Irgb6 has a role in the cell-autonomous response against T. gondii and involves ubiquitination and breakdown of vacuoles via binding of a specific phospholipid on the parasitophorous vacuole membrane.
A transmission-blocking vaccine (TBV) against Plasmodium falciparum is likely to be a valuable tool in a malaria eradication program. Pfs230 is one of the major TBV candidates, and multiple Pfs230-based vaccines induced antibodies, which prevented oocyst formation in mosquitoes as determined by a standard membrane-feeding assay (SMFA). Pfs230 is a >300 kDa protein consisting of 14 cysteine motif (CM) domains, and the size and cysteine-rich nature of the molecule have hampered its production as an intact protein. Except for one early study with maltose-binding protein fusion Pfs230 constructs expressed in Esherichia coli, all other studies have focused on only the first four CM domains in the Pfs230 molecule. To identify all possible TBV candidate domains, we systematically produced either single-CM-domain (a total of 14), 2-CM-domain (7), or 4-CM-domain (6) recombinant protein fragments using a eukaryotic wheat germ cell-free expression system (WGCFS). In addition, two more constructs which covered previously published regions, and an N-terminal prodomain construct spanning the natural cleavage site of Pfs230 were produced. Antisera against each fragment were generated in mice and we evaluated the reactivity to native Pfs230 protein by Western blots and immunofluorescence assay (IFA), and functionality by SMFA. All 30 WGCFS-produced Pfs230 constructs were immunogenic in mice. Approximately half of the mouse antibodies specifically recognized native Pfs230 by Western blots with variable band intensities. Among them, seven antibodies showed higher reactivities against native Pfs230 determined by IFA. Interestingly, antibodies against all protein fragments containing CM domain 1 displayed strong inhibitions in SMFA, while antibodies generated using constructs without CM domain 1 showed no inhibition. The results strongly support the concept that future Pfs230-based vaccine development should focus on the Pfs230 CM domain 1.
Upon invasion, Plasmodium falciparum exports hundreds of proteins across its surrounding parasitophorous vacuole membrane (PVM) to remodel the infected erythrocyte. Although this phenomenon is crucial for the parasite growth and virulence, elucidation of precise steps in the export pathway is still required. A translocon protein complex, PTEX, is the only known pathway that mediates passage of exported proteins across the PVM. P. falciparum Parasitophorous Vacuolar protein 1 (PfPV1), a previously reported parasitophorous vacuole (PV) protein, is considered essential for parasite growth. In this study, we characterized PfPV1 as a novel merozoite dense granule protein. Structured illumination microscopy (SIM) analyses demonstrated that PfPV1 partially co-localized with EXP2, suggesting the protein could be a PTEX accessory molecule. Furthermore, PfPV1 and exported protein PTP5 co-immunoprecipitated with anti-PfPV1 antibody. Surface plasmon resonance (SPR) confirmed the proteins’ direct interaction. Additionally, we identified a PfPV1 High-affinity Region (PHR) at the C-terminal side of PTP5 where PfPV1 dominantly bound. SIM analysis demonstrated an export arrest of PTP5ΔPHR, a PTP5 mutant lacking PHR, suggesting PHR is essential for PTP5 export to the infected erythrocyte cytosol. The overall results suggest that PfPV1, a novel dense granule protein, plays an important role in protein export at PV.
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...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply.
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.