How to starve a parasite: Manipulating CoA biosynthesis to control Plasmodium development in the mosquito

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Project Summary Malaria parasites require pantothenate (Pan) from both the insect and mammalian hosts to synthesize coenzyme A (CoA) and acetyl-CoA (AC). Further, mosquito-stage parasites cannot take up preformed CoA from the insect host, so they are entirely dependent on mosquito Pan availability. Thus, we hypothesize that reducing Pan stores in the mosquito by increasing Pan kinase (PanK) activity and, in turn, CoA biosynthesis will limit parasite survival in the mosquito, without impacting the availability of CoA/AC to the mosquito itself. PanK is the rate-limiting enzyme in the CoA biosynthesis pathway and a logical target for our approach. In this study we will focus on increasing PanK activity in the mosquito to convert Pan into CoA and starve the malaria parasite of this essential precursor. To accomplish this we will utilize PanK-targeted small molecules or pantazines and genetic manipulation of PanK in our study host Anopheles stephensi. We will screen pantazines from a library of compounds developed by our collaborators at St. Jude Children’s Hospital. In Aim 1, we will use a Go-No Go strategy for pantazine screening that culminates in testing the capacity of selected pantazines to reduce P. falciparum and Plasmodium yoelii infections in A. stephensi. Aim 2 will validate the bioactivity and specificity of candidate pantazines identified in the screen in Aim 1. The specificity of candidate pantazines to activate PanK will be assessed through RNAi or CRISPR/Cas9 knockdown of PanK, followed by a characterization of the impact on Pan, CoA, AC and parasite infection success. Concurrent with Aims 1 and 2, we will generate transgenic A. stephensi with increased PanK activity and determine the impact on Pan levels and parasite survival in Aim 3. The generation of transgenic mosquitoes with increased midgut PanK expression will contribute to our assessment of PanK-dependent depletion of Pan stores on parasite infection as well as other aspects of mosquito biology related to vectorial capacity. Specifically, we will define the effects of mosquito PanK activation, via both pantazine treatment and molecular manipulations, on A. stephensi lifespan, stress responses, metabolism and reproduction. These studies will reveal important new insights into nutrient-driven mosquito-parasite interactions that drive parasite infection success and they will support future efforts to optimize pantazines and novel transgenic lines as distinct strategies for mosquito-targeted malaria control.