Translational repression of the CPW-WPC multigene family in the malaria parasite Plasmodium

The technical challenges of working with the sexual stages of the malaria parasite Plasmodium have hindered the characterization of putative parasite surface antigens in the quest for a successful malaria transmission-blocking vaccine. One such predicted uncharacterized extracellular protein, the CPW-WPC family of proteins, is conserved across Apicomplexa , and has been named for a characteristic domain that contains two conserved motifs CPxxW and WPC. We detected nine cpw-wpc transcripts in P. falciparum gametocytes and eight cpw-wpc transcripts in P. berghei gametocytes. With the exception of one cpw-wpc gene we observed no transcript expression in asexual stages. Using immune sera generated against a recombinant CPW-WPC protein, as well as a transgenic P. berghei parasite line expressing an epitope-tagged CPW-WPC protein, we were unable to detect protein expression in intraerythrocytic asexual and sexual stage parasites, suggesting translational control of some or all members of the family. RNA immunoprecipitation (RIP) followed by Reverse Transcriptase-PCR using transgenic DOZI::GFP and CITH::GFP-expressing P. berghei lines confirmed that all cpw-wpc transcripts were bound by both translational repressors in gametocytes, and thus likely translationally repressed until a later life cycle stage. In order to analyze translational repression in P. falciparum gametocytes the expression of luciferase reporters regulated by cpw-wpc and control regulatory regions was used as a proxy for CPW-WPC protein expression. This analysis demonstrated that cpw-wpc regulated transcripts were likely translationally silent in gametocytes but translationally activated following gametocyte activation.