RALP1 Is a Rhoptry Neck Erythrocyte-Binding Protein of Plasmodium falciparum Merozoites and a Potential Blood-Stage Vaccine Candidate Antigen

ABSTRACTErythrocyte invasion by merozoites is an obligatory stage ofPlasmodiuminfection and is essential to disease progression. Proteins in the apical organelles of merozoites mediate the invasion of erythrocytes and are potential malaria vaccine candidates. Rhoptry-associated, leucine zipper-like protein 1 (RALP1) ofPlasmodium falciparumwas previously found to be specifically expressed in schizont stages and localized to the rhoptries of merozoites by immunofluorescence assay (IFA). Also, RALP1 has been refractory to gene knockout attempts, suggesting that it is essential for blood-stage parasite survival. These characteristics suggest that RALP1 can be a potential blood-stage vaccine candidate antigen, and here we assessed its potential in this regard. Antibodies were raised against recombinant RALP1 proteins synthesized by using the wheat germ cell-free system. Immunoelectron microscopy demonstrated for the first time that RALP1 is a rhoptry neck protein of merozoites. Moreover, our IFA data showed that RALP1 translocates from the rhoptry neck to the moving junction during merozoite invasion. Growth and invasion inhibition assays revealed that anti-RALP1 antibodies inhibit the invasion of erythrocytes by merozoites. The findings that RALP1 possesses an erythrocyte-binding epitope in the C-terminal region and that anti-RALP1 antibodies disrupt tight-junction formation, are evidence that RALP1 plays an important role during merozoite invasion of erythrocytes. In addition, human sera collected from areas in Thailand and Mali where malaria is endemic recognized this protein. Overall, our findings indicate that RALP1 is a rhoptry neck erythrocyte-binding protein and that it qualifies as a potential blood-stage vaccine candidate.

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Discovery of GAMA, a Plasmodium falciparum Merozoite Micronemal Protein, as a Novel Blood-Stage Vaccine Candidate Antigen

Infection and Immunity ◽

10.1128/iai.05412-11 ◽

2011 ◽

Vol 79 (11) ◽

pp. 4523-4532 ◽

Cited By ~ 45

Author(s):

Thangavelu U. Arumugam ◽

Satoru Takeo ◽

Tsutomu Yamasaki ◽

Amporn Thonkukiatkul ◽

Kazutoyo Miura ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Vaccine Candidate ◽

Gene Knockout ◽

Blood Stage ◽

Dependent Manner ◽

Free System ◽

Content Type ◽

Vaccine Candidate Antigen ◽

Erythrocyte Binding ◽

Candidate Antigen

ABSTRACTOne of the solutions for reducing the global mortality and morbidity due to malaria is multivalent vaccines comprising antigens of several life cycle stages of the malarial parasite. Hence, there is a need for supplementing the current set of malaria vaccine candidate antigens. Here, we aimed to characterize glycosylphosphatidylinositol (GPI)-anchored micronemal antigen (GAMA) encoded by the PF08_0008 gene inPlasmodium falciparum. Antibodies were raised against recombinant GAMA synthesized by using a wheat germ cell-free system. Immunoelectron microscopy demonstrated for the first time that GAMA is a microneme protein of the merozoite. Erythrocyte binding assays revealed that GAMA possesses an erythrocyte binding epitope in the C-terminal region and it binds a nonsialylated protein receptor on human erythrocytes. Growth inhibition assays revealed that anti-GAMA antibodies can inhibitP. falciparuminvasion in a dose-dependent manner and GAMA plays a role in the sialic acid (SA)-independent invasion pathway. Anti-GAMA antibodies in combination with anti-erythrocyte binding antigen 175 exhibited a significantly higher level of invasion inhibition, supporting the rationale that targeting of both SA-dependent and SA-independent ligands/pathways is better than targeting either of them alone. Human sera collected from areas of malaria endemicity in Mali and Thailand recognized GAMA. Since GAMA inP. falciparumis refractory to gene knockout attempts, it is essential to parasite invasion. Overall, our study indicates that GAMA is a novel blood-stage vaccine candidate antigen.

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PfRON3 is an erythrocyte-binding protein and a potential blood-stage vaccine candidate antigen

Malaria Journal ◽

10.1186/1475-2875-13-490 ◽

2014 ◽

Vol 13 (1) ◽

pp. 490 ◽

Cited By ~ 9

Author(s):

Xin Zhao ◽

Zhiguang Chang ◽

Zhiwei Tu ◽

Shengchao Yu ◽

Xiaoyan Wei ◽

...

Keyword(s):

Vaccine Candidate ◽

Binding Protein ◽

Blood Stage ◽

Vaccine Candidate Antigen ◽

Erythrocyte Binding ◽

Candidate Antigen

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Bacterially Expressed Full-Length Recombinant Plasmodium falciparum RH5 Protein Binds Erythrocytes and Elicits Potent Strain-Transcending Parasite-Neutralizing Antibodies

Infection and Immunity ◽

10.1128/iai.00970-13 ◽

2013 ◽

Vol 82 (1) ◽

pp. 152-164 ◽

Cited By ~ 54

Author(s):

K. Sony Reddy ◽

Alok K. Pandey ◽

Hina Singh ◽

Tajali Sahar ◽

Amlabu Emmanuel ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Neutralizing Antibodies ◽

Malaria Vaccine ◽

Expression System ◽

Full Length ◽

Blood Stage ◽

Parasite Protein ◽

Content Type ◽

Erythrocyte Binding ◽

Blood Stage Malaria

ABSTRACTPlasmodium falciparumreticulocyte binding-like homologous protein 5 (PfRH5) is an essential merozoite ligand that binds with its erythrocyte receptor, basigin. PfRH5 is an attractive malaria vaccine candidate, as it is expressed by a wide number ofP. falciparumstrains, cannot be genetically disrupted, and exhibits limited sequence polymorphisms. Viral vector-induced PfRH5 antibodies potently inhibited erythrocyte invasion. However, it has been a challenge to generate full-length recombinant PfRH5 in a bacterial-cell-based expression system. In this study, we have produced full-length recombinant PfRH5 inEscherichia colithat exhibits specific erythrocyte binding similar to that of the native PfRH5 parasite protein and also, importantly, elicits potent invasion-inhibitory antibodies against a number ofP. falciparumstrains. Antibasigin antibodies blocked the erythrocyte binding of both native and recombinant PfRH5, further confirming that they bind with basigin. We have thus successfully produced full-length PfRH5 as a functionally active erythrocyte binding recombinant protein with a conformational integrity that mimics that of the native parasite protein and elicits potent strain-transcending parasite-neutralizing antibodies.P. falciparumhas the capability to develop immune escape mechanisms, and thus, blood-stage malaria vaccines that target multiple antigens or pathways may prove to be highly efficacious. In this regard, antibody combinations targeting PfRH5 and other key merozoite antigens produced potent additive inhibition against multiple worldwideP. falciparumstrains. PfRH5 was immunogenic when immunized with other antigens, eliciting potent invasion-inhibitory antibody responses with no immune interference. Our results strongly support the development of PfRH5 as a component of a combination blood-stage malaria vaccine.

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