The molecular mechanism of cytoadherence to placental or tumor cells through VAR2CSA from Plasmodium falciparum.
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ABSTRACT:
SUBMITTER: Wang W
PROVIDER: S-EPMC8523661 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Plasmodium falciparum VAR2CSA binds to chondroitin sulfate A (CSA) on the surface of the syncytiotrophoblast during placental malaria. This interaction facilitates placental sequestration of malaria parasites resulting in severe health outcomes for both the mother and her offspring. Furthermore, CSA is presented by diverse cancer cells and specific targeting of cells by VAR2CSA may become a viable approach for cancer treatment. In the present study, we determined the cryo-electron microscopy structures of the full-length ectodomain of VAR2CSA from P. falciparum strain NF54 in complex with CSA, and VAR2CSA from a second P. falciparum strain FCR3. The architecture of VAR2CSA is composed of a stable core flanked by a flexible arm. CSA traverses the core domain by binding within two channels and CSA binding does not induce major conformational changes in VAR2CSA. The CSA-binding elements are conserved across VAR2CSA variants and are flanked by polymorphic segments, suggesting immune selection outside the CSA-binding sites. This work provides paths for developing interventions against placental malaria and cancer.
| S-EPMC7914210 | biostudies-literature
Project description:BACKGROUND: Protection against pregnancy associated malaria (PAM) is associated with high levels of anti-VAR2CSA antibodies. This protection is obtained by the parity dependent acquisition of anti-VAR2CSA antibodies. Distinct parity-associated molecular signatures have been identified in VAR2CSA domains. These two observations combined point to the importance of identifying VAR2CSA sequence variation, which facilitate parasitic evasion or subversion of host immune response. Highly conserved domains of VAR2CSA such as DBL5? are likely to contain conserved epitopes, and therefore do constitute attractive targets for vaccine development. METHODOLOGY/PRINCIPAL FINDINGS: VAR2CSA DBL5?-domain sequences obtained from cDNA of 40 placental isolates were analysed by a combination of experimental and in silico methods. Competition ELISA assays on two DBL5? variants, using plasma samples from women from two different areas and specific mice hyperimmune plasma, indicated that DBL5? possess conserved and cross-reactive B cell epitopes. Peptide ELISA identified conserved areas that are recognised by naturally acquired antibodies. Specific antibodies against these peptides labelled the native proteins on the surface of placental parasites. Despite high DBL5? sequence homology among parasite isolates, sequence analyses identified motifs in DBL5? that discriminate parasites according to donor's parity. Moreover, recombinant proteins of two VAR2CSA DBL5? variants displayed diverse recognition patterns by plasma from malaria-exposed women, and diverse proteoglycan binding abilities. CONCLUSIONS/SIGNIFICANCE: This study provides insights into conserved and exposed B cell epitopes in DBL5? that might be a focus for cross reactivity. The importance of sequence variation in VAR2CSA as a critical challenge for vaccine development is highlighted. VAR2CSA conformation seems to be essential to its functionality. Therefore, identification of sequence variation sites in distinct locations within VAR2CSA, affecting antigenicity and/or binding properties, is critical to the effort of developing an efficient VAR2CSA-based vaccine. Motifs associated with parasite segregation according to parity constitute one such site.
| S-EPMC2948511 | biostudies-literature
Project description:BACKGROUND: In Plasmodium falciparum malaria endemic areas placental malaria (PM) is an important complication of malaria. The recurrence of malaria in primigravidae women irrespective of acquired protection during childhood is caused by the interaction between the parasite-expressed VAR2CSA antigen and chondroitin sulfate A (CSA) in the placental intervillous space and lack of protective antibodies. PM impairs fetal development mainly by excessive inflammation processes. After infections during pregnancy women acquire immunity to PM conferred by antibodies against VAR2CSA. Ideally, a vaccine against PM will induce antibody-mediated immune responses that block the adhesion of infected erythrocytes (IE) in the placenta. PRINCIPAL FINDINGS: We have previously shown that antibodies raised in rat against individual domains of VAR2CSA can block IE binding to CSA. In this study we have immunized mice, rats and rabbits with each individual domain and the full-length protein corresponding to the FCR3 VAR2CSA variant. We found there is an inherently higher immunogenicity of C-terminal domains compared to N-terminally located domains. This was irrespective of whether antibodies were induced against single domains or the full-length protein. Species-specific antibody responses were also found, these were mainly directed against single domains and not the full-length VAR2CSA protein. CONCLUSIONS/SIGNIFICANCE: Binding inhibitory antibodies appeared to be against conformational B-cell epitopes. Non-binding inhibitory antibodies reacted highly against the C-terminal end of the VAR2CSA molecule especially the highly polymorphic DBL6? domain. Differential species-specific induction of antibody responses may allow for more direct analysis of functional versus non-functional B-cell epitopes.
