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During blood-stage infection, malaria parasites export numerous proteins to the host erythrocyte. The Poly-Helical Interspersed Sub-Telomeric (PHIST) proteins are an exported family that share a common 'PRESAN' domain, and include numerous members in P. falciparum, P. vivax and P. knowlesi. In P. falciparum, PHIST proteins have been implicated in p...
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... characterise the localisation of exported PHISTb pro- teins, we expressed a range PHISTb family members [31] with C-terminal GFP tags, in P. falciparum parasites. The cloned genes encoded full-length PF3D7 0201600, PF3D7 0401800 (PFD80), PF3D7 0424600, PF3D7 0532400, PF3D7 0936600, PF3D7 1102500, PF3D7 1252700 and PF3D7 1476200 (see Table 1 for details on alternative gene identifiers). ...
Citations
... In the sporozoite stage, those include thrombospondin-related anonymous protein (TRAP), also known as sporozoite surface protein 2 (SSP2), apical membrane antigen 1 (AMA1), liver stage antigen 1 (LSA1), and exported protein 1 (Exp-1), also known as circumsporozoite-related antigen (CRA). Additionally, we included known surface proteins that can be found in other stages of the infection cycle like the family of monomeric serine-threonine protein kinases (FIKK, Anil Kumar et al. 2021), the helical intersperse sub-telomeric family of exported proteins (PHIST, Tarr et al. 2014), and the multigene family of cytoadherence linked asexual gene (CLAG, Gupta et al. 2015). ...
Motivation:
Machine learning methods can be used to support scientific discovery in healthcare-related research fields. However, these methods can only be reliably used if they can be trained on high-quality and curated datasets. Currently, no such dataset for the exploration of Plasmodium falciparum protein antigen candidates exists. The parasite P.falciparum causes the infectious disease malaria. Thus, identifying potential antigens is of utmost importance for the development of antimalarial drugs and vaccines. Since exploring antigen candidates experimentally is an expensive and time-consuming process, applying machine learning methods to support this process has the potential to accelerate the development of drugs and vaccines, which are needed for fighting and controlling malaria.
Results:
We developed PlasmoFAB, a curated benchmark that can be used to train machine learning methods for the exploration of P.falciparum protein antigen candidates. We combined an extensive literature search with domain expertise to create high-quality labels for P.falciparum specific proteins that distinguish between antigen candidates and intracellular proteins. Additionally, we used our benchmark to compare different well-known prediction models and available protein localization prediction services on the task of identifying protein antigen candidates. We show that available general-purpose services are unable to provide sufficient performance on identifying protein antigen candidates and are outperformed by our models that were trained on this tailored data.
Availability and implementation:
PlasmoFAB is publicly available on Zenodo with DOI 10.5281/zenodo.7433087. Furthermore, all scripts that were used in the creation of PlasmoFAB and the training and evaluation of machine learning models are open source and publicly available on GitHub here: https://github.com/msmdev/PlasmoFAB.
... These proteins peripherally-localised in infected erythrocytes and in P. falciparum are involved in functions such as protein trafficking, membrane rigidity and intercellular signalling. 90 Other loci identified included the leucine-rich repeat protein (pvlrr8) and the surface protein pvmsp1 along with a paralog pvmsp1p-19 91 (Table S13). Within South America, we looked for signals of positive selection at the country level (for countries with >10 isolates). ...
Background
Brazil is a unique and understudied setting for malaria, with complex foci of transmission associated with human and environmental conditions. An understanding of the population genomic diversity of P. vivax parasites across Brazil can support malaria control strategies.
Methods
Through whole genome sequencing of P. vivax isolates across 7 Brazilian states, we use population genomic approaches to compare genetic diversity within country (n = 123), continent (6 countries, n = 315) and globally (26 countries, n = 885).
Findings
We confirm that South American isolates are distinct, have more ancestral populations than the other global regions, with differentiating mutations in genes under selective pressure linked to antimalarial drugs (pvmdr1, pvdhfr-ts) and mosquito vectors (pvcrmp3, pvP45/48, pvP47). We demonstrate Brazil as a distinct parasite population, with signals of selection including ABC transporter (PvABCI3) and PHIST exported proteins.
Interpretation
Brazil has a complex population structure, with evidence of P. simium infections and Amazonian parasites separating into multiple clusters. Overall, our work provides the first Brazil-wide analysis of P. vivax population structure and identifies important mutations, which can inform future research and control measures.
