Zinc metabolism and transcriptome and metabolome profiling in Plasmodium falciparum: The effects of traditional antimalarials and metal chelators.

摘要

Zinc and other transition metals are essential for cellular chemistry in all organisms. Part of the work described in this thesis focuses on three aspects of zinc metabolism in the malaria parasite, Plasmodium falciparum.; The first aspect is to study zinc transport by isolated malaria parasites and parasites within the infected erythrocyte. We find that despite living within an environment containing rich sources of zinc, the malaria parasite still needs to obtain extracellular zinc. Zinc transport was time, temperature, concentration and pH dependent and also exhibited saturable kinetics. In addition, labile pools of zinc were present within parasites but not within erythrocytes suggesting zinc transport processes may provide bioavailable zinc to the parasite. Also, we show that parasites are susceptible to both zinc chelation and excess zinc.; Secondly, the expression and localization of a zinc transporter was also examined. PfZnT, by sequence and structural homology belongs to the Cation Diffusion Family (CDF) of zinc transporters involved in cellular zinc detoxification. The expression of PfZnT was studied in parasites as well as in a heterologous mammalian system. We find that PfZnT partially colocalizes with a trans-golgi network marker to a peri-nuclear region in COS cells. We show that the intracellular soluble domain of PfZnT containing conserved histidine residues binds zinc, copper, cobalt and zinc in vitro.; Another area of zinc metabolism studied is zinc protoporphyrin IX (ZnPPIX) inhibition of neutral lipid mediated heme crystallization. Inhibition was temperature and pH dependent and unlike the quinoline antimalarials, irreversible.; Finally, the effects of metal chelators and antimalarials on the transcriptome and metabolome of the parasite was studied. We discovered that the transcriptome of P. falciparum is rigid and unresponsive to drug perturbations. Preliminary analysis on the metabolome indicate that metabolite fluxes change with drug treatment and can be measured in isolated parasites or in infected erythrocytes.; Taken together, the studies presented in this thesis work, has lead to a better understanding of zinc metabolism in the malaria parasite. Preliminary work on metabolic profiling in P. falciparum, presented here, has laid the foundation for more rigorous analysis of metabolite fluxes in response to drug.