Phenotypic screens reveal Plasmodium falciparum genetic factors associated with infection of sickle-trait cells

Abstract

Background: Malaria causes over 200 million cases and more than half a million deaths annually. In many African regions, hemoglobinopathies, such as sickle cell trait (HbAS), confer partial protection against severe P. falciparum malaria. HbAS significantly reduces the risk of severe, life-threatening malaria by over 90 %. This study aims to describe a new analysis for the piggyBac transposon-based mutagenesis phenotypic screen to identify genes that influence the mechanisms behind this protection and tolerance of P. falciparum to the HbAS intracellular microenvironment, providing insights into potential new targets for malaria intervention and the evolutionary relationship between host and parasite. Methods: We optimized and successfully employed a phenotypic screen using a piggyBac transposon-mutant library of P. falciparum to identify genetic factors essential for parasite survival in HbAS RBCs. Parasites were cultured in vitro in HbAS and control HbAA RBCs. Parasite growth was assessed via Quantitative Insertion Site Sequencing (QIseq) to determine sensitivity of each mutant in response to the conditions of HbAS RBCs identifying sensitive and tolerant mutants. Finally, a pairwise comparison was performed between HbAS and previously published piggyBac screens to infer potential links between HbAS infection and parasite responses to heat-shock, antimalarial drugs and oxidative stress. Results: Our findings revealed that P. falciparum mutants sensitive to HbAS growth are associated with genes involved in signaling pathways, exported proteins, and host-interaction genes. These genetic factors overlap with those involved in the parasite's response to oxidative stress and antimalarial drug sensitivity, such as artemisinin derivates and proteasome inhibitor. Conclusions: Our study identifies genetic factors influencing P. falciparum infection in HbAS RBCs, shedding light on how HbAS may counteract with the parasite, suggesting a connection between sickle-trait infections and other stress responses, such as heat-shock, artemisinin and oxidative stress.