of the National Academy of Sciences
Date: 20-09-2010
Abstract: Plasmodium falciparum is responsible for the most severe form of malaria disease in humans, causing more than 1 million deaths each year. As an obligate intracellular parasite, P. falciparum ’s ability to invade erythrocytes is essential for its survival within the human host. P. falciparum invades erythrocytes using multiple host receptor–parasite ligand interactions known as invasion pathways. Here we show that CR1 is the host erythrocyte receptor for PfRh4, a major P. falciparum ligand essential for sialic acid–independent invasion. PfRh4 and CR1 interact directly, with a K d of 2.9 μM. PfRh4 binding is strongly correlated with the CR1 level on the erythrocyte surface. Parasite invasion via sialic acid–independent pathways is reduced in low-CR1 erythrocytes due to limited availability of this receptor on the surface. Furthermore, soluble CR1 can competitively block binding of PfRh4 to the erythrocyte surface and specifically inhibit sialic acid–independent parasite invasion. These results demonstrate that CR1 is an erythrocyte receptor used by the parasite ligand PfRh4 for P. falciparum invasion.
Publisher: American Society for Microbiology
Date: 04-2004
DOI: 10.1128/IAI.72.4.2321-2328.2004
Abstract: Merozoite surface protein 1 (MSP1) is a highly polymorphic Plasmodium falciparum merozoite surface protein implicated in the invasion of human erythrocytes during the asexual cycle. It forms a complex with MSP6 and MSP7 on the merozoite surface, and this complex is released from the parasite around the time of erythrocyte invasion. MSP1 and many other merozoite surface proteins contain dimorphic elements in their protein structures, and here we show that MSP6 is also dimorphic. The sequences of eight MSP6 genes indicate that the alleles of each dimorphic form of MSP6 are highly conserved. The smaller 3D7-type MSP6 alleles are detected in parasites from all malarious regions of the world, whereas K1-type MSP6 alleles have only been detected in parasites from mainland Southeast Asia. Cleavage of MSP6, which produces the p36 fragment in 3D7-type MSP6 and associates with MSP1, also occurs in K1-type MSP6 but at a different site in the protein. Anti-3D7 MSP6 antibodies weakly inhibited erythrocyte invasion by homologous 3D7 merozoites but did not inhibit a parasite line expressing the K1-type MSP6 allele. Antibodies from hyperimmune in iduals affinity purified on an MSP3 peptide cross-reacted with MSP6 therefore, MSP6 may also be a target of antibody-dependent cellular inhibition.
Publisher: Public Library of Science (PLoS)
Date: 16-12-2005
Publisher: Elsevier BV
Date: 09-1992
DOI: 10.1016/0166-6851(92)90113-X
Abstract: We have cloned and sequenced a homologue of the ring-infected erythrocyte surface antigen (RESA) gene from Plasmodium falciparum designated RESA-2. Two reading frames with high homology to exon 1 and exon 2 of RESA at both the nucleotide and amino acid levels were identified in the RESA-2 sequence. However, RESA-2 does not contain either of the blocks of tandem repeats present in RESA. The lack of an RNA transcript in either asexual or sexual stage parasites and the presence of an in-frame stop codon in the second reading frame suggests RESA-2 could be a pseudogene. Its lack of expression in asexual stages demonstrates that it does not complement the RESA deletion in isolate FCR3.
Publisher: Springer Science and Business Media LLC
Date: 2006
Publisher: Public Library of Science (PLoS)
Date: 22-12-2015
Publisher: Elsevier BV
Date: 04-1997
DOI: 10.1016/S0092-8674(00)80207-X
Abstract: Knobs at the surface of erythrocytes infected with Plasmodium falciparum have been proposed to be important in adherence of these cells to the vascular endothelium. This structure contains the knob-associated histidine-rich protein (KAHRP) and the adhesion receptor P. falciparum erythrocyte membrane protein 1. We have disrupted the gene encoding KAHRP and show that it is essential for knob formation. Knob-transfectants adhere to CD36 in static assays when tested under flow conditions that mimic those of postcapillary venules, however, the binding to CD36 was dramatically reduced. These data suggest that knobs on P. falciparum-infected erythrocytes exert an important influence on adherence of parasitized-erythrocytes to microvascular endothelium, an important process in the pathogenesis of P. falciparum infections.
Publisher: Elsevier BV
Date: 09-2003
Publisher: eLife Sciences Publications, Ltd
Date: 2017
Publisher: Elsevier BV
Date: 05-1990
DOI: 10.1016/0014-4894(90)90124-U
Abstract: DNA probes were used to detect variation in subpopulations of virulent and serially passaged Babesia bovis. Two distinct patterns were evident after hybridization to genomic DNA the first was a basic profile typical of virulent B. bovis and the second, a more variable array, was characteristic of B. bovis after various stages of attenuation. Tick transmission of avirulent B. bovis causes reversion to the virulent genomic pattern, suggesting that selective enrichment of a small residual subpopulation caused reversion to a virulent profile of subpopulations. Certain genomic fragments, predominant in either virulent or avirulent parasite forms, are putative "markers" or actual elements responsible for these biological characteristics.
Publisher: American Society for Microbiology
Date: 04-2005
Publisher: Springer Science and Business Media LLC
Date: 02-2010
DOI: 10.1038/NATURE08728
Publisher: Proceedings of the National Academy of Sciences
Date: 25-03-2008
Abstract: The transfer of genes from an endosymbiont to its host typically requires acquisition of targeting signals by the gene product to ensure its return to the endosymbiont for function. Many hundreds of plastid-derived genes must have acquired transit peptides for successful relocation to the nucleus. Here, we explore potential evolutionary origins of plastid transit peptides in the malaria parasite Plasmodium falciparum . We show that exons of the P. falciparum genome could serve as transit peptides after exon shuffling. We further demonstrate that numerous randomized peptides and even whimsical sequences based on English words can also function as transit peptides in vivo . Thus, facile acquisition of transit peptides from existing sequence likely expedited endosymbiont integration through intracellular gene transfer.
Publisher: Portland Press Ltd.
Date: 22-12-2022
DOI: 10.1042/BCJ20220554
Abstract: Transmission blocking interventions can stop malaria parasite transmission from mosquito to human by inhibiting parasite infection in mosquitos. One of the most advanced candidates for a malaria transmission blocking vaccine is Pfs230. Pfs230 is the largest member of the 6-cysteine protein family with 14 consecutive 6-cysteine domains and is expressed on the surface of gametocytes and gametes. Here, we present the crystal structure of the first two 6-cysteine domains of Pfs230. We identified high affinity Pfs230-specific nanobodies that recognized gametocytes and bind to distinct sites on Pfs230, which were isolated from immunized alpacas. Using two non-overlapping Pfs230 nanobodies, we show that these nanobodies significantly blocked P. falciparum transmission and reduced the formation of exflagellation centers. Crystal structures of the transmission blocking nanobodies with the first 6-cysteine domain of Pfs230 confirm that they bind to different epitopes. In addition, these nanobodies bind to Pfs230 in the absence of the prodomain, in contrast with the binding of known Pfs230 transmission blocking antibodies. These results provide additional structural insight into Pfs230 domains and elucidate a mechanism of action of transmission blocking Pfs230 nanobodies.
Publisher: Proceedings of the National Academy of Sciences
Date: 02-04-2003
Abstract: The Plasmodium falciparum erythrocyte-binding antigen 175 (EBA-175) is a ligand for merozoite invasion into human erythrocytes that binds to glycophorin A in a sialic acid-dependent manner. P. falciparum strain W2mef depends on sialic acid for invasion of erythrocytes, whereas 3D7 is sialic acid-independent. We generated parasites that lack expression or express truncated forms of EBA-175 in W2mef and 3D7. Lack of EBA-175 expression in W2mef parasites was associated with a switch to sialic acid-independent invasion. 3D7 parasites lacking expression of EBA-175 showed no alteration in their ability to utilize sialic acid-independent pathways. Strikingly, both W2mef and 3D7 parasites lacking EBA-175 expression invaded chymotrypsin-treated erythrocytes inefficiently compared with the parental lines. This loss of function suggests that the EBA-175/glycophorin A ligand–receptor interaction is the major chymotrypsin-resistant invasion pathway. Parasite lines with truncated EBA-175 had invasion phenotypes equivalent to parasites lacking expression of EBA-175. The EBA-175 ligand is functional in erythrocyte invasion by merozoites that utilize either sialic acid-dependent or -independent invasion pathways. This finding suggests a model where a minimal affinity supplied by multiple ligand–receptor interactions is required for successful invasion and has implications for EBA-175 as a malaria vaccine candidate.
Publisher: Elsevier BV
Date: 02-2003
Publisher: Elsevier BV
Date: 02-2005
DOI: 10.1016/J.MOLBIOPARA.2004.09.012
Abstract: The merozoite surface of the pathogenic malaria parasite Plasmodium falciparum is comprised of proteins that are important for the identification and invasion of human red cells. Merozoite surface protein (MSP)3 is a polymorphic protein associated with the surface of merozoites and is also a vaccine candidate. A distinct feature of the MSP3 sequence is three blocks of alanine-rich heptad repeats that are predicted to form an intramolecular coiled-coil. Three orthologues of MSP3 that also contain alanine-rich heptad repeats have been described in P. vivax and we therefore searched the P. falciparum genome database for MSP3 paralogues. We have identified two genes, H101 and H103 related to MSP3, however like another MSP3 paralogue, MSP6, H101 and H103 do not contain heptad repeats. H101 and H103 are expressed during the asexual cycle and immunofluorescence indicates H103 localises to the merozoite surface as a peripheral membrane protein. Transfected parasite lines that express truncated forms of H101 or H103 were viable and grew at the same rate as the parental parasite line. This result may reflect redundancy in function among members of the MSP3/MSP6 gene family as has been described for other families of paralogue genes in P. falciparum.
Publisher: American Society for Microbiology
Date: 03-2011
DOI: 10.1128/IAI.01021-10
Abstract: Plasmodium falciparum causes the most severe form of malaria in humans and invades erythrocytes using multiple ligand-receptor interactions. Two important protein families involved in erythrocyte binding are the erythrocyte binding-like (EBL) and the reticulocyte binding-like (RBL or P. falciparum Rh [PfRh]) proteins. We constructed P. falciparum lines lacking expression of EBL proteins by creating single and double knockouts of the corresponding genes for eba-175 , eba-181 , and eba-140 and show that the EBL and PfRh proteins function cooperatively, consistent with them playing a similar role in merozoite invasion. We provide evidence that PfRh and EBL proteins functionally interact, as loss of function of EBA-181 ablates the ability of PfRh2a/b protein antibodies to inhibit merozoite invasion. Additionally, loss of function of some ebl genes results in selection for increased transcription of the PfRh family. This provides a rational basis for considering PfRh and EBL proteins for use as a combination vaccine against P. falciparum . We immunized rabbits with combinations of PfRh and EBL proteins to test the ability of antibodies to block merozoite invasion in growth inhibition assays. A combination of EBA-175, PfRh2a/b, and PfRh4 recombinant proteins induced antibodies that potently blocked merozoite invasion. This validates the use of a combination of these ligands as a potential vaccine that would have broad activity against P. falciparum .
Publisher: Elsevier BV
Date: 06-2018
DOI: 10.1016/J.EJMECH.2018.05.022
Abstract: Plasmepsin V is an aspartyl protease that plays a critical role in the export of proteins bearing the Plasmodium export element (PEXEL) motif (RxLxQ/E/D) to the infected host erythrocyte, and thus the survival of the malaria parasite. Previously, development of transition state PEXEL mimetic inhibitors of plasmepsin V have primarily focused on demonstrating the importance of the P
Publisher: American Society of Hematology
Date: 05-10-2006
DOI: 10.1182/BLOOD-2006-08-043364
Abstract: A key feature of Plasmodium falciparum, the parasite causing the most severe form of malaria in humans, is its ability to export parasite molecules onto the surface of the erythrocyte. The major virulence factor and variant surface protein PfEMP1 (P falciparum erythrocyte membrane protein 1) acts as a ligand to adhere to endothelial receptors avoiding splenic clearance. Because the erythrocyte is devoid of protein transport machinery, the parasite provides infrastructure for trafficking across membranes it traverses. In this study, we show that the P falciparum skeleton-binding protein 1 (PfSBP1) is required for transport of PfEMP1 to the P falciparum–infected erythrocyte surface. We present evidence that PfSBP1 functions at the parasitophorous vacuole membrane to load PfEMP1 into Maurer clefts during formation of these structures. Furthermore, the major reactivity of antibodies from malaria-exposed multigravid women is directed toward PfEMP1 because this is abolished in the absence of PfSBP1.
Publisher: Springer Science and Business Media LLC
Date: 07-04-2022
DOI: 10.1038/S42003-022-03290-3
Abstract: RhopH complexes consists of Clag3, RhopH2 and RhopH3 and are essential for growth of Plasmodium falciparum inside infected erythrocytes. Proteins are released from rhoptry organelles during merozoite invasion and trafficked to the surface of infected erythrocytes and enable uptake of nutrients. RhopH3, unlike other RhopH proteins, is required for parasite invasion, suggesting some cellular processes RhopH proteins function as single players rather than a complex. We show the RhopH complex has not formed during merozoite invasion. Clag3 is directly released into the host cell cytoplasm, whilst RhopH2 and RhopH3 are released into the nascent parasitophorous vacuole. Export of RhopH2 and RhopH3 from the parasitophorous vacuole into the infected erythrocyte cytoplasm enables assembly of Clag3/RhopH2/RhopH3 complexes and incorporation into the host cell membrane concomitant with activation of nutrient uptake. This suggests compartmentalisation prevents premature channel assembly before intact complex is assembled at the host cell membrane.
Publisher: Springer Science and Business Media LLC
Date: 09-12-2002
DOI: 10.1038/NM807
Publisher: Public Library of Science (PLoS)
Date: 04-09-2009
Publisher: Springer Science and Business Media LLC
Date: 05-2009
DOI: 10.1038/NRMICRO2110
Abstract: Exported proteins of the malaria parasite Plasmodium falciparum interact with proteins of the erythrocyte membrane and induce substantial changes in the morphology, physiology and function of the host cell. These changes underlie the pathology that is responsible for the deaths of 1-2 million children every year due to malaria infections. The advent of molecular transfection technology, including the ability to generate deletion mutants and to introduce fluorescent reporter proteins that track the locations and dynamics of parasite proteins, has increased our understanding of the processes and machinery for export of proteins in P. falciparum-infected erythrocytes and has provided us with insights into the functions of the parasite protein exportome. We review these developments, focusing on parasite proteins that interact with the erythrocyte membrane skeleton or that promote delivery of the major virulence protein, PfEMP1, to the erythrocyte membrane.
Publisher: Elsevier BV
Date: 12-2020
Publisher: Elsevier BV
Date: 08-2006
DOI: 10.1016/J.MIB.2006.06.015
Abstract: The secretory pathway in the malaria parasite Plasmodium falciparum has many unique aspects in terms of protein destinations and trafficking mechanisms. Recently, several exciting insights into protein trafficking within this intracellular parasite have been unveiled: these include signals that are required for targeting of proteins to the red blood cell and the relict plastid (known as the apicoplast) and the elucidation of the pathways of the haemoglobin proteases targeted to the food vacuole. Protein-targeting to the apical organelles in P. falciparum, however, is still not very well understood, but available research offers a tantalising glimpse of the system.
Publisher: American Society of Hematology
Date: 02-2002
Abstract: Red blood cells (RBCs) parasitized by Plasmodium falciparum are rigid and poorly deformable and show abnormal circulatory behavior. During parasite development, knob-associated histidine-rich protein (KAHRP) and P falciparum erythrocyte membrane protein 3 (PfEMP3) are exported from the parasite and interact with the RBC membrane skeleton. Using micropipette aspiration, the membrane shear elastic modulus of RBCs infected with transgenic parasites (with kahrp or pfemp3 genes deleted) was measured to determine the contribution of these proteins to the increased rigidity of parasitized RBCs (PRBCs). In the absence of either protein, the level of membrane rigidification was significantly less than that caused by the normal parental parasite clone. KAHRP had a significantly greater effect on rigidification than PfEMP3, contributing approximately 51% of the overall increase that occurs in PRBCs compared to 15% for PfEMP3. This study provides the first quantitative information on the contribution of specific parasite proteins to altered mechanical properties of PRBCs.
Publisher: Annual Reviews
Date: 08-09-2013
DOI: 10.1146/ANNUREV-MICRO-092412-155730
Abstract: One of the most fascinating and remarkable features of Plasmodium parasites, which cause malaria, is their choice of erythrocytes as the principal host cells in which to reside during infection of a vertebrate host. Parasites completely renovate the terminally differentiated cells, which lack most of the normal organelles and functions of other cells, such as a nucleus and the machinery to express and transport proteins to subcellular locations. Erythrocyte remodeling begins immediately after invasion by the Plasmodium parasite, by expression and export of many hundreds of proteins that assemble into molecular machinery in the host cell that permit protein trafficking, harvesting of nutrients, and mechanisms to evade host immune responses. In this review, we discuss recent studies on erythrocyte remodeling, including mechanisms of protein export as well as the identity, functions, and subcellular locations of key exported proteins.
Publisher: Springer Science and Business Media LLC
Date: 12-12-2018
DOI: 10.1038/S41586-018-0779-6
Abstract: Plasmodium falciparum causes the severe form of malaria that has high levels of mortality in humans. Blood-stage merozoites of P. falciparum invade erythrocytes, and this requires interactions between multiple ligands from the parasite and receptors in hosts. These interactions include the binding of the Rh5-CyRPA-Ripr complex with the erythrocyte receptor basigin
Publisher: Elsevier BV
Date: 11-2016
Publisher: American Society for Microbiology
Date: 28-02-2018
Abstract: The malaria parasite is a massive burden in several parts of the world. Worryingly, the parasite has become resistant to several of the drugs commonly used to treat the disease, and at this time, there is no commercial vaccine. It is therefore critical to identify new targets for the development of antimalarials. To survive in the human body, the malaria parasite needs to invade red blood cells. For this, it uses a variety of effectors stored in organelles forming a structure called the apical complex. The mechanisms behind how the parasite generates the apical complex are poorly understood. In this study, we present evidence that a transmembrane protein called sortilin potentially acts as an escorter to transport proteins from the Golgi apparatus to the rhoptries, a component of the apical complex. Our study provides new insight into the biogenesis of a critical structure of the malaria parasite.
Publisher: Oxford University Press (OUP)
Date: 09-2001
DOI: 10.1016/S0035-9203(01)90023-9
Abstract: Resistance to antimalarial chemotherapy is a major concern for malaria control in Viet Nam. In this study undertaken in 1998, 65 patients with uncomplicated Plasmodium falciparum malaria were monitored for 28 days after completion of a 5-day treatment course with artemisinin. Overall 36.9% (24/65) of patients had recurrent parasitaemia during the surveillance period. P. falciparum isolates were tested for sensitivity in vitro to chloroquine, mefloquine, quinine, sulfadoxine-pyrimethamine and results were compared to those from a similar study in 1995. Increased parasite sensitivity to sulfadoxine-pyrimethamine, chloroquine and quinine was demonstrated, with significantly lower mean EC50 and EC99 values in 1998 compared to 1995. Parasite sensitivity to mefloquine did not differ significantly in the 2 surveys. Isolates were also tested for sensitivity in vitro to artemisinin in the 1998 survey. The mean EC50 was 0.03 mumol/L and the EC99 was 0.94 mumol/L. Parasite sensitivity to artemisinin will need to be monitored in view of its increasing use in Viet Nam.
Publisher: Wiley
Date: 05-02-2009
Publisher: Wiley
Date: 26-06-2000
DOI: 10.1016/S0014-5793(00)01703-8
Abstract: Plasmodium falciparum causes the most lethal form of malaria in humans and is responsible for over two million deaths per year. The development of a vaccine against this parasite is an urgent priority and potential protein targets include those on the surface of the asexual merozoite stage, the form that invades the host erythrocyte. The development of methods to transfect P. falciparum has enabled the construction of gain-of-function and loss-of-function mutants and provided new strategies to analyse the role of parasite proteins. In this review, we describe the use of this technology to examine the role of merozoite antigens in erythrocyte invasion and to address their potential as vaccine candidates.
Publisher: Elsevier BV
Date: 02-2009
DOI: 10.1016/J.IJPARA.2008.10.006
Abstract: Invasion of erythrocytes is a prerequisite in the life history of the malaria parasite. Members of the reticulocyte-binding homologue family (PfRh) have been implicated in the invasion process and in some cases have been shown to act as adhesins, binding to specific receptors on the erythrocyte surface. We have identified a further, putatively essential, PfRh family member in the most virulent human malaria Plasmodium falciparum, called PfRh5, which binds to an unknown class of glycosylated receptors on the erythrocyte surface. This protein is an atypical PfRh family member, being much smaller than others and lacking a transmembrane and cytosolic region at the C-terminus. This suggests it may be part of a functional protein complex. PfRh5 localises to the rhoptries in merozoites and follows the tight junction during the process of erythrocyte invasion. Furthermore, rabbit immune serum raised against a portion of the ecto-domain, inhibits parasite invasion in vitro. We hypothesise an essential role for the PfRh5 adhesin in erythrocyte selection and commitment to invasion. Given its small size, we believe PfRh5 may prove to be a valuable candidate for inclusion in a multi-component anti-malarial vaccine.
Publisher: Hindawi Limited
Date: 24-04-2019
DOI: 10.1111/CMI.13030
Publisher: Springer Science and Business Media LLC
Date: 03-2002
DOI: 10.1007/S00249-001-0202-2
Abstract: The wide availability of the confocal microscope and the emergence of green fluorescent protein (GFP) transfection technology has led to the increasing use of photobleaching studies to examine aspects of cellular dynamics in living cells. In this review, we examine the theory and practice of performing photobleaching studies using a confocal microscope. We illustrate the application of photobleaching protocols using our own measurements of fluorescently labelled red blood cells and of malaria parasite-infected erythrocytes expressing GFP fusions and examine other ex les from the literature.
Publisher: eLife Sciences Publications, Ltd
Date: 14-02-2017
DOI: 10.7554/ELIFE.21347
Abstract: Plasmodium falciparum causes malaria in humans with over 450,000 deaths annually. The asexual blood stage involves invasion of erythrocytes by merozoites, in which they grow and ide to release daughter merozoites, which in turn invade new erythrocytes perpetuating the cycle responsible for malaria. A key step in merozoite invasion is the essential binding of PfRh5/CyRPA/PfRipr complex to basigin, a step linked to the formation of a pore between merozoites and erythrocytes. We show CyRPA interacts directly with PfRh5. An invasion inhibitory monoclonal antibody to CyRPA blocks binding of CyRPA to PfRh5 and complex formation thus illuminating the molecular mechanism for inhibition of parasite growth. We determined the crystal structures of CyRPA alone and in complex with an antibody Fab fragment. CyRPA has a six-bladed β-propeller fold, and we identify the region that interacts with PfRh5. This functionally conserved epitope is a potential target for vaccines against P. falciparum.
Publisher: Elsevier BV
Date: 08-2007
DOI: 10.1016/J.IJPARA.2007.02.009
Abstract: The malaria parasite, Plasmodium falciparum, exports proteins beyond the confines of its own plasma membrane, however there is debate regarding the machinery used for these trafficking events. We have generated transgenic parasites expressing chimeric proteins and used immunofluorescence studies to determine the locations of plasmodial homologues of the COPII component, Sar1p, and the Golgi-docking protein, Bet3p. The P. falciparum Sar1p (PfSar1p) chimeras bind to the endoplasmic reticulum surface and define a network of membranes wrapped around parasite nuclei. As the parasite matures, the endomembrane systems of in idual merozoites remain interconnected until very late in schizogony. Antibodies raised against plasmodial Bet3p recognise two foci of reactivity in early parasite stages that increase in number as the parasite matures. Some of the P. falciparum Bet3p (PfBet3p) compartments are juxtaposed to compartments defined by the cis Golgi marker, PfGRASP, while others are distributed through the cytoplasm. The compartments defined by the trans Golgi marker, PfRab6, are separate, suggesting that the Golgi is dispersed. Bet3p-green fluorescent protein (GFP) is partly associated with punctate structures but a substantial population diffuses freely in the parasite cytoplasm. By contrast, yeast Bet3p is very tightly associated with immobile structures. This study challenges the view that the COPII complex and the Golgi apparatus are exported into the infected erythrocyte cytoplasm.
Publisher: eLife Sciences Publications, Ltd
Date: 26-09-2017
DOI: 10.7554/ELIFE.28673
Abstract: The study of antigenic targets of naturally-acquired immunity is essential to identify and prioritize antigens for further functional characterization. We measured total IgG antibodies to 38 P. vivax antigens, investigating their relationship with prospective risk of malaria in a cohort of 1–3 years old Papua New Guinean children. Using simulated annealing algorithms, the potential protective efficacy of antibodies to multiple antigen-combinations, and the antibody thresholds associated with protection were investigated for the first time. High antibody levels to multiple known and newly identified proteins were strongly associated with protection (IRR 0.44–0.74, p .001–0.041). Among five-antigen combinations with the strongest protective effect ( %), EBP, DBPII, RBP1a, CyRPA, and PVX_081550 were most frequently identified several of them requiring very low antibody levels to show a protective association. These data identify in idual antigens that should be prioritized for further functional testing and establish a clear path to testing a multicomponent P. vivax vaccine.
Publisher: Elsevier BV
Date: 09-2005
DOI: 10.1016/J.VETPAR.2005.07.002
Abstract: The apicomplexan parasites Babesia and Plasmodium are related, yet phylogenetically distinct haemoprotozoa that infect red blood cells and cause severe diseases of major human and veterinary importance. A variety of cellular and molecular interactions are pivotal in many aspects of the pathogenicity of these two parasites. Comparison of the cellular and molecular mechanisms that culminate in accumulation of parasitised red blood cells in the microvasculature of cattle infected with Babesia bovis (babesiosis) and humans infected with Plasmodium falciparum (falciparum malaria) is particularly instructive given the striking similarities in the pathophysiology of these two important medical and veterinary diseases. While such adhesive phenomena have been studied extensively in malaria, they have received relatively little attention in babesiosis. In this review, we summarise the findings of more than 25 years of research into cellular adhesive phenomena in malaria and speculate on how this body of work can now be applied to Babesia parasites. Such information is fundamental if we are to learn more about the biology of Babesia parasites, the cellular and molecular mechanisms by which they cause infection and disease and how to develop novel therapeutic strategies or vaccines for both Babesia and malaria infections.
Publisher: American Society for Microbiology
Date: 24-12-2019
Abstract: The malaria parasite has a voracious appetite, requiring large amounts of glucose and nutrients for its rapid growth and proliferation inside human red blood cells. The host cell is resource rich, but this is a double-edged sword nutrient excess can lead to undesirable metabolic reactions and harmful by-products. Here, we demonstrate that the parasite possesses a metabolite repair enzyme (PGP) that suppresses harmful metabolic by-products (via substrate dephosphorylation) and allows the parasite to maintain central carbon metabolism. Loss of PGP leads to the accumulation of two damaged metabolites and causes a domino effect of metabolic dysregulation. Accumulation of one damaged metabolite inhibits an essential enzyme in the pentose phosphate pathway, leading to substrate accumulation and secondary inhibition of glycolysis. This work highlights how the parasite coordinates metabolic flux by eliminating harmful metabolic by-products to ensure rapid proliferation in its resource-rich niche.
Publisher: Springer Science and Business Media LLC
Date: 05-1990
DOI: 10.1038/345255A0
Abstract: The lethal form of human malaria caused by Plasmodium falciparum is virtually uncontrollable in many areas because of the development of drug resistance, in particular chloroquine resistance (CQR). CQR is biologically similar to the multiple drug resistance phenotype (MDR) of mammalian tumour cells, as both involve expulsion of drug from the cell and both can be reversed by calcium channel antagonists. A homologue (pfmdr1) of the mammalian multidrug resistance gene has been implicated in CQR because it is lified in some CQR isolates of P. falciparum as is an mdr gene in MDR tumour cells. We show here that the complete sequences of pfmdr1 genes from 2 CQ sensitive (CQS) P. falciparum isolates are identical. In 5 CQR isolates, 1-4 key nucleotide differences resulted in amino acid substitutions. On the basis of these substitutions, we have correctly predicted the CQS/CQR status of a further 34 out of 36 isolates. This is a paradox as CQR arises much less frequently than would be predicted if single point mutations were sufficient. We conclude that a mutated pfmdr1 gene is one of at least two mutated genes required for CQR.
Publisher: Public Library of Science (PLoS)
Date: 27-07-2012
Publisher: Frontiers Media SA
Date: 30-06-2014
Publisher: Wiley
Date: 28-08-2007
Publisher: American Society for Microbiology
Date: 2003
DOI: 10.1128/AAC.47.1.297-301.2003
Abstract: It has long been held that the malaria parasite, Plasmodium sp., is incapable of de novo fatty acid synthesis. This view has recently been overturned with the emergence of data for the presence of a fatty acid biosynthetic pathway in the relict plastid of P. falciparum (known as the apicoplast). This pathway represents the type II pathway common to plant chloroplasts and bacteria but distinct from the type I pathway of animals including humans. Specific inhibitors of the type II pathway, thiolactomycin and triclosan, have been reported to target this Plasmodium pathway. Here we report further inhibitors of the plastid-based pathway that inhibit Plasmodium parasites. These include several analogues of thiolactomycin, two with sixfold-greater efficacy than thiolactomycin. We also report that parasites respond very rapidly to such inhibitors and that the greatest sensitivity is seen in ring-stage parasites. This study substantiates the importance of fatty acid synthesis for blood-stage parasite survival and shows that this pathway provides scope for the development of novel antimalarial drugs.
Publisher: Wiley
Date: 15-10-2001
Publisher: Elsevier BV
Date: 07-1995
DOI: 10.1016/0166-6851(94)00104-U
Abstract: Genomic structure has been determined for a gene encoding a host-protective antigen of the parasitic platyhelminth Taenia ovis. An incomplete cDNA, known as 45W, encodes the protective antigen. Southern hybridisation experiments using 45W cDNA as a probe, revealed that the 45W gene was a member of a multigene family. Differential Southern hybridisation and rapid lification of cDNA end (RACE) experiments were used to characterise the related genes, allowing the full-length coding region of the 45W encoded antigen to be determined. The gene family comprises a minimum of four members per haploid genome with each member showing varying degrees of 5' and 3' homology with respect to the 45W cDNA. A close homologue of the 45W gene, designated 45S, differed from 45W at 11 of 985 nt comprising the full-length mRNA. Sequencing of several independent RACE products for both 45W and 45S identified a cDNA which may be a product of homologous recombination between these genes, suggesting that the two genes may be alleles. Homologous recombination in genes which encode a host protective antigen such as 45W would provide a mechanism by which antigenic variants could arise.
Publisher: American Society for Microbiology
Date: 10-2004
DOI: 10.1128/IAI.72.10.6095-6105.2004
Abstract: Following invasion of human erythrocytes, the malaria parasite, Plasmodium falciparum , exports proteins beyond the confines of its own plasma membrane to modify the properties of the host red cell membrane. These modifications are critical to the pathogenesis of malaria. Analysis of the P. falciparum genome sequence has identified a large number of molecules with putative atypical signal sequences. The signals remain poorly characterized however, a number of molecules with these motifs localize to the host erythrocyte. To examine the role of these atypical signal sequences in the export of parasite proteins, we have generated transfected parasites expressing a chimeric protein comprising the N-terminal region of the P. falciparum ring-infected erythrocyte surface antigen (RESA) appended to green fluorescent protein (GFP). This N-terminal region contains a hydrophobic stretch of amino acids that is presumed to act as a noncanonical secretory signal sequence. Modulation of the timing of transgene expression demonstrates that trafficking of malaria proteins into the host erythrocyte is dependant on both the presence of an appropriate transport signal and the timing of expression. Transgene expression under the control of a trophozoite-specific promoter mistargets the chimeric molecule to the parasitophorous vacuole surrounding the parasite. However, expression of RESA-GFP in schizont stages, under the control of the RESA promoter, enables correct trafficking of a population of the chimeric protein to the host erythrocyte.
Publisher: American Society for Microbiology
Date: 03-2019
DOI: 10.1128/AAC.01804-18
Abstract: A series of 4-amino 2-anilinoquinazolines optimized for activity against the most lethal malaria parasite of humans, Plasmodium falciparum , was evaluated for activity against other human Plasmodium parasites and related apicomplexans that infect humans and animals. Four of the most promising compounds from the 4-amino 2-anilinoquinazoline series were equally as effective against the asexual blood stages of the zoonotic P. knowlesi , suggesting that they could also be effective against the closely related P. vivax , another important human pathogen.
Publisher: The Royal Society
Date: 29-01-2002
Abstract: Malaria is a major human health problem and is responsible for over 2 million deaths per year. It is caused by a number of species of the genus Plasmodium , and Plasmodium falciparum is the causative agent of the most lethal form. Consequently, the development of a vaccine against this parasite is a priority. There are a number of stages of the parasite life cycle that are being targeted for the development of vaccines. Important candidate antigens include proteins on the surface of the asexual merozoite stage, the form that invades the host erythrocyte. The development of methods to manipulate the genome of Plasmodium species has enabled the construction of gain–of–function and loss–of–function mutants and provided new strategies to analyse the role of parasite proteins. This has provided new information on the role of merozoite antigens in erythrocyte invasion and also allows new approaches to address their potential as vaccine candidates.
Publisher: Springer Science and Business Media LLC
Date: 27-07-2015
DOI: 10.1038/NSMB.3061
Abstract: Plasmepsin V, an essential aspartyl protease of malaria parasites, has a key role in the export of effector proteins to parasite-infected erythrocytes. Consequently, it is an important drug target for the two most virulent malaria parasites of humans, Plasmodium falciparum and Plasmodium vivax. We developed a potent inhibitor of plasmepsin V, called WEHI-842, which directly mimics the Plasmodium export element (PEXEL). WEHI-842 inhibits recombinant plasmepsin V with a half-maximal inhibitory concentration of 0.2 nM, efficiently blocks protein export and inhibits parasite growth. We obtained the structure of P. vivax plasmepsin V in complex with WEHI-842 to 2.4-Å resolution, which provides an explanation for the strict requirements for substrate and inhibitor binding. The structure characterizes both a plant-like fold and a malaria-specific helix-turn-helix motif that are likely to be important in cleavage of effector substrates for export.
Publisher: Elsevier BV
Date: 07-2022
DOI: 10.1016/J.STR.2022.03.018
Abstract: Plasmepsins IX (PMIX) and X (PMX) are essential aspartyl proteases for Plasmodium spp. egress, invasion, and development. WM4 and WM382 inhibit PMIX and PMX in Plasmodium falciparum and P. vivax. WM4 inhibits PMX, while WM382 is a dual inhibitor of PMIX and PMX. To understand their function, we identified protein substrates. Enzyme kinetic and structural analyses identified interactions responsible for drug specificity. PMIX and PMX have similar substrate specificity however, there are distinct differences for peptide and protein substrates. Differences in WM4 and WM382 binding for PMIX and PMX map to variations in the S' region and engagement of the active site S3 pocket. Structures of PMX reveal interactions and mechanistic detail of drug binding important for development of clinical candidates against these targets.
Publisher: Springer Science and Business Media LLC
Date: 24-06-2010
Abstract: Artemisinin derivatives have been used for malaria treatment in Vietnam since 1989. Reported malaria cases have decreased from 1,672,000 with 4,650 deaths in 1991, to 91,635 with 43 deaths in 2006. Current national guidelines recommend artemisinin-based combination therapy (ACT), although artesunate is still available as monotherapy through the private sector. Recent reports suggest that effectiveness of ACT and artesunate monotherapy has declined in western Cambodia. This study examined Plasmodium falciparum resistance patterns over 10 years in southwest Vietnam in infected patients treated with artemisinin compounds. The study was conducted in two communes in Phuoc Long district, Binh Phuoc province, 100 km west of the Cambodian border. This was chosen as a likely site for emerging artemisinin resistance because of the high prevalence of P. falciparum malaria, and the length of time that artemisinin had been in use. In vivo and in vitro monitoring of P. falciparum susceptibility to anti-malarial drugs was conducted in 1998, 2001, 2004/5, and 2008/9. Patients with confirmed P. falciparum malaria received therapy with 5 or 7 days of artemisinin (1998 and 2001 respectively) or 7 days of artesunate In the four surveys, 270 patients were recruited and treated. The mean parasite clearance times differed between 1998, 2001 and 2004/5 (1.8, 2.3 and 2.1 days, P 0.01) but not between 1998 and 2008/2009. The mean parasite clearance times were correlated with parasite density at day 0 (r = 0.4 P 0.001). Treatment failure rates after PCR adjustment were 13.8%, 2.9%, 1.2%, and 0% respectively. Susceptibility of P. falciparum to artemisinin in in vitro tests was stable during the period, except for a rise in EC90 and EC99 in 2001. This study showed stable levels of P. falciparum sensitivity to artemisinin compounds in the two sites over a ten-year period. The introduction of ACT in this area in 2003 may have protected against the development of artemisinin resistance. Adherence to the latest WHO and Vietnamese guidelines, which recommend ACT as first-line therapy in all malarious areas, and continued monitoring along the Vietnam-Cambodia border will be essential to prevent the spread of artemisinin resistance in Vietnam.
Publisher: Wiley
Date: 11-2000
Publisher: Elsevier BV
Date: 09-2014
Publisher: Wiley
Date: 03-03-2003
DOI: 10.1093/EMBOJ/CDG096
Publisher: The American Association of Immunologists
Date: 02-2016
Abstract: The human complement system is the frontline defense mechanism against invading pathogens. The coexistence of humans and microbes throughout evolution has produced ingenious molecular mechanisms by which microorganisms escape complement attack. A common evasion strategy used by erse pathogens is the hijacking of soluble human complement regulators to their surfaces to afford protection from complement activation. One such host regulator is factor H (FH), which acts as a negative regulator of complement to protect host tissues from aberrant complement activation. In this report, we show that Plasmodium falciparum merozoites, the invasive form of the malaria parasites, actively recruit FH and its alternative spliced form FH-like protein 1 when exposed to human serum. We have mapped the binding site in FH that recognizes merozoites and identified Pf92, a member of the six-cysteine family of Plasmodium surface proteins, as its direct interaction partner. When bound to merozoites, FH retains cofactor activity, a key function that allows it to downregulate the alternative pathway of complement. In P. falciparum parasites that lack Pf92, we observed changes in the pattern of C3b cleavage that are consistent with decreased regulation of complement activation. These results also show that recruitment of FH affords P. falciparum merozoites protection from complement-mediated lysis. Our study provides new insights on mechanisms of immune evasion of malaria parasites and highlights the important function of surface coat proteins in the interplay between complement regulation and successful infection of the host.
Publisher: American Society for Clinical Investigation
Date: 22-07-2021
Publisher: Wiley
Date: 06-2000
Publisher: Research Square Platform LLC
Date: 24-02-2027
DOI: 10.21203/RS.3.RS-2410384/V1
Abstract: Plasmodium falciparum causes the most severe form of malaria in humans. The protozoan parasite develops within erythrocytes to mature schizonts, that contain more than 16 merozoites, which egress and invade fresh erythrocytes. The aspartic protease plasmepsin X (PMX), processes proteins and proteases essential for merozoite egress from the schizont and invasion of the host erythrocyte, including the leading vaccine candidate PfRh5. PfRh5 is anchored to the merozoite surface through a 5-membered complex (PCRCR), consisting of Plasmodium thrombospondin-related apical merozoite protein (PTRAMP), cysteine-rich small secreted protein (CSS), Rh5-interacting protein (PfRipr) and cysteine-rich protective antigen (CyRPA). We show that PCRCR is processed by PMX in micronemes to remove the N-terminal prodomain of PhRh5 and this activates the function of the complex unmasking a form that can bind basigin on the erythrocyte membrane and mediate merozoite invasion. The ability to activate PCRCR at a specific time in merozoite invasion most likely masks potential deleterious effects of its function until they are required. These results provide an important understanding of the essential role of PMX and the fine regulation of PCRCR function in P. falciparum biology.
Publisher: Public Library of Science (PLoS)
Date: 10-04-2012
Publisher: American Association for the Advancement of Science (AAAS)
Date: 26-08-2005
Abstract: The malaria parasite, Plasmodium falciparum , exploits multiple ligand-receptor interactions, called invasion pathways, to invade the host erythrocyte. Strains of P. falciparum vary in their dependency on sialated red cell receptors for invasion. We show that switching from sialic acid–dependent to –independent invasion is reversible and depends on parasite ligand use. Expression of P. falciparum reticulocyte–binding like homolog 4 (PfRh4) correlates with sialic acid–independent invasion, and PfRh4 is essential for switching invasion pathways. Differential activation of PfRh4 represents a previously unknown mechanism to switch invasion pathways and provides P. falciparum with exquisite adaptability in the face of erythrocyte receptor polymorphisms and host immune responses.
Publisher: American Society for Microbiology
Date: 06-2009
DOI: 10.1128/IAI.00048-09
Abstract: Plasmodium falciparum invasion into human erythrocytes relies on the interaction between multiple parasite ligands and their respective erythrocyte receptors. The sialic acid-independent invasion pathway is dependent on the expression of P. falciparum reticulocyte binding protein-like homologue 4 (PfRh4), as disruption of the gene abolishes the ability of parasites to switch to this pathway. We show that PfRh4 is present as an invasion ligand in culture supernatants as a 160-kDa proteolytic fragment. We confirm that PfRh4 binds to the surfaces of erythrocytes through recognition of an erythrocyte receptor that is neuraminidase resistant but trypsin and chymotrypsin sensitive. Serum antibodies from malaria-exposed in iduals show reactivity against the binding domain of PfRh4. Purified immunoglobulin G raised in rabbits against the binding domain of PfRh4 blocked the binding of native PfRh4 to the surfaces of erythrocytes and inhibited erythrocyte invasion of parasites using sialic acid-independent invasion pathways and grown in neuraminidase-treated erythrocytes. Our results suggest PfRh4 is a potential vaccine candidate.
Publisher: Wiley
Date: 27-02-2013
DOI: 10.1111/TRA.12053
Abstract: Plasmodium falciparum exports several hundred effector proteins that remodel the host erythrocyte and enable parasites to acquire nutrients, sequester in the circulation and evade immune responses. The majority of exported proteins contain the Plasmodium export element (PEXEL RxLxE/Q/D) in their N-terminus, which is proteolytically cleaved in the parasite endoplasmic reticulum by Plasmepsin V, and is necessary for export. Several exported proteins lack a PEXEL or contain noncanonical motifs. Here, we assessed whether Plasmepsin V could process the N-termini of erse protein families in P. falciparum. We show that Plasmepsin V cleaves N-terminal sequences from RIFIN, STEVOR and RESA multigene families, the latter of which contain a relaxed PEXEL (RxLxxE). However, Plasmepsin V does not cleave the N-terminal sequence of the major exported virulence factor erythrocyte membrane protein 1 (PfEMP1) or the PEXEL-negative exported proteins SBP-1 or REX-2. We probed the substrate specificity of Plasmepsin V and determined that lysine at the PEXEL P3 position, which is present in PfEMP1 and other putatively exported proteins, blocks Plasmepsin V activity. Furthermore, isoleucine at position P1 also blocked Plasmepsin V activity. The specificity of Plasmepsin V is therefore exquisitely confined and we have used this novel information to redefine the predicted P. falciparum PEXEL exportome.
Publisher: Elsevier BV
Date: 04-2011
Publisher: Public Library of Science (PLoS)
Date: 15-02-2019
Publisher: Oxford University Press (OUP)
Date: 20-04-2009
DOI: 10.1093/NAR/GKP239
Publisher: Elsevier BV
Date: 09-2009
Publisher: Elsevier BV
Date: 05-2013
DOI: 10.1016/J.CELL.2013.04.029
Abstract: Cell-cell communication is an important mechanism for information exchange promoting cell survival for the control of features such as population density and differentiation. We determined that Plasmodium falciparum-infected red blood cells directly communicate between parasites within a population using exosome-like vesicles that are capable of delivering genes. Importantly, communication via exosome-like vesicles promotes differentiation to sexual forms at a rate that suggests that signaling is involved. Furthermore, we have identified a P. falciparum protein, PfPTP2, that plays a key role in efficient communication. This study reveals a previously unidentified pathway of P. falciparum biology critical for survival in the host and transmission to mosquitoes. This identifies a pathway for the development of agents to block parasite transmission from the human host to the mosquito.
Publisher: American Society for Microbiology
Date: 06-2001
DOI: 10.1128/IAI.69.6.3635-3645.2001
Abstract: The 235-kDa family of rhoptry proteins in Plasmodium yoelii and the two reticulocyte binding proteins of P. vivax comprise a family of proteins involved in host cell selection and erythrocyte invasion. Here we described a member of the gene family found in P. falciparum ( PfRH3 ) that is transcribed in its entirety, under stage-specific control, with correct splicing of the intron, but appears not to be translated, probably due to two reading frameshifts at the 5′ end of the gene.
Publisher: American Association for the Advancement of Science (AAAS)
Date: 31-01-2003
Abstract: Transit peptides mediate protein targeting into plastids and are only poorly understood. We extracted amino acid features from transit peptides that target proteins to the relict plastid (apicoplast) of malaria parasites. Based on these amino acid characteristics, we identified 466 putative apicoplast proteins in the Plasmodium falciparum genome. Altering the specific charge characteristics in a model transit peptide by site-directed mutagenesis severely disrupted organellar targeting in vivo. Similarly, putative Hsp70 (DnaK) binding sites present in the transit peptide proved to be important for correct targeting.
Publisher: Wiley
Date: 03-2004
Publisher: Wiley
Date: 16-01-2014
DOI: 10.1111/MMI.12496
Abstract: Intra-erythrocytic stages of the malaria parasite, Plasmodium falciparum, are thought to be dependent on de novo synthesis of phosphatidylinositol, as red blood cells (RBC) lack the capacity to synthesize this phospholipid. The myo-inositol headgroup of PI can either be synthesized de novo or scavenged from the RBC. An untargeted metabolite profiling of P. falciparum infected RBC showed that trophozoite and schizont stages accumulate high levels of myo-inositol-3-phosphate, indicating increased de novo biosynthesis of myo-inositol from glucose 6-phosphate. Metabolic labelling studies with (13) C-U-glucose in the presence and absence of exogenous inositol confirmed that de novo myo-inositol synthesis occurs in parallel with myo-inositol salvage pathways. Unexpectedly, while both endogenous and scavenged myo-inositol was used to synthesize bulk PI, only de novo-synthesized myo-inositol was incorporated into GPI glycolipids. Moreover, gene disruption studies suggested that the INO1 gene, encoding myo-inositol 3-phosphate synthase, is essential in asexual parasite stages. Together these findings suggest that P. falciparum asexual stages are critically dependent on de novo myo-inositol biosynthesis for assembly of a sub-pool of PI species and GPI biosynthesis. These findings highlight unexpected complexity in phospholipid biosynthesis in P. falciparum and a lack of redundancy in some nutrient salvage versus endogenous biosynthesis pathways.
No related organisations have been discovered for Alan Cowman.
Start Date: 02-2012
End Date: 12-2012
Amount: $350,000.00
Funder: Australian Research Council
View Funded ActivityStart Date: 09-2004
End Date: 08-2009
Amount: $1,500,000.00
Funder: Australian Research Council
View Funded ActivityStart Date: 04-2017
End Date: 12-2017
Amount: $850,000.00
Funder: Australian Research Council
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