PV1, a novel Plasmodium falciparum merozoite dense granule protein, interacts with exported protein in infected erythrocytes

Morita, Masayuki; Nagaoka, Hikaru; Ntege, Edward H.; Kanoi, Bernard N.; Ito, Daisuke; Nakata, Takashi; Lee, Jiwon; Tokunaga, Kazutoshi; Iimura, Tadahiro; Torii, Motomi; Tsuboi, Takafumi; Takashima, Eizo

doi:10.1038/s41598-018-22026-0

Cited by 32 publications

(36 citation statements)

References 37 publications

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“…1C). In merozoites, mCherry fluorescence was concentrated in a punctate intraparasitic region, perhaps signifying storage of PbPV5 in the dense granules, as has been demonstrated for several other important PV proteins (30)(31)(32), but this fraction was minimal as compared to the protein contained in the residual body. Quantification of the mCherry-fluorescence intensity in live parasites indicated that PbPV5 is much more abundant in mature parasite stages than in rings and merozoites, suggesting substantial levels of de novo synthesis throughout parasite maturation (Fig.…”

Section: Resultsmentioning

confidence: 57%

A lipocalin mediates unidirectional heme biomineralization in malaria parasites

Matz

Drepper

Blum

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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During blood-stage development, malaria parasites are challenged with the detoxification of enormous amounts of heme released during the proteolytic catabolism of erythrocytic hemoglobin. They tackle this problem by sequestering heme into bioinert crystals known as hemozoin. The mechanisms underlying this biomineralization process remain enigmatic. Here, we demonstrate that both rodent and human malaria parasite species secrete and internalize a lipocalin-like protein, PV5, to control heme crystallization. Transcriptional deregulation ofPV5in the rodent parasitePlasmodium bergheiresults in inordinate elongation of hemozoin crystals, while conditionalPV5inactivation in the human malaria agentPlasmodium falciparumcauses excessive multidirectional crystal branching. Although hemoglobin processing remains unaffected, PV5-deficient parasites generate less hemozoin. Electron diffraction analysis indicates that despite the distinct changes in crystal morphology, neither the crystalline order nor unit cell of hemozoin are affected by impaired PV5 function. Deregulation ofPV5expression rendersP. bergheihypersensitive to the antimalarial drugs artesunate, chloroquine, and atovaquone, resulting in accelerated parasite clearance following drug treatment in vivo. Together, our findings demonstrate thePlasmodium-tailored role of a lipocalin family member in hemozoin formation and underscore the heme biomineralization pathway as an attractive target for therapeutic exploitation.

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Section: Resultsmentioning

confidence: 57%

A lipocalin mediates unidirectional heme biomineralization in malaria parasites

Matz

Drepper

Blum

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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“…Therefore, the molecular mediated parasites residing in the host cell might be fewer and more infertile for Babesia than Plasmodium spp. In Plasmodium spp., many proteins were reported to be discharged into the cytoplasm or cytoplasmic face of iRBC to internalize and stabilize the host cell environment for parasite survival and replication [ 16 – 18 , 52 – 54 ]. For example, ring-infected erythrocyte surface antigen (RESA) proteins were secreted by dense granules, and then were located to the cytoplasmic face of iRBC through binding the repeat 16 of beta-chain for stabilizing the spectrin tetramer against mechanical and thermal damage [ 16 , 17 , 54 ].…”

Section: Discussionmentioning

confidence: 99%

“…Dense granules are reported to release proteins into the parasitophorous vacuole (PV) shortly after invasion, which may play a key role in PV membrane modifications and is assumed to be associated with the nutrient acquisition. Likewise, the proteins secreted by dense granules are also translocated to the cytoplasmic side of infected red blood cell (iRBC) and participate in stabilizing spectrin tetramers [ 15 – 18 ]. However, in Babesia spp.…”

Section: Introductionmentioning

confidence: 99%

Characterization of a novel secretory spherical body protein in Babesia orientalis and Babesia orientalis-infected erythrocytes

Guo

Sun

et al. 2018

Parasites Vectors

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BackgroundThe spherical body, a membrane bound organelle localized in the apical organelle complex, is unique to Babesia and Theileria spp. The spherical body proteins (SBPs) secreted by spherical bodies include SBP1, SBP2, SBP3 and SBP4. Up to now, only SBP3 has been characterized in Babesia orientalis.MethodsThe BoSBP4 gene was amplified from cDNA and gDNA and cloned into the pGEX-6P-1 vector by homologous recombination, sequenced and analyzed by bioinformatics tools. The amino acid (aa) sequence of BoSBP4 was compared with that of Babesia bovis and Babesia bigemina as well as SBP3 of B. orientalis. The immunoreactivity was evaluated by incubating recombinant BoSBP4 (rBoSBP4) with the serum of B. orientalis-infected water buffalo. The native form of BoSBP4 was identified by incubating lysate of B. orientalis-infected water buffalo erythrocytes with the anti-rBoSBP4 mouse serum. The cellular localization of BoSBP4 was determined by indirect immunofluorescence assay.ResultsThe full length of the BoSBP4 gene was estimated to be 945 bp without introns, encoding a 314 aa polypeptide with a predicted molecular weight of 37 kDa. The truncated recombinant protein was expressed from 70 to 945 bp as a GST fusion protein with a practical molecular weight of 70 kDa. BoSBP4 shared a 40% and 30% identity with B. bovis and B. bigemina, respectively. Furthermore, it was 31% identical to SBP3 of B. orientalis. BoSBP4 was identified in the lysate of B. orientalis-infected water buffalo erythrocytes with a molecular weight of 37 kDa, corresponding to the expected molecular mass of BoSBP4. The result of rBoSBP4 with positive serum revealed that BoSBP4 can elicit an immune response to B. orientalis-infected water buffalo. The cellular localization of BoSBP4 was detected to be adjacent to the merozoite nucleus in the intracellular phase, followed by the diffusion of the fluorescence of BoSBP4 into the cytoplasm of B. orientalis-infected erythrocytes as puncta-like specks and a gradual increase of the fluorescence.ConclusionsIn this study, SBP4 in B. orientalis was characterized for the first time. It may play a key role in interaction with the host cell by being secreted into the cytoplasm of the B. orientalis-infected erythrocytes to facilitate parasite growth and reproduction.

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“…To generate a parasite line expressing PfRipr-HA, full length pfripr was amplified by PCR with the primers indicated in Table S2 (Ripr-tran F and R). The DNA fragments were digested with SpeI and PstI, and ligated into pD3HA plasmids with a Pfef1-alpha promoter for episomal expression 52,53 . For transfection, parasitized erythrocytes were resuspended in 200 µl of cytomix (120 mM KCl, 0.15 mM CaCl 2 , 2 mM EGTA, 5 mM MgCl 2 , 10 mM K 2 HPO 4 /KH 2 PO 4 , 25 mM HEPES, pH 7.4) containing 100 µg of plasmid DNA.…”

Section: Scientific Reports |mentioning

confidence: 99%

Antibodies against a short region of PfRipr inhibit Plasmodium falciparum merozoite invasion and PfRipr interaction with Rh5 and SEMA7A

Nagaoka

Kanoi

Ntege

et al. 2020

Sci Rep

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Plasmodium falciparum merozoite invasion into erythrocytes is an essential step of the blood-stage cycle, survival of parasites, and malaria pathogenesis. P. falciparum merozoite Rh5 interacting protein (PfRipr) forms a complex with Rh5 and CyRPA in sequential molecular events leading to erythrocyte invasion. Recently we described pfRipr as a conserved protein that induces strain-transcending growth inhibitory antibodies in in vitro assays. However, being a large and complex protein of 1086 amino acids (aa) with 87 cysteine residues, PfRipr is difficult to express in conventional expression systems towards vaccine development. in this study we sought to identify the most potent region of pfRipr that could be developed to overcome difficulties related to protein expression, as well as to elucidate the invasion inhibitory mechanism of anti-pfRipr antibodies. Using the wheat germ cell-free system, ecto-pfRipr and truncates of approximately 200 aa were expressed as soluble proteins. We demonstrate that antibodies against PfRipr truncate 5 (PfRipr_5: C 720 -D 934 ), a region within the pfRipr c-terminal eGflike domains, potently inhibit merozoite invasion. furthermore, the antibodies strongly block pfRipr/ Rh5 interaction, as well as that between PfRipr and its erythrocyte-surface receptor, SEMA7A. Taken together, PfRipr_5 is a potential candidate for further development as a blood-stage malaria vaccine.Plasmodium falciparum malaria remains a serious challenge to global health. In 2016, more than 3 billion people were reportedly at risk of infection, with an estimated 200 million cases and more than 400,000 deaths, primarily in young children living in sub-Saharan Africa 1 . Development of a malaria vaccine of high efficacy is considered a critical global agenda towards the achievement of malaria control and elimination. However, this progress has been greatly hampered by antigen polymorphism and low efficacy among target antigens 2,3 . Relatively conserved antigens that induce broadly cross-reactive antibodies and cell-mediated immune responses may provide long lasting and more efficacious protection 4-8 . It is also suggested that the next generation vaccines should incorporate multi-stage and/or multivalent targets aimed at inducing both humoral and cellular immunity 9 .The process of merozoite invasion of erythrocytes, that marks the beginning of the blood stage cycle of P. falciparum infections, takes less than 2 min and is characterized by dynamic molecular and cellular events 10-12 . Upon egress from the infected erythrocyte the merozoite is exposed to low potassium levels which trigger intracellular calcium release. The release activates secretion of adhesins and invasins, localized in the micronemes, onto the parasite surface [13][14][15] . Following the initial recognition of the host erythrocyte the parasite orients itself so that its apical end is directly facing the target erythrocyte membrane. The rhoptries are subsequently triggered to release invasion ligands that interact with erythrocyte receptors 16...

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PV1, a novel Plasmodium falciparum merozoite dense granule protein, interacts with exported protein in infected erythrocytes

Cited by 32 publications

References 37 publications

A lipocalin mediates unidirectional heme biomineralization in malaria parasites

A lipocalin mediates unidirectional heme biomineralization in malaria parasites

Characterization of a novel secretory spherical body protein in Babesia orientalis and Babesia orientalis-infected erythrocytes

Antibodies against a short region of PfRipr inhibit Plasmodium falciparum merozoite invasion and PfRipr interaction with Rh5 and SEMA7A

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