| S-EPMC3064590 | biostudies-literature
Project description:BackgroundPlacental malaria is caused by Plasmodium falciparum-infected erythrocytes (IEs) that surface-express VAR2CSA and bind chondroitin sulfate A. The inflammatory response to placenta-sequestered parasites is associated with poor pregnancy outcomes, and protection may be mediated in part by VAR2CSA antibodies that block placental IE adhesion.MethodsIn this study, we used a new approach to assess VAR2CSA domains for functional epitopes recognized by naturally acquired antibodies. Antigen-specific immunoglobulin (Ig) G targeting Duffy binding-like (DBL) domains from different alleles were sequentially purified from plasma pooled from multigravid women and then characterized using enzyme-linked immunosorbent assay, flow cytometry, and antiadhesion assays.ResultsDifferent DBL domain-specific IgGs could react to homologous as well as heterologous antigens and parasites, suggesting that conserved epitopes are shared between allelic variants. Homologous blocking of IE binding was observed with ID1-DBL2-ID2a-, DBL4-, and DBL5-specific IgG (range, 42%-75%), whereas partial cross-inhibition activity was observed with purified IgG specific to ID1-DBL2-ID2a and DBL4 antigens. Plasma retained broadly neutralizing activity after complete depletion of these VAR2CSA specificities.ConclusionsBroadly neutralizing antibodies of multigravidae are not depleted on VAR2CSA recombinant antigens, and hence development of VAR2CSA vaccines based on a single construct and variant might induce antibodies with limited broadly neutralizing activity.
| S-EPMC4957442 | biostudies-literature
Project description:BackgroundThe ability of mature forms of Plasmodium falciparum infected erythrocytes to bind to a range of host receptors including those displayed on endothelial cells has been associated with the pathology of this infection. Investigations into this adhesive phenomenon have used protein and cell-based adhesion assays to quantify the ability of infected red blood cells to bind. These adhesion assays tend to have relatively high inherent variability and so require multiple experiments in order to provide good quantitation. This means that investigators doing these experiments must count many fields of adherent parasites, a task that is time-consuming and laborious. To address this issue and to facilitate cytoadherence research, developed automated protocols were developed for counting parasite adhesion.MethodsParasite adhesion assays were mainly carried out under static conditions using purified receptors, which is the simplest form of these assays and is translatable to the field. Two different software platforms were used, one commercial (Image Pro-Plus (Media Cybernetics)) and one available in the public domain (ImageSXM) based on the freely available NIH Image software. The adhesion assays were performed and parasite binding quantified using standard manual techniques. Images were also captured using video microscopy and analysed using the two automated systems. The results generated by each system were compared using the Bland and Altman method for assessing the agreement between two methods.ResultsBoth automated counting programs showed concordance compared to the 'gold standard' manual counting within the normal range of adhesion seen with these assays, although the ImageSXM technique had some systematic bias. There was some fall-off in accuracy at very high parasite densities, but this can be resolved through good design of the experiments. Cell based assays were also used as inputs to one of the automated systems (ImageSXM) and produced variable, but encouraging, results.ConclusionsThe automated counting programs are an accurate and practical way of quantifying static parasite binding assays to purified proteins. They are less accurate when applied to cell based systems, but can still provide a reasonable level of accuracy to give a semi-quantitative readout.
| S-EPMC3094228 | biostudies-literature
Project description:Over 30 million women living in P. falciparum endemic areas are at risk of developing malaria during pregnancy every year. Placental malaria is characterized by massive accumulation of infected erythrocytes in the intervillous space of the placenta, accompanied by infiltration of immune cells, particularly monocytes. The consequent local inflammation and the obstruction of the maternofetal exchanges can lead to severe clinical outcomes for both mother and child. Even if protection against the disease can gradually be acquired following successive pregnancies, the malaria parasite has developed a large panel of evasion mechanisms to escape from host defense mechanisms and manipulate the immune system to its advantage. Infected erythrocytes isolated from placentas of women suffering from placental malaria present a unique phenotype and express the pregnancy-specific variant VAR2CSA of the Plasmodium falciparum Erythrocyte Membrane Protein (PfEMP1) family at their surface. The polymorphic VAR2CSA protein is able to mediate the interaction of infected erythrocytes with a variety of host cells including placental syncytiotrophoblasts and leukocytes but also with components of the immune system such as non-specific IgM. This review summarizes the described VAR2CSA-mediated host defense evasion mechanisms employed by the parasite during placental malaria to ensure its survival and persistence.