Funding
AI is funded by an MRC LiD PhD studentship. TGC is funded by the Medical Research Council (Grant no. MR/M01360X/1, MR/N010469/1, MR/R025576/1, MR/R020973/1 and MR/X005895/1). SC is funded by Medical Research Council UK grants (MR/M01360X/1, MR/R025576/1, MR/R020973/1 and MR/X005895/1) and Bloomsbury SET (ref. CCF17-7779). FN is funded by The Shloklo Malaria Research Unit - part of the Mahidol Oxford Research Unit, supported by the Wellcome Trust (Grant no. 220211). ARSB is funded by São Paulo Research Foundation - FAPESP (Grant no. 2002/09546–1). RLDM is funded by Brazilian National Council for Scientific and Technological Development - CNPq (Grant no. 302353/2003–8 and 471605/2011–5); CRFM is funded by FAPESP (Grant no. 2020/06747–4) and CNPq (Grant no. 302917/2019–5 and 408636/2018–1); JGD is funded by FAPESP fellowships (2016/13465–0 and 2019/12068–5) and CNPq (Grant no. 409216/2018–6).
... In the sporozoite stage, those include thrombospondin-related anonymous protein (TRAP), also known as sporozoite surface protein 2 (SSP2), apical membrane antigen 1 (AMA1), liver stage antigen 1 (LSA1), and exported protein 1 (Exp-1), also known as circumsporozoite-related antigen (CRA). Additionally, we included known surface proteins that can be found in other stages of the infection cycle like the family of monomeric serine-threonine protein kinases (FIKK, AK et al. (2021)), the helical intersperse sub-telomeric family of exported proteins (PHIST, Tarr et al. (2014)), and the multigene family of cytoadherence linked asexual gene (CLAG, Gupta et al. (2015)). ...
Motivation: Machine learning methods can be used to support scientific discovery in healthcare-related research fields. However, these methods can only be reliably used if they can be trained on high-quality and curated datasets. Currently, no such dataset for the exploration of Plasmodium falciparum protein antigen candidates exists. The parasite Plasmodium falciparum causes the infectious disease malaria. Thus, identifying potential antigens is of utmost importance for the development of antimalarial drugs and vaccines. Since exploring antigen candidates experimentally is an expensive and time-consuming process, applying machine learning methods to support this process has the potential to accelerate the development of drugs and vaccines which are needed for fighting and controlling malaria. Results: We developed PlasmoFAB, a curated benchmark that can be used to train machine learning methods for the exploration of Plasmodium falciparum protein antigen candidates. We combined an extensive literature search with domain expertise to create high-quality labels for Plasmodium falciparum specific proteins that distinguish between antigen candidates and intracellular proteins. Additionally, we used our benchmark to compare different well-known prediction models and available protein localization prediction services on the task of identifying protein antigen candidates. We show that available general-purpose services are unable to provide sufficient performance on identifying protein antigen candidates and are outperformed by models that were trained on specialized data.
... We found other interesting genes in this list. First, PF3D7_1372000 is a Plasmodium exported protein of the Poly-Helical Interspersed Sub-Telomeric (PHIST) protein family (Tarr et al., 2014;Warncke et al., 2016), also known as the PRESAN family (Oakley et al., 2007;Sargeant et al., 2006). Although detailed functions of most Plasmodium exported proteins are yet to be revealed, in general, the parasite exported proteins are pivotal for parasite survival by interacting and interfering activities of the infected cells (Maier et al., 2008). ...
... Pfmdr1 (PF3D7_0523000), P. falciparum multidrug drug resistance gene 1, was identified among the shared top genes by both in vivo and in vitro datasets, consistent with previous reports stating that it plays an essential role in the response processes of a broad range of ACT antimalarials (Chavchich et al., 2010;Dahlströ m et al., 2009;Eastman et al., 2016;Gupta et al., 2014;Holmgren et al., 2006Holmgren et al., , 2007Imwong et al., 2010;Ngalah et al., 2015;Ould Ahmedou Salem et al., 2017;Sidhu et al., 2006;Sisowath et al., 2007;Ursing et al., 2006). PF3D7_1372000, a Plasmodium exported protein of the Poly-Helical Interspersed Sub-Telomeric (PHIST) protein family (Tarr et al., 2014;Warncke et al., 2016), was also identified among the shared top genes. Literature has reported that the parasite exported proteins are pivotal for parasite survival by interacting and interfering activities of the infected cells (Maier et al., 2008). ...
Drug resistance has been rapidly evolving with regard to the first-line malaria treatment, artemisinin-based combination therapies. It has been an open question whether predictive models for this drug resistance status can be generalized across in vivo-in vitro transcriptomic measurements. In this study, we present a model that predicts artemisinin treatment resistance developed with transcriptomic information of Plasmodium falciparum. We demonstrated the robustness of this model across in vivo clearance rate and in vitro IC50 measurement, and based on different microarray and data processing modalities. The validity of the algorithm is further supported by its first-placement in the DREAM Malaria Challenge. We identified transcription biomarkers to artemisinin treatment resistance that can predict artemisinin resistance and are conserved in their expression modules. This is a critical step in the research of malaria treatment as it demonstrated the potential of a platform-robust, personalized model for artemisinin resistance using molecular biomarkers.
... Among the exported proteins, the PHIST family, containing 89 proteins, plays a significant role in enhancing the cytoadhesion of the reconstructed host cell [60]. The authors showed that sulfated polysaccharides can deactivate the proteins exported by the parasite, associated with control of the main virulent factor of the parasite P. falciparum erythrocyte membrane protein (PfEMP1) [61]. ...
This review presents materials characterizing sulfated polysaccharides (SPS) of marine hydrobionts (algae and invertebrates) as potential means for the prevention and treatment of protozoa and helminthiasis. The authors have summarized the literature on the pathogenetic targets of protozoa on the host cells and on the antiparasitic potential of polysaccharides from red, brown and green algae as well as certain marine invertebrates. Information about the mechanisms of action of these unique compounds in diseases caused by protozoa has also been summarized. SPS is distinguished by high antiparasitic activity, good solubility and an almost complete absence of toxicity. In the long term, this allows for the consideration of these compounds as effective and attractive candidates on which to base drugs, biologically active food additives and functional food products with antiparasitic activity.
... plasm odb. org) and are unified by possessing a single domain termed PHIST that is predicted to be composed solely of alpha helices 52,53 . This family of exported proteins are conserved across the human Plasmodium species; P. falciparum, P. vivax and P. knowlesi 52 . ...
... org) and are unified by possessing a single domain termed PHIST that is predicted to be composed solely of alpha helices 52,53 . This family of exported proteins are conserved across the human Plasmodium species; P. falciparum, P. vivax and P. knowlesi 52 . As we identified a transmembrane domain and GPI anchors, we expect phistb to be surface anchored 54 . ...
Genetic diversity of surface exposed and stage specific Plasmodium falciparum immunogenic proteins pose a major roadblock to developing an effective malaria vaccine with broad and long-lasting immunity. We conducted a prospective genetic analysis of candidate antigens ( msp1, ama1, rh5, eba175, glurp, celtos, csp, lsa3, Pfsea, trap, conserved chrom3 , hyp9 , hyp10 , phistb, surfin8.2, and surfin14.1 ) for malaria vaccine development on 2375 P. falciparum sequences from 16 African countries. We described signatures of balancing selection inferred from positive values of Tajima’s D for all antigens across all populations except for glurp. This could be as a result of immune selection on these antigens as positive Tajima’s D values mapped to regions with putative immune epitopes. A less diverse phistb antigen was characterised with a transmembrane domain, glycophosphatidyl anchors between the N and C- terminals, and surface epitopes that could be targets of immune recognition. This study demonstrates the value of population genetic and immunoinformatic analysis for identifying and characterising new putative vaccine candidates towards improving strain transcending immunity, and vaccine efficacy across all endemic populations.
... Indirect immunofluorescence assays were used to confirm the localisation of the 13 proteins ( Fig. 1). RESA1, PF3D7_1201000, PF3D7_0401800, PF3D7_0424600 and LyMP displayed a strong signal at the surface of the iRBC, which was often accompanied by a weaker signal in the cytoplasm (Fig. 1i), in agreement with previous studies (Proellocks et al., 2014, Tarr et al., 2014, Tiburcio et al., 2015, Davies et al., 2016, Moreira et al., 2016. PF3D7_0532300 has been previously suggested to localise to the MCs (Moreira et al., 2016), which we confirm here through co-localisation with the MC marker REX1 but we also observed signal lining the iRBC surface (Fig. 1i). ...
... Here we confirm previous observations for the localisation of: RESA1, PF3D7_0401800, PF3D7_0424600, PF3D7_1201000, LyMP, PF3D7_0301700 ( Fig. 5) (Proellocks et al., 2014, Tarr et al., 2014, Schulze et al., 2015, Tiburcio et al., 2015, Davies et al., 2016, Moreira et al., 2016. Furthermore, we have also been able to confirm the prediction studies suggesting that PF3D7_0532300, PTP4, PF3D7_1477500, Hyp1, PF3D7_0113200, PF3D7_0501000 and PF3D7_1401200 are exported into the host cell cytoplasm, and demonstrate that they have diffuse or punctate localisation. ...
... PF3D7_0532300 only interacted with RhopH components under weaker detergent conditions indicating this association is likely weak, transient or indirect (Fig. 3). RESA1, PF3D7_1201000, PF3D7_0401800 and LyMP all localise to the iRBC surface and have been shown previously to interact with cytoskeletal components (Fig. 5) (Foley et al., 1991, Da Silva et al., 1994, Silva et al., 2005, Pei et al., 2007, Proellocks et al., 2014, Tarr et al., 2014. Only RESA1 and PF3D7_1201000 showed stronger association to cytoskeletal components compared to that of the other exported proteins targeted here and therefore might be associating with RhopH components due to their localisation and not direct association (Table 1B). ...
During its intraerythrocytic life cycle, the human malaria parasite Plasmodium falciparum supplements its nutritional requirements by scavenging substrates from the plasma through the new permeability pathways (NPPs) installed in the red blood cell (RBC) membrane. Parasite proteins of the RhopH complex: CLAG3, RhopH2, RhopH3, have been implicated in NPP activity. Here we studied 13 exported proteins previously hypothesised to interact with RhopH2, to study their potential contribution to the function of NPPs. NPP activity assays revealed that the 13 proteins do not appear to be individually important for NPP function, as conditional knockdown of these proteins had no effect on sorbitol uptake. Intriguingly, reciprocal immunoprecipitation assays showed that five of the 13 proteins interact with all members of the RhopH complex, with PF3D7_1401200 showing the strongest association. Mass spectrometry‐based proteomics further identified new protein complexes; a cytoskeletal complex and a Maurer’s clefts/J‐dot complex, which overall helps clarify protein‐protein interactions within the infected RBC (iRBC) and is suggestive of the potential trafficking route of the RhopH complex itself to the RBC membrane.
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... Different PHIST members are located in distinct subcellular regions. Several are exported to the iRBC surface (Oberli et al., 2014;Tarr et al., 2014;Kumar et al., 2018), while PFE1605w comigrates with PfEMP1 within iRBCs before being located at knobs, where it likely functions as an anchor of PfEMP1 (Oberli et al., 2014). However, PFI1780w was found to Affinity between PF3D7_1372300 and ATS measured by dot blot. ...
Plasmodium falciparum extensively remodels host cells by translocating numerous proteins into the cytoplasm of red blood cells (RBCs) after invasion. Among these exported proteins, members of the Plasmodium helical interspersed subtelomeric (PHIST) family are crucial for host cell remodeling and host-parasite interactions, and thereby contribute to malaria pathogenesis. Herein, we explored the function of PF3D7_1372300, a member of the PHIST/PHISTa-like subfamily. PF3D7_1372300 was highly transcribed and expressed during the blood stage of P. falciparum, and distributed throughout RBCs, but most abundant at the erythrocyte membrane. Specific interaction of PF3D7_1372300 with the cytoplasmic tail of P. falciparum erythrocyte membrane protein 1 (PfEMP1) was revealed by immunofluorescence assay, in vitro intermolecular interaction assays. The interaction sites of PF3D7_1372300 with PfEMP1 ATS domain were found involved more than 30 amino acids (aa) at several positions. The findings deepen our understanding of host-parasite interactions and malaria pathogenesis.
... The PHIST proteins are known to be exported to various locations including both the host cell periphery and cytosol, as well as the parasite's parasitophorous vacuole, Maurer's cleft and merozoite surface [64][65][66][67] . They function by interacting with host erythrocyte cytoskeletal components 64,65,68,69 as well as parasite-specific proteins such as PfEMP1, skeleton binding protein 1 (SBP1), and knob associated histidine rich protein (KAHRP) 66,70,71 , ultimately leading to cytoadherension and resistance to stress 68,72 . ...
... The PHIST proteins are known to be exported to various locations including both the host cell periphery and cytosol, as well as the parasite's parasitophorous vacuole, Maurer's cleft and merozoite surface [64][65][66][67] . They function by interacting with host erythrocyte cytoskeletal components 64,65,68,69 as well as parasite-specific proteins such as PfEMP1, skeleton binding protein 1 (SBP1), and knob associated histidine rich protein (KAHRP) 66,70,71 , ultimately leading to cytoadherension and resistance to stress 68,72 . PHIST proteins such as members of the PFEMP1 trafficking protein (PTP) contribute to cytoadherence by mediating the successful trafficking of PfEMP1 from the Maurer's cleft to the host cell surface 43 . ...
The central role that erythrocyte invasion plays in Plasmodium falciparum survival and reproduction makes this process an attractive target for therapeutic or vaccine development. However, multiple invasion-related genes with complementary and overlapping functions afford the parasite the plasticity to vary ligands used for invasion, leading to phenotypic variation and immune evasion. Overcoming the challenge posed by redundant ligands requires a deeper understanding of conditions that select for variant phenotypes and the molecular mediators. While host factors including receptor heterogeneity and acquired immune responses may drive parasite phenotypic variation, we have previously shown that host-independent changes in invasion phenotype can be achieved by continuous culturing of the W2mef and Dd2 P. falciparum strains in moving suspension as opposed to static conditions. Here, we have used a highly biologically replicated whole transcriptome sequencing approach to identify the molecular signatures of variation associated with the phenotype switch. The data show increased expression of particular invasion-related genes in switched parasites, as well as a large number of genes encoding proteins that are either exported or form part of the export machinery. The genes with most markedly increased expression included members of the erythrocyte binding antigens (EBA), reticulocyte binding homologues (RH), surface associated interspersed proteins (SURFIN), exported protein family 1 (EPF1) and Plasmodium Helical Interspersed Sub-Telomeric (PHIST) gene families. The data indicate changes in expression of a repertoire of genes not previously associated with erythrocyte invasion phenotypes, suggesting the possibility that moving suspension culture may also select for other traits.
... Amongst the proteins known or predicted to be exported, 89 proteins belong to the Plasmodium helical interspersed subtelomeric (PHIST) family (Hiller et al., 2004;Marti et al., 2004;Sargeant et al., 2006;Warncke, Vakonakis, & Beck, 2016). Several PHIST proteins have been shown to localise and interact with the erythrocyte cytoskeleton in asexual stages of P. falciparum (Oberli et al., 2014;Proellocks et al., 2014;Tarr, Moon, Hardege, & Osborne, 2014;Warncke et al., 2016) and thus, have been proposed to play a role in the remodelling of the erythrocyte cytoskeleton. ...
... The 10 erythrocytes or parasite proteins with the lowest p value are shown in Figure 2c. All parasite proteins included in Figure 2c have been shown or are predicted to be exported (Boddey et al., 2013;Oberli et al., 2014;Tarr et al., 2014), and one (PF3D7_0114000 or GEXP06) has previously been identified as a gametocyte exported protein (Silvestrini et al., 2010). In parallel, we performed another co-IP experiment under native conditions without crosslinking, and only four proteins were found to be enriched, namely, Ankyrin, Band 3, and both subunits of Spectrin, all of which are part of the human Band 3 complex that anchors the erythrocyte cytoskeleton to the membrane ( Figure 2d; Bennett & Stenbuck, 1979). ...
... In fact, we identified two PHIST proteins that colocalised with GEXP02 and might fulfill identical or similar roles as GEXP02. It is noteworthy that all hitherto examined PHISTb proteins target the host cytoskeleton as has been shown with several episomally overexpressed PHISTb-GFP fusion proteins (including GEXP02; Tarr et al., 2014). It is also important to realise that all gametocyte experiments reported here were performed in vitro, and it is conceivable that GEXP02 may play an important role in the biology of gametocytes in vivo, which is currently extremely difficult if not impossible to be measured for reasons of technical limitations. ...
A hallmark of the biology of Plasmodium falciparum blood stage parasites is their extensive host cell remodeling, facilitated by parasite proteins that are exported into the erythrocyte. Although this area has received extensive attention, only a few exported parasite proteins have been analyzed in detail and much of this remodeling process remains unknown, particularly for gametocyte development. Recent advances to induce high rates of sexual commitment enable the production of large numbers of gametocytes. We used this approach to study the Plasmodium helical interspersed subtelomeric (PHIST) protein GEXP02, which is expressed during sexual development. We show by immunofluorescence that GEXP02 is exported to the gametocyte‐infected host cell periphery. Co‐immunoprecipitation revealed potential interactions between GEXP02 and components of the erythrocyte cytoskeleton as well as other exported parasite proteins. This indicates that GEXP02 targets the erythrocyte cytoskeleton and is likely involved in its remodeling. GEXP02 knock‐out parasites show no obvious phenotype during gametocyte maturation, transmission through mosquitoes, and hepatocyte infection, suggesting auxiliary or redundant functions for this protein. In summary, we performed a detailed cellular and biochemical analysis of a sexual stage‐specific exported parasite protein using a novel experimental approach that is broadly applicable to study the biology of P. falciparum gametocytes.
