Gene Disruption of Plasmodium falciparum p52 Results in Attenuation of Malaria Liver Stage Development in Cultured Primary Human Hepatocytes

Schaijk, Ben C. L. van; Janse, Chris J.; Gemert, Geert-Jan van; Dijk, Melissa R. van; Gego, Audrey; Franetich, Jean‐François; Vegte‐Bolmer, Marga van de; Yalaoui, Samir; Silvie, Olivier; Hoffman, Stephen L.; Waters, Andrew P.; Mazier, Dominique; Sauerwein, Robert W.; Khan, Shahid M.

doi:10.1371/journal.pone.0003549

Cited by 88 publications

(94 citation statements)

References 64 publications

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“…However, the knockouts showed clear defects in liver-stage development. Recently, in an independent study, disruption of the P52 gene in P. falciparum has been achieved by single-cross-over integration (32). The observed phenotype was in agreement with the p52 Ϫ phenotype presented here.…”

Section: Discussionsupporting

confidence: 88%

“…The observed phenotype was in agreement with the p52 Ϫ phenotype presented here. However, single-cross-over recombination insertions are prone to reversion by plasmid excision, and indeed, the study by van Schaijk et al (32) observed reverting parasites that had restored WT growth. Our strategy for gene deletions was to use double-cross-over recombination gene replacement, which makes it impossible for the parasite to genet- Fig.…”

Section: Discussionmentioning

confidence: 99%

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Preerythrocytic, live-attenuated Plasmodium falciparum vaccine candidates by design

VanBuskirk

O'Neill

Vega

et al. 2009

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

153

150

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Falciparum malaria is initiated when Anopheles mosquitoes transmit the Plasmodium sporozoite stage during a blood meal. Irradiated sporozoites confer sterile protection against subsequent malaria infection in animal models and humans. This level of protection is unmatched by current recombinant malaria vaccines. However, the live-attenuated vaccine approach faces formidable obstacles, including development of accurate, reproducible attenuation techniques. We tested whether Plasmodium falciparum could be attenuated at the early liver stage by genetic engineering. The P. falciparum genetically attenuated parasites (GAPs) harbor individual deletions or simultaneous deletions of the sporozoiteexpressed genes P52 and P36. Gene deletions were done by double-cross-over recombination to avoid genetic reversion of the knockout parasites. The gene deletions did not affect parasite replication throughout the erythrocytic cycle, gametocyte production, mosquito infections, and sporozoite production rates. However, the deletions caused parasite developmental arrest during hepatocyte infection. The double-gene deletion line exhibited a more severe intrahepatocytic growth defect compared with the single-gene deletion lines, and it did not persist. This defect was assessed in an in vitro liver-stage growth assay and in a chimeric mouse model harboring human hepatocytes. The strong phenotype of the double knockout GAP justifies its human testing as a whole-organism vaccine candidate using the established sporozoite challenge model. GAPs might provide a safe and reproducible platform to develop an efficacious whole-cell malaria vaccine that prevents infection at the preerythrocytic stage.genetically attenuated parasites ͉ malaria vaccine ͉ P36 ͉ P52 ͉ sporozoite M alaria is a formidable global health problem, affecting 300 million to 500 million people worldwide annually (1). The resulting Ϸ1 million deaths per year are mainly caused by Plasmodium falciparum infections. Eradication of malaria will in large part depend on an effective vaccine that prevents infection by Plasmodium, but such a vaccine has remained elusive. The parasites' preerythrocytic stages, encompassing the mosquito-inoculated sporozoites and liver stages that develop from sporozoites after their invasion of hepatocytes, are attractive targets for antiinfection vaccines, because at this stage the number of infected host cells is low, and further transmission of the parasite is not yet possible. Occurrence of blood-stage infection after sporozoite challenge is completely preventable by immunization with radiation-attenuated sporozoites in mouse models of malaria (2). This was a landmark finding that set the standards for malaria preerythrocytic vaccine development. Radiation-attenuated sporozoites arrest in development during hepatocyte infection, but their safety and efficacy are dependent on a precise irradiation dose. Humans immunized with P. falciparum radiation-attenuated sporozoites have been effectively protected from subsequent challenge with homologous an...

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Section: Discussionsupporting

confidence: 88%

Section: Discussionmentioning

confidence: 99%

Preerythrocytic, live-attenuated Plasmodium falciparum vaccine candidates by design

VanBuskirk

O'Neill

Vega

et al. 2009

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

153

150

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“…Gliding assays and inhibition of sporozoite invasion assays were performed as described previously (38).…”

Section: Methodsmentioning

confidence: 99%

Vaccine-induced monoclonal antibodies targeting circumsporozoite protein prevent Plasmodium falciparum infection

Foquet¹,

Hermsen²,

Gemert³

et al. 2013

J. Clin. Invest.

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142

123

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“…The functions of these two proteins appear to be different because the deletion of either gene influences the ability of the parasite to efficiently infect hepatocytes, with no effect on their ability to invade salivary gland cells in the mosquito (22,23). In P. berghei there is also evidence that these genes may be required for invasion of hepatocytes and to promote normal development in the liver.…”

mentioning

confidence: 99%

Structure of thePlasmodium6-cysteine s48/45 domain

Arredondo

Cai²,

Takayama³

et al. 2012

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

107

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The s48/45 domain was first noted in Plasmodium proteins more than 15 y ago. Previously believed to be unique to Plasmodium , the s48/45 domain is present in other aconoidasidans. In Plasmodium , members of the s48/45 family of proteins are localized on the surface of the parasite in different stages, mostly by glycosylphosphatydylinositol-anchoring. Members such as P52 and P36 seem to play a role in invasion of hepatocytes, and Pfs230 and Pfs48/45 are involved in fertilization in the sexual stages and have been consistently studied as targets of transmission-blocking vaccines for years. In this report, we present the molecular structure for the s48/45 domain corresponding to the C-terminal domain of the blood-stage protein Pf12 from Plasmodium falciparum , obtained by NMR. Our results indicate that this domain is a β-sandwich formed by two sheets with a mixture of parallel and antiparallel strands. Of the six conserved cysteines, two pairs link the β-sheets by two disulfide bonds, and the third pair forms a bond outside the core. The structure of the s48/45 domain conforms well to the previously defined surface antigen 1 (SAG1)-related-sequence (SRS) fold observed in the SAG family of surface antigens found in Toxoplasma gondii . Despite extreme sequence divergence, remarkable spatial conservation of one of the disulfide bonds is observed, supporting the hypothesis that the domains have evolved from a common ancestor. Furthermore, a homologous domain is present in ephrins, raising the possibility that the precursor of the s48/45 and SRS domains emerged from an ancient transfer to Apicomplexa from metazoan hosts.

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Gene Disruption of Plasmodium falciparum p52 Results in Attenuation of Malaria Liver Stage Development in Cultured Primary Human Hepatocytes

Cited by 88 publications

References 64 publications

Preerythrocytic, live-attenuated Plasmodium falciparum vaccine candidates by design

Preerythrocytic, live-attenuated Plasmodium falciparum vaccine candidates by design

Vaccine-induced monoclonal antibodies targeting circumsporozoite protein prevent Plasmodium falciparum infection

Structure of thePlasmodium6-cysteine s48/45 domain

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