| S-EPMC7900151 | biostudies-literature
Project description:Two vaccines based on Plasmodium falciparum protein VAR2CSA are currently in clinical evaluation to prevent placental malaria (PM), but a deeper understanding of var2csa variability could impact vaccine design. Here we identified atypical extended or truncated VAR2CSA extracellular structures and confirmed one extended structure in a Malian maternal isolate, using a novel protein fragment assembly method for RNA-seq and DNA-seq data. Extended structures included one or two additional DBL domains downstream of the conventional NTS-DBL1X-6ɛ domain structure, with closest similarity to DBLɛ in var2csa and non-var2csa genes. Overall, 4/82 isolates displayed atypical VAR2CSA structures. The maternal isolate expressing an extended VAR2CSA bound to CSA, but its recombinant VAR2CSA bound less well to CSA than VAR2CSANF54 and showed lower reactivity to naturally acquired parity-dependent antibody. Our protein fragment sequence assembly approach has revealed atypical VAR2CSA domain architectures that impact antigen reactivity and function, and should inform the design of VAR2CSA-based vaccines.
| S-EPMC6897902 | biostudies-literature
Project description:In areas of endemicity pregnancy-associated malaria is an important cause of maternal anemia, stillbirth, and delivery of low-birth-weight children. The syndrome is precipitated by the accumulation of Plasmodium falciparum-infected erythrocytes in the placenta, mediated through an interaction between a parasite protein expressed on erythrocytes named variant surface antigen 2-chondroitin sulfate A (VAR2CSA) and CSA on syncytiotrophoblasts. VAR2CSA is a large polymorphic protein consisting of six Duffy binding-like (DBL), domains and with current constraints on recombinant protein production it is not possible to produce entire VAR2CSA recombinant proteins. Furthermore, the presence of polymorphisms has raised the question of whether it is feasible to define VAR2CSA antigens eliciting broadly protective antibodies. Thus, the challenge for vaccine development is to define smaller parts of the molecule which induce antibodies that inhibit CSA binding of different parasite strains. In this study, we produced a large panel of VAR2CSA proteins and raised antibodies against these antigens. We show that antibodies against the DBL4 domain effectively inhibit parasite binding. As the inhibition was not limited to homologous parasite strains, it seems feasible to base a protective malaria vaccine on a single VAR2CSA DBL domain.
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Project description:Malaria is associated with significant microcirculation disorders, especially when the infection reaches its severe stage. This can lead to a range of fatal conditions, from cerebral malaria to multiple organ failure, of not fully understood pathogenesis. It has recently been proposed that a breakdown of the glycocalyx, the carbohydrate-rich layer lining the vascular endothelium, plays a key role in severe malaria, but direct evidence supporting this hypothesis is still lacking. Here, the interactions between Plasmodium falciparum infected red blood cells ( PfRBCs) and endothelial glycocalyx are investigated by developing an in vitro, physiologically relevant model of human microcirculation based on microfluidics. Impairment of the glycocalyx is obtained by enzymatic removal of sialic acid residues, which, due to their terminal location and net negative charge, are implicated in the initial interactions with contacting cells. We show a more than twofold increase of PfRBC adhesion to endothelial cells upon enzymatic treatment, relative to untreated endothelial cells. As a control, no effect of enzymatic treatment on healthy red blood cell adhesion is found. The increased adhesion of PfRBCs is also associated with cell flipping and reduced velocity as compared to the untreated endothelium. Altogether, these results provide a compelling evidence of the increased cytoadherence of PfRBCs to glycocalyx-impaired vascular endothelium, thus supporting the advocated role of glycocalyx disruption in the pathogenesis of this disease.
| S-EPMC6303788 | biostudies-literature
Project description:Development of severe disease in Plasmodium falciparum malaria infection is thought to be, at least in part, due to the sequestration of trophozoite-stage infected red blood cells in the microvasculature. The process of cytoadherence is mediated by binding of the parasite protein PfEMP-1 on the surface of infected red blood cells to endothelial cell receptors. Although antimalarial treatments rapidly kill parasites, significant mortality is still seen in severe malaria, particularly within 24h of hospital admission. We find that cytoadherence of infected red blood cells continues for several hours after killing of the parasite by antimalarials; after 24h treatment using a range of antimalarials binding is approximately one-third the level of untreated parasite cultures. This is consistent with the maintained presence of PfEMP-1 on the surface of drug-treated infected red blood cells. A specific advantage of artesunate over other treatments tested is seen on addition of this drug to younger ring stage parasites, which do not mature to the cytoadherent trophozoite-stage. These findings show that cytoadherence, a potential pathogenic property of P. falciparum infected red blood cells, continues long after the parasite has been killed. These data support the development of adjunctive therapies to reverse the pathophysiological consequences of cytoadherence.
| S-EPMC2814047 | biostudies-literature