Genome-Scale Identification of Essential Metabolic Processes for Targeting the Plasmodium Liver Stage

Rebecca R Stanway,Vassily Hatzimanikatis,Blandine Franke-Fayard,Reto Caldelari,Theo Sanderson,Anush Chiappino-Pepe,Vikash Pandey,Frank Schwach,Ellen Bushell,Séverine Chevalley,Magali Roques,Oliver Billker,Chris J Janse,Tom Metcalf,Murielle Golomingi,Julian C Rayner,Colin Herd,Jai Ramesar,Mary Nyonda,Dominique Soldati‐Favre ,Paul‐Christian Burda ,Volker Heussler

doi:10.1016/j.cell.2019.10.030

Abstract

SummaryPlasmodium gene functions in mosquito and liver stages remain poorly characterized due to limitations in the throughput of phenotyping at these stages. To fill this gap, we followed more than 1,300 barcoded P. berghei mutants through the life cycle. We discover 461 genes required for efficient parasite transmission to mosquitoes through the liver stage and back into the bloodstream of mice. We analyze the screen in the context of genomic, transcriptomic, and metabolomic data by building a thermodynamic model of P. berghei liver-stage metabolism, which shows a major reprogramming of parasite metabolism to achieve rapid growth in the liver. We identify seven metabolic subsystems that become essential at the liver stages compared with asexual blood stages: type II fatty acid synthesis and elongation (FAE), tricarboxylic acid, amino sugar, heme, lipoate, and shikimate metabolism. Selected predictions from the model are individually validated in single mutants to provide future targets for drug development.

Highlights

Malaria, caused by parasites of the genus Plasmodium, remains a disease of major significance to global public health
Validating Barseq for Analysis of Gene Knockout Mutants in Non-erythrocytic Stages Only the asexual blood stages of Plasmodium parasites can be propagated continuously to drug-select for knockout mutants, meaning that only genes that are not required for blood-stage development can be investigated at later stages of the cycle using barseq
Using knockout vectors targeting 15 genes with known functions at the liver stage and 19 control and test genes, a pool of mutant parasites was generated by transfection and used to infect three mice

Summary

Introduction

Malaria, caused by parasites of the genus Plasmodium, remains a disease of major significance to global public health. Over the course of two to five days, depending on the Plasmodium species, the parasite increases dramatically in size and eventually gives rise to thousands of daughter merozoites With this immense and rapid expansion, parasites need to be highly metabolically active, despite their dependence on the host cell for nutrient acquisition. The merozoites are released into the bloodstream, where they invade red blood cells and undergo repeated rounds of asexual replication, each round culminating in the release of further invasive merozoites It is the blood phase of the parasite life cycle that leads to the symptoms of malaria and, in the case of Plasmodium falciparum, can cause fatal disease (reviewed in Cowman et al, 2016). Motile sporozoites are liberated into the haemocoel of the mosquito and eventually accumulate in the salivary glands, where they await injection into a new mammalian host

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