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Abstract

Stage-specific sterile protection against malaria can be achieved in rodents and human volunteers by administration of Plasmodium parasites that attenuate during liver-stage development by either radiation (RAS) or genetic modification (GAP). In this study we focussed on the identification and characterisation of potential antigens involved in GAP-mediated immunity. We identified within this study novel potential antigens involved in GAP-mediated immunity. We compared the transcript repertoire expressed of Plasmodium liver-stage parasites from GAP and RAS versus wild-type in an attempt to detect antigens that are uniquely or differentially expressed in GAP. One of the most abundant transcript up-regulated in RAS and GAP coded for a P. berghei C2-containing protein (C2CP). C2 domains are generally involved in Ca2+-sensing and -signalling and characteristic domains of the ferlin protein family. Database search revealed beside the Pb C2CP one annotated Pb ferlin (FER) and a ferlin-like protein (FLP). The cellular functions of Plasmodial ferlins, however, are unknown. We therefore also aimed at the cellbiological characterisation of the identified protein family. Targeted gene disruption experiments revealed an essential role of Pb C2CP and Pb FER during blood-stage development. The main objective of this study, however, focussed on the investigation of the antigenic potential of this novel pre-erythrocytic antigen. Interestingly epitope prediction identified several high-binding Pb C2CP-specific H2b-restricted CD8+ T-cell epitopes. Indeed, we could show that Pb C2CP-specific cells are recognised and lysed in immunised C57BL/6 mice. Moreover, we were able to specifically restimulate T cells in the livers of immunised animals with the Pb C2CP-derived peptide T9L. Altogether, both the dissection of novel antigens from the protection-inducing attenuated parasites and the immunogenic capacity as shown by a selected critical target antigen Pb C2CP for the first time allows specific restimulation of T cells in the experimental GAP vaccination model. Therefore this work clears the way for studying effector mechanisms underlying GAP-induced immunity. This work is of fundamental importance to understanding the mechanisms of pre-erythrocytic immunity to malaria and may further pave the way for the composition of an anti-infective multi-component subunit malarial vaccine.