title: The subcellular location of ovalbumin in Plasmodium berghei blood stages influences the magnitude of T-cell responses.
abstract: Model antigens are frequently introduced into pathogens to study determinants that influence T-cell responses to infections. To address whether an antigen's subcellular location influences the nature and magnitude of antigen-specific T-cell responses, we generated Plasmodium berghei parasites expressing the model antigen ovalbumin (OVA) either in the parasite cytoplasm or on the parasitophorous vacuole membrane (PVM). For cytosolic expression, OVA alone or conjugated to mCherry was expressed from a strong constitutive promoter (OVAhsp70 or OVA::mCherryhsp70); for PVM expression, OVA was fused to HEP17/EXP1 (OVA::Hep17hep17). Unexpectedly, OVA expression in OVAhsp70 parasites was very low, but when OVA was fused to mCherry (OVA::mCherryhsp70), it was highly expressed. OVA expression in OVA::Hep17hep17 parasites was strong but significantly less than that in OVA::mCherryhsp70 parasites. These transgenic parasites were used to examine the effects of antigen subcellular location and expression level on the development of T-cell responses during blood-stage infections. While all OVA-expressing parasites induced activation and proliferation of OVA-specific CD8(+) T cells (OT-I) and CD4(+) T cells (OT-II), the level of activation varied: OVA::Hep17hep17 parasites induced significantly stronger splenic and intracerebral OT-I and OT-II responses than those of OVA::mCherryhsp70 parasites, but OVA::mCherryhsp70 parasites promoted stronger OT-I and OT-II responses than those of OVAhsp70 parasites. Despite lower OVA expression levels, OVA::Hep17hep17 parasites induced stronger T-cell responses than those of OVA::mCherryhsp70 parasites. These results indicate that unconjugated cytosolic OVA is not stably expressed in Plasmodium parasites and, importantly, that its cellular location and expression level influence both the induction and magnitude of parasite-specific T-cell responses. These parasites represent useful tools for studying the development and function of antigen-specific T-cell responses during malaria infection.
authors.affiliation: Leiden Malaria Research Group, Parasitology, Leiden University Medical Centre, Leiden, The Netherlands.
authors.forename: Jing-Wen
authors.initials: JW
authors.lastname: Lin
authors.affiliation: Faculty of Life Sciences, University of Manchester, Manchester, United Kingdom.
authors.forename: Tovah N
authors.initials: TN
authors.lastname: Shaw
authors.affiliation: Leiden Malaria Research Group, Parasitology, Leiden University Medical Centre, Leiden, The Netherlands.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Annoura
authors.affiliation: Leiden Malaria Research Group, Parasitology, Leiden University Medical Centre, Leiden, The Netherlands.
authors.forename: Aurélie
authors.initials: A
authors.lastname: Fougère
authors.affiliation: Leiden Malaria Research Group, Parasitology, Leiden University Medical Centre, Leiden, The Netherlands.
authors.forename: Pascale
authors.initials: P
authors.lastname: Bouchier
authors.affiliation: Leiden Malaria Research Group, Parasitology, Leiden University Medical Centre, Leiden, The Netherlands.
authors.forename: Séverine
authors.initials: S
authors.lastname: Chevalley-Maurel
authors.affiliation: Leiden Malaria Research Group, Parasitology, Leiden University Medical Centre, Leiden, The Netherlands.
authors.forename: Hans
authors.initials: H
authors.lastname: Kroeze
authors.affiliation: Leiden Malaria Research Group, Parasitology, Leiden University Medical Centre, Leiden, The Netherlands.
authors.forename: Blandine
authors.initials: B
authors.lastname: Franke-Fayard
authors.affiliation: Leiden Malaria Research Group, Parasitology, Leiden University Medical Centre, Leiden, The Netherlands.
authors.forename: Chris J
authors.initials: CJ
authors.lastname: Janse
authors.affiliation: Faculty of Life Sciences, University of Manchester, Manchester, United Kingdom.
authors.forename: Kevin N
authors.initials: KN
authors.lastname: Couper
authors.affiliation: Leiden Malaria Research Group, Parasitology, Leiden University Medical Centre, Leiden, The Netherlands S.M.Khan@lumc.nl.
authors.forename: Shahid M
authors.initials: SM
authors.lastname: Khan
mesh_terms: D000818:Animals; D005260:Female; D005786:Gene Expression Regulation / Q000502:physiology*; D008288:Malaria / Q000097:blood / Q000469:parasitology*; D051379:Mice; D030781:Organisms, Genetically Modified; D010047:Ovalbumin / Q000235:genetics / Q000378:metabolism*; D010962:Plasmodium berghei / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology*; D013154:Spleen / Q000166:cytology; D013601:T-Lymphocytes / Q000502:physiology
chemical_list: D010047:Ovalbumin