Investigating the link between nutrient sensing and gametocyte formation in the malaria parasite Plasmodium falciparum

Malaria is an infectious disease and causes over 600’000 deaths annually. To ensure host-to-host transmission, malaria parasites rely on specialized stages, called gametocytes. The formation of these sexual precursor cells is initiated by commitment of asexual blood stage parasites to the sexual differentiation pathway. Plasmodium falciparum, the most virulent of six parasite species infecting humans, employs nutrient sensing to adapt the rate of sexual commitment. The parasite monitors the levels of the host-derived lipid lysophosphatidylcholine (lysoPC) to control the epigenetically regu-lated expression of the transcription factor AP2-G, the master regulator of sexual commitment. The molecular mechanisms linking lysoPC sensing to AP2-G expression, however, are poorly under-stood. In this PhD project, I aimed at identifying factors linking nutrient sensing and sexual commit-ment in P. falciparum. To this end, I investigated the effect of a set of chemical compounds and 12 selected genes on sexual commitment rates. In a first study, I assessed the effect of drug treatment on sexual commitment rates using a high content imaging-based assay to clarify the longstanding question of whether antimalarial drug treatment increases gametocyte formation. By screening a comprehensive set of pharmaceuticals using both drug sensitive and drug resistant parasite strains, we show that drug treatment indeed can increase sexual commitment rates, but only rarely results in a net increase in gametocyte production. Our results suggest that general stress, rather than drug-specific activities, promotes sexual commitment. This finding was crucial to interpret the results of a second study, in which we screened compounds of inhibitor libraries to identify molecules influenc-ing sexual commitment. We then employed mass spectrometry-based cellular thermal shift assays (MS-CETSA) to identify the cellular target of selected hit compounds. Thereby, we identified high-confidence candidate targets and assessed their role during sexual commitment using reverse genet-ics approaches. We found indication for an involvement of cyclic adenosine monophosphate (cAMP)-dependent signaling and protein kinase A (PKA) in regulating commitment to gametocyte formation. Based on reverse genetics experiments and chemical perturbation assays, we hypothesize that PKA acts as a repressor of AP2-G expression under lysoPC-free conditions and thus links nutri-ent sensing to the epigenetic regulation of AP2-G. However, more experiments are required to con-firm this hypothesis. In a third project, we assessed the function the FK506-binding protein 35 (FKBP35) – another putative target of a commitment-inducing compound – by combining reverse genetics with transcriptional and proteomic profiling. While we did not find evidence for a role in sexual commitment, we show that limiting FKBP35 levels are lethal to P. falciparum and result in a delayed death-like phenotype that is characterized by defective ribosome homeostasis and stalled protein synthesis. Our data furthermore suggest that the antiplasmodial activity of the well-characterized FKBP inhibitor FK506 is independent of FKBP35, which has implications for the de-velopment of FK506-derived molecules as antimicrobial drugs. In summary, this thesis offers a test-able model to further investigate the link between nutrient sensing and sexual commitment and pro-vides new insights into the function of FKBP35.