Immunoscreening of Plasmodium falciparum proteins expressed in a wheat germ cell-free system reveals a novel malaria vaccine candidate

Morita, Masayuki; Takashima, Eizo; Ito, Daisuke; Miura, Kazutoyo; Thongkukiatkul, Amporn; Diouf, Ababacar; Fairhurst, Rick M.; Diakité, Mahamadou; Long, Carole A.; Torii, Motomi; Tsuboi, Takafumi

doi:10.1038/srep46086

Cited by 47 publications

(39 citation statements)

References 39 publications

(53 reference statements)

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…We chose differentially expressed genes encoding proteins containing predicted transmembrane domains or signal peptides, thereby likely to enter the parasite secretory pathway, excluding genes previously characterised as variably expressed or members of multi-gene families (S2 Table). Eight genes were selected for assay (S2 Fig), two of which encode known antigens (merozoite-associated tryptophan-rich antigen [34] liver-stage antigen-3 [35]). These gene transcripts were quantified by RT-qPCR in 58 RNA preparations from the laboratory clones and clinical isolates that had been analysed by RNA-seq.…”

Section: Resultsmentioning

confidence: 99%

“…In all, eight genes were identified for further characterisation (Table 3), which included three genes that were also differentially expressed among laboratory-adapted isolate comparisons (PF3D7_0423900, PF3D7_1252900 and PF3D7_1476500). Putative merozoite surface proteins encoded by PF3D7_0102700 (merozoite-associated tryptophan-rich antigen, MaTrA [48]) and PF3D7_0220000 (liver-stage antigen-3, LSA-3 [49]), were also among the subset of genes identified for further characterisation.…”

Section: Resultsmentioning

confidence: 99%

“…Focusing on genes for which variable expression has not been previously studied, we selected a panel of eight highly differentially expressed genes that encode secreted proteins. Of these, the merozoite-associated tryptophan-rich antigen and liver stage antigen 3 have been previously identified as merozoite proteins [34,35], and another gene (PF3D7_0423900) is adjacent to loci encoding the cysteine-rich protective antigen (CyRPA) and reticulocyte binding homologue 5 (Rh5) which are targets of antibodies that inhibit merozoite invasion. We extensively validated the RNA-seq data by RT-qPCR for these genes, and extended quantitation to additional ex vivo clinical isolate samples.…”

Section: Discussionmentioning

confidence: 99%

“…In order to focus on transcriptional variation as a mechanism of immune evasion, we identified a panel of eight highly differentially expressed genes that encode secreted proteins, since these may be targets of immunity. In particular, PF3D7_0102700 (merozoite-associated tryptophan-rich antigen), PF3D7_0220000 (liver stage antigen 3) have both been identified as merozoite proteins [48, 49]. Another gene identified in out study, PF3D7_0423900, sits near the genes encoding cysteine-rich protective antigen (CyRPA) and reticulocyte binding homolog 5 (PfRh5) on chromosome 4.…”

Section: Discussionmentioning

confidence: 99%

See 3 more Smart Citations

Plasmodium falciparum schizont stage transcriptome variation among clinical isolates and laboratory-adapted clones

Díaz-Ingelmo

et al. 2018

Preprint

View full text Add to dashboard Cite

Malaria parasite genes can exhibit variation in sequence or transcriptional activity. A wealth of information is available on sequence polymorphism in clinical malaria isolates, but there are few quantitative whole-transcriptome studies of natural variation at specific developmental stages. It is challenging to obtain adequately precise and well-replicated transcriptome measurements in order to account for technical and biological variation among preparations, which can otherwise introduce noise or bias in analyses. We address the issue by obtaining of RNA-seq profiles of multiple independently cultured replicates of mature schizont-stage malaria parasites from a panel of clinical isolates and laboratory-adapted lines. With a goal of robustly identifying variably expressed genes, we show that increasing the biological sample replication improves the true positive discovery rate, and that six independent replicates of each isolate is significantly superior to lower numbers. Focusing on genes that are more highly expressed on average improves the discovery rate when fewer biological replicates are available. We identify genes encoding transcription factors and proteins implicated in gametocytogenesis that differ in expression between cultured clinical and laboratory adapted lines.We confirm the variable expression of known merozoite invasion ligands, and identify previously uncharacterised genes as highly differentially expressed among isolates. RT-qPCR assays confirm the variation, and extend quantitation of expression of these genes to a wider panel of ex vivo clinical isolate samples. This highlights new candidates for investigation as potential markers of alternative developmental pathways or targets of immunity. Author summaryWe analyse the transcriptomes of mature Plasmodium falciparum schizonts using RNA-sequencing.We use large numbers of replicates per sample to minimise the impact of inter-replicate biological variation on observed patterns of differential expression. We identify genes that are differentially expressed between isolates. We extensively validate our findings for novel putative targets of immunity. For a panel of ex vivo clinical isolates, we show that expression levels of these candidate genes in fall within the ranges observed for the cultured isolates. These genes constitute novel targets for characterisation for merozoite-stage intervention.

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 2 more Smart Citations

Plasmodium falciparum schizont stage transcriptome variation among clinical isolates and laboratory-adapted clones

Díaz-Ingelmo

et al. 2018

Preprint

View full text Add to dashboard Cite

show abstract

“…It presents flexibility in experimental conditions, requires no codon optimization, allows post-translational modifications and can generate milligram amounts of high-quality proteins. This technology has been successfully applied to find novel malaria vaccine candidates [37][38][39][40][41].…”

Section: The Wheat Germ Cell-free Systemmentioning

confidence: 99%