Structural and mechