scholarly journals Role of Plasmodium falciparum Protein GEXP07 in Maurer’s Cleft Morphology, Knob Architecture, and P. falciparum EMP1 Trafficking

mBio ◽  
2020 ◽  
Vol 11 (2) ◽  
Author(s):  
Emma McHugh ◽  
Olivia M. S. Carmo ◽  
Adam Blanch ◽  
Oliver Looker ◽  
Boyin Liu ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Fluorescent Protein ◽  
Placental Malaria ◽  
Protein Composition ◽  
Surface Expression ◽  
Content Type ◽  
Virulence Protein ◽  
Infected Rbcs ◽  
Loading And Unloading ◽  
Maurer's Clefts

ABSTRACT The malaria parasite Plasmodium falciparum traffics the virulence protein P. falciparum erythrocyte membrane protein 1 (PfEMP1) to the surface of infected red blood cells (RBCs) via membranous organelles, known as the Maurer’s clefts. We developed a method for efficient enrichment of Maurer’s clefts and profiled the protein composition of this trafficking organelle. We identified 13 previously uncharacterized or poorly characterized Maurer’s cleft proteins. We generated transfectants expressing green fluorescent protein (GFP) fusions of 7 proteins and confirmed their Maurer’s cleft location. Using co-immunoprecipitation and mass spectrometry, we generated an interaction map of proteins at the Maurer’s clefts. We identified two key clusters that may function in the loading and unloading of PfEMP1 into and out of the Maurer’s clefts. We focus on a putative PfEMP1 loading complex that includes the protein GEXP07/CX3CL1-binding protein 2 (CBP2). Disruption of GEXP07 causes Maurer’s cleft fragmentation, aberrant knobs, ablation of PfEMP1 surface expression, and loss of the PfEMP1-mediated adhesion. ΔGEXP07 parasites have a growth advantage compared to wild-type parasites, and the infected RBCs are more deformable and more osmotically fragile. IMPORTANCE The trafficking of the virulence antigen PfEMP1 and its presentation at the knob structures at the surface of parasite-infected RBCs are central to severe adhesion-related pathologies such as cerebral and placental malaria. This work adds to our understanding of how PfEMP1 is trafficked to the RBC membrane by defining the protein-protein interaction networks that function at the Maurer’s clefts controlling PfEMP1 loading and unloading. We characterize a protein needed for virulence protein trafficking and provide new insights into the mechanisms for host cell remodeling, parasite survival within the host, and virulence.

scholarly journals The Plasmodium falciparum protein VCAP1 controls Maurer’s cleft morphology, knob architecture and PfEMP1 trafficking

2019 ◽  
Author(s):  
Emma McHugh ◽  
Olivia Carmo ◽  
Adam Blanch ◽  
Oliver Looker ◽  
Boyin Liu ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Protein Interaction ◽  
Protein Trafficking ◽  
Placental Malaria ◽  
Protein Composition ◽  
Surface Expression ◽  
Virulence Protein ◽  
Infected Rbcs ◽  
Loading And Unloading ◽  
Maurer's Clefts

AbstractThe malaria parasite, Plasmodium falciparum, traffics the virulence protein, P. falciparum erythrocyte membrane protein 1 (PfEMP1) to the surface of infected red blood cells (RBCs) via membranous organelles, known as the Maurer’s clefts. We developed a method for efficient enrichment of Maurer’s clefts and profiled the protein composition of this trafficking organelle. We identified 13 previously uncharacterised or poorly characterised Maurer’s cleft proteins. We generated transfectants expressing GFP-fusions of 7 proteins and confirmed their Maurer’s cleft location. Using co-immunoprecipitation and mass spectrometry we have generated a protein interaction map of proteins at the Maurer’s clefts. We identified two key clusters that may function in the loading and unloading of PfEMP1 into and out of the Maurer’s clefts. We focus on a putative PfEMP1 loading complex that includes the newly characterised virulence complex assembly protein 1 (VCAP1). Disruption of VCAP1 causes Maurer’s cleft fragmentation, aberrant knobs, ablation of PfEMP1 surface expression and loss of the PfEMP1 directed adhesion. ΔVCAP1 parasite lines have a growth advantage compared to wildtype parasites; and the infected RBCs are more deformable and more osmotically fragile.ImportanceThe trafficking of the virulence antigen PfEMP1 and its presentation at the knob structures at the surface of parasite infected RBCs is central to severe adhesion related pathologies such as cerebral and placental malaria. This work adds to our understanding of how PfEMP1 is trafficked to the RBC membrane by defining the protein-protein interaction networks that function at the Maurer’s clefts controlling PfEMP1 loading and unloading. This work adds significantly to our understanding of virulence protein trafficking and will provide crucial knowledge that will be required to determine the mechanisms underpinning parasite driven host cell remodelling, parasite survival within the host and virulence mechanisms.

scholarly journals Antibodies to Cryptic Epitopes in Distant Homologues Underpin a Mechanism of Heterologous Immunity between Plasmodium vivax PvDBP and Plasmodium falciparum VAR2CSA

mBio ◽  
2019 ◽  
Vol 10 (5) ◽  
Author(s):  
Catherine J. Mitran ◽  
Angie Mena ◽  
Sedami Gnidehou ◽  
Shanna Banman ◽  
Eliana Arango ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Plasmodium Vivax ◽  
Molecular Mechanism ◽  
Neutralizing Antibodies ◽  
Natural Infection ◽  
Placental Malaria ◽  
Content Type ◽  
Heterologous Immunity ◽  
Cryptic Epitopes

ABSTRACT Many pathogens evolve extensive genetic variation in virulence proteins as a strategy to evade host immunity. This poses a significant challenge for the host to develop broadly neutralizing antibodies. In Plasmodium falciparum, we show that a mechanism to circumvent this challenge is to elicit antibodies to cryptic epitopes that are not under immune pressure. We previously discovered that antibodies to the Plasmodium vivax invasion protein, PvDBP, cross-react with P. falciparum VAR2CSA, a distantly related virulence factor that mediates placental malaria. Here, we describe the molecular mechanism underlying this cross-species immunity. We identified an epitope in subdomain 1 (SD1) within the Duffy binding-like (DBL) domain of PvDBP that gives rise to cross-reactive antibodies to VAR2CSA and show that human antibodies affinity purified against a synthetic SD1 peptide block parasite adhesion to chondroitin sulfate A (CSA) in vitro. The epitope in SD1 is subdominant and highly conserved in PvDBP, and in turn, SD1 antibodies target cryptic epitopes in P. falciparum VAR2CSA. The epitopes in VAR2CSA recognized by vivax-derived SD1 antibodies (of human and mouse origin) are distinct from those recognized by VAR2CSA immune serum. We mapped two peptides in the DBL5ε domain of VAR2CSA that are recognized by SD1 antibodies. Both peptides map to regions outside the immunodominant sites, and antibodies to these peptides are not elicited following immunization with VAR2CSA or natural infection with P. falciparum in pregnancy, consistent with the cryptic nature of these target epitopes. IMPORTANCE In this work, we describe a molecular mechanism of heterologous immunity between two distant species of Plasmodium. Our results suggest a mechanism that subverts the classic parasite strategy of presenting highly polymorphic epitopes in surface antigens to evade immunity to that parasite. This alternative immune pathway can be exploited to protect pregnant women from falciparum placental malaria by designing vaccines to cryptic epitopes that elicit broadly inhibitory antibodies against variant parasite strains.

scholarly journals Antibodies to Escherichia coli-Expressed C-Terminal Domains of Plasmodium falciparum Variant Surface Antigen 2-Chondroitin Sulfate A (VAR2CSA) Inhibit Binding of CSA-Adherent Parasites to Placental Tissue

2013 ◽  
Vol 81 (4) ◽  
pp. 1031-1039 ◽  
Author(s):  
Tracy Saveria ◽  
Andrew V. Oleinikov ◽  
Kathryn Wiliamson ◽  
Richa Chaturvedi ◽  
Joe Lograsso ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Plasmodium Falciparum ◽  
Chondroitin Sulfate ◽  
Surface Antigen ◽  
Transmembrane Protein ◽  
Placental Malaria ◽  
Placental Tissue ◽  
Content Type ◽  
Variant Surface Antigen ◽  
Chondroitin Sulfate A

ABSTRACTPlacental malaria (PM) is characterized by infected erythrocytes (IEs) that selectively bind to chondroitin sulfate A (CSA) and sequester in placental tissue. Variant surface antigen 2-CSA (VAR2CSA), aPlasmodium falciparumerythrocyte membrane protein 1 (PfEMP1) protein family member, is expressed on the surface of placental IEs and mediates adherence to CSA on the surface of syncytiotrophoblasts. This transmembrane protein contains 6 Duffy binding-like (DBL) domains which might contribute to the specific adhesive properties of IEs. Here, we use laboratory isolate 3D7 VAR2CSA DBL domains expressed inEscherichia colito generate antibodies specific for this protein. Flow cytometry results showed that antibodies generated against DBL4ε, DBL5ε, DBL6ε, and tandem double domains of DBL4-DBL5 and DBL5-DBL6 all bind to placental parasite isolates and to lab strains selected for CSA binding but do not bind to children's parasites. Antisera to DBL4ε and to DBL5ε inhibit maternal IE binding to placental tissue in a manner comparable to that for plasma collected from multigravid women. These antibodies also inhibit binding to CSA of several field isolates derived from pregnant women, while antibodies to double domains do not enhance the functional immune response. These data support DBL4ε and DBL5ε as vaccine candidates for pregnancy malaria and demonstrate thatE. coliis a feasible tool for the large-scale manufacture of a vaccine based on these VAR2CSA domains.

scholarly journals Multiple Plasmodium falciparum Erythrocyte Membrane Protein 1 Variants per Genome Can Bind IgM via Its Fc Fragment Fcμ

2015 ◽  
Vol 83 (10) ◽  
pp. 3972-3981 ◽  
Author(s):  
Anine Jeppesen ◽  
Sisse Bolm Ditlev ◽  
Vladyslav Soroka ◽  
Liz Stevenson ◽  
Louise Turner ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Membrane Protein ◽  
Erythrocyte Membrane ◽  
Placental Malaria ◽  
Clinical Manifestations ◽  
Content Type ◽  
Erythrocyte Membrane Protein ◽  
C Terminus ◽  
Adhesive Proteins ◽  
New Evidence

ThePlasmodium falciparumerythrocyte membrane protein 1 (PfEMP1) adhesive proteins expressed on the surfaces of infected erythrocytes (IEs) are of key importance in the pathogenesis ofP. falciparummalaria. Several structurally and functionally defined PfEMP1 types have been associated with severe clinical manifestations, such as cerebral malaria in children and placental malaria in pregnant women. PfEMP1 that can bind the Fc part of IgM (Fcμ) characterizes one such type, although the functional significance of this IgM binding to PfEMP1 remains unclear. In this study, we report the identification and functional analysis of five IgM-binding PfEMP1 proteins encoded byP. falciparumNF54. In addition to the VAR2CSA-type PFL0030c protein, already known to bind Fcμ and to mediate chondroitin sulfate A (CSA)-specific adhesion of IEs in the placenta, we found four PfEMP1 proteins not previously known to bind IgM this way. Although they all contained Duffy binding-like ε (DBLε) domains similar to those in VAR2CSA-type PfEMP1, they did not mediate IE adhesion to CSA, and IgM binding did not shield IEs from phagocytosis of IgG-opsonized IEs. In this way, these new IgM-binding PfEMP1 proteins resemble the rosette-mediating and IgM-binding PfEMP1 HB3VAR06, but none of them mediated formation of rosettes. We could map the capacity for Fc-specific IgM binding to DBLε domains near the C terminus for three of the four PfEMP1 proteins tested. Our study provides new evidence regarding Fc-dependent binding of IgM to PfEMP1, which appears to be a common and multifunctional phenotype.

scholarly journals mSphere of Influence: Structural Insights into the Molecular Mechanism Underlying Placental Malaria

mSphere ◽  
2021 ◽  
Vol 6 (3) ◽  
Author(s):  
Maria del Pilar Quintana
Keyword(s):  
Plasmodium Falciparum ◽  
Chondroitin Sulfate ◽  
Placental Malaria ◽  
Structural Basis ◽  
Content Type ◽  
Link Type ◽  
Structural Details ◽  
Inhibitory Antibodies ◽  
Structural Insights ◽  
Insight Into

ABSTRACT Maria del Pilar Quintana works on immunology and pathogenesis of severe malaria. In this mSphere of Influence article, she reflects on how the papers “Structural basis for placental malaria mediated by Plasmodium falciparum VAR2CSA” (R. Ma, T. Lian, R. Huang, J. P. Renn, J. D. Petersen, J. Zimmerberg, P. E. Duffy, N. H. Tolia, Nat Microbiol 6:380–391, 2021, https://doi.org/10.1038/s41564-020-00858-9) and “Cryo-EM reveals the architecture of placental malaria VAR2CSA and provides molecular insight into chondroitin sulfate binding” (K. Wang, R. Dagil, T. Lavsten, S. K. Misra, C. B. Spliid, Y. Wang, T. Gustavsson, D. R. Sandoval, E. E. Vidal-Calvo, S. Choudary, M. O. Agerback, K. Lindorff-Larsen, M. A. Nielsen, T. G. Theander, J. S. Sharp, T. M. Clausen, P. Gourdon, A. Salanti [Research Square preprint], 2021, https://doi.org/10.21203/rs.3.rs-121821/v1) shed light on the precise structural details behind Plasmodium falciparum VAR2CSA binding to chondroitin sulfate A (CSA) in the placenta and how these novel insights have changed the way she will approach her work toward the discovery of new broadly cross-reactive/inhibitory antibodies targeting VAR2CSA.

scholarly journals CD47-SIRPα Interactions Regulate Macrophage Uptake of Plasmodium falciparum-Infected Erythrocytes and Clearance of MalariaIn Vivo

2016 ◽  
Vol 84 (7) ◽  
pp. 2002-2011 ◽  
Author(s):  
Kodjo Ayi ◽  
Ziyue Lu ◽  
Lena Serghides ◽  
Jenny M. Ho ◽  
Constance Finney ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Fusion Protein ◽  
Regulatory Protein ◽  
Content Type ◽  
Fc Fusion Protein ◽  
Infected Rbcs ◽  
Deficient Mice ◽  
Macrophage Uptake

CD47 engagement by the macrophage signal regulatory protein alpha (SIRPα) inhibits phagocytic activity and protects red blood cells (RBCs) from erythrophagocytosis. The role of CD47-SIRPα in the innate immune response toPlasmodium falciparuminfection is unknown. We hypothesized that disruption of SIRPα signaling may enhance macrophage uptake of malaria parasite-infected RBCs. To test this hypothesis, we examinedin vivoclearance in CD47-deficient mice infected withPlasmodium bergheiANKA andin vitrophagocytosis ofP. falciparum-infected RBCs by macrophages from SHP-1-deficient (Shp-1−/−) mice and NOD.NOR-Idd13.Prkdcscid(NS-Idd13) mice, as well as human macrophages, following disruption of CD47-SIRPα interactions with anti-SIRPα antibodies or recombinant SIRPα-Fc fusion protein. Compared to their wild-type counterparts,Cd47−/−mice displayed significantly lower parasitemia, decreased endothelial activation, and enhanced survival. Using macrophages from SHP-1-deficient mice or from NS-Idd13mice, which express a SIRPα variant that does not bind human CD47, we showed that altered SIRPα signaling resulted in enhanced phagocytosis ofP. falciparum-infected RBCs. Moreover, disrupting CD47-SIRPα engagement using anti-SIRPα antibodies or SIRPα-Fc fusion protein also increased phagocytosis ofP. falciparum-infected RBCs. These results indicate an important role for CD47-SIRPα interactions in innate control of malaria and suggest novel targets for intervention.

scholarly journals Genesis of and Trafficking to the Maurer's Clefts of Plasmodium falciparum-Infected Erythrocytes

2006 ◽  
Vol 26 (11) ◽  
pp. 4074-4085 ◽  
Author(s):  
Cornelia Spycher ◽  
Melanie Rug ◽  
Nectarios Klonis ◽  
David J. P. Ferguson ◽  
Alan F. Cowman ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Fluorescent Protein ◽  
Transmembrane Domain ◽  
Imaging Techniques ◽  
Parasitophorous Vacuole ◽  
Time Lapse ◽  
Cell Periphery ◽  
Parasitophorous Vacuole Membrane ◽  
Vacuole Membrane ◽  
Maurer's Clefts

ABSTRACT Malaria parasites export proteins beyond their own plasma membrane to locations in the red blood cells in which they reside. Maurer's clefts are parasite-derived structures within the host cell cytoplasm that are thought to function as a sorting compartment between the parasite and the erythrocyte membrane. However, the genesis of this compartment and the signals directing proteins to the Maurer's clefts are not known. We have generated Plasmodium falciparum-infected erythrocytes expressing green fluorescent protein (GFP) chimeras of a Maurer's cleft resident protein, the membrane-associated histidine-rich protein 1 (MAHRP1). Chimeras of full-length MAHRP1 or fragments containing part of the N-terminal domain and the transmembrane domain are successfully delivered to Maurer's clefts. Other fragments remain trapped within the parasite. Fluorescence photobleaching and time-lapse imaging techniques indicate that MAHRP1-GFP is initially trafficked to isolated subdomains in the parasitophorous vacuole membrane that appear to represent nascent Maurer's clefts. The data suggest that the Maurer's clefts bud from the parasitophorous vacuole membrane and diffuse within the erythrocyte cytoplasm before taking up residence at the cell periphery.

scholarly journals Environmentally Regulated Glycosome Protein Composition in the African Trypanosome

2013 ◽  
Vol 12 (8) ◽  
pp. 1072-1079 ◽  
Author(s):  
Sarah Bauer ◽  
James C. Morris ◽  
Meredith T. Morris
Keyword(s):  
Environmental Conditions ◽  
Fluorescent Protein ◽  
Protein Composition ◽  
Bimodal Distribution ◽  
Live Cells ◽  
Reporter System ◽  
Content Type ◽  
Parasite Survival ◽  
Low Glucose ◽  
Parasite Populations

ABSTRACT Trypanosomes compartmentalize many metabolic enzymes in glycosomes, peroxisome-related microbodies that are essential to parasite survival. While it is understood that these dynamic organelles undergo profound changes in protein composition throughout life cycle differentiation, the adaptations that occur in response to changes in environmental conditions are less appreciated. We have adopted a fluorescent-organelle reporter system in procyclic Trypanosoma brucei by expressing a fluorescent protein (FP) fused to a glycosomal targeting sequence (peroxisome-targeting sequence 2 [PTS2]). In these cell lines, PTS2-FP is localized within import-competent glycosomes, and organelle composition can be analyzed by microscopy and flow cytometry. Using this reporter system, we have characterized parasite populations that differ in their glycosome composition. In glucose-rich medium, two parasite populations are observed; one population harbors glycosomes bearing the full repertoire of glycosome proteins, while the other parasite population contains glycosomes that lack the usual glycosome-resident proteins but do contain the glycosome membrane protein TbPEX11. Interestingly, these cells lack TbPEX13, a protein essential for the import of proteins into the glycosome. This bimodal distribution is lost in low-glucose medium. Furthermore, we have demonstrated that changes in environmental conditions trigger changes in glycosome protein composition. These findings demonstrate a level of procyclic glycosome diversity heretofore unappreciated and offer a system by which glycosome dynamics can be studied in live cells. This work adds to our growing understanding of how the regulation of glycosome composition relates to environmental sensing.

scholarly journals Virulence determinant, PTP7, controls vesicle budding from the Maurer’s clefts, adhesin protein trafficking and host cell remodeling in Plasmodium falciparum

2021 ◽  
Author(s):  
Olivia M. S. Carmo ◽  
Gerald J Shami ◽  
Dezerae Cox ◽  
Boyin Liu ◽  
Adam J Blanch ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Cell Surface ◽  
Blood Cell ◽  
Red Blood Cell ◽  
Protein Trafficking ◽  
Low Complexity ◽  
Vesicle Budding ◽  
Cell Remodeling ◽  
Virulence Protein ◽  
Maurer's Clefts

Presentation of the variant antigen, Plasmodium falciparum erythrocyte membrane protein 1 (EMP1), at knob-like protrusions on the surface of infected red blood cells, underpins P. falciparum malaria pathogenicity. Here we describe a protein PF3D7_0301700 (PTP7), that functions at the nexus between the intermediate trafficking organelle, the Maurer’s cleft, and the red blood cell surface. Genetic disruption of PTP7 leads to accumulation of vesicles at the Maurer’s clefts, grossly aberrant knob morphology, and failure to deliver EMP1 to the red blood cell surface.  We show that an expanded low complexity sequence in the C-terminal region of PTP7, found only in the Laverania clade of Plasmodium , is critical for efficient virulence protein trafficking.

scholarly journals Tracking Glideosome-Associated Protein 50 Reveals the Development and Organization of the Inner Membrane Complex of Plasmodium falciparum

2011 ◽  
Vol 10 (4) ◽  
pp. 556-564 ◽  
Author(s):  
Jeffrey A. Yeoman ◽  
Eric Hanssen ◽  
Alexander G. Maier ◽  
Nectarios Klonis ◽  
Bohumil Maco ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Fluorescent Protein ◽  
Early Stage ◽  
Tail Domain ◽  
Structured Illumination Microscopy ◽  
Membrane Anchor ◽  
Inner Membrane ◽  
Content Type ◽  
Membrane Complex ◽  
Inner Membrane Complex

ABSTRACT The most deadly of the human malaria parasites, Plasmodium falciparum , has different stages specialized for invasion of hepatocytes, erythrocytes, and the mosquito gut wall. In each case, host cell invasion is powered by an actin-myosin motor complex that is linked to an inner membrane complex (IMC) via a membrane anchor called the glideosome-associated protein 50 (PfGAP50). We generated P. falciparum transfectants expressing green fluorescent protein (GFP) chimeras of PfGAP50 (PfGAP50-GFP). Using immunoprecipitation and fluorescence photobleaching, we show that C-terminally tagged PfGAP50-GFP can form a complex with endogenous copies of the linker protein PfGAP45 and the myosin A tail domain-interacting protein (MTIP). Full-length PfGAP50-GFP is located in the endoplasmic reticulum in early-stage parasites and then redistributes to apical caps during the formation of daughter merozoites. In the final stage of schizogony, the PfGAP50-GFP profile extends further around the merozoite surface. Three-dimensional (3D) structured illumination microscopy reveals the early-stage IMC as a doubly punctured flat ellipsoid that separates to form claw-shaped apposed structures. A GFP fusion of PfGAP50 lacking the C-terminal membrane anchor is misdirected to the parasitophorous vacuole. Replacement of the acid phosphatase homology domain of PfGAP50 with GFP appears to allow correct trafficking of the chimera but confers a growth disadvantage.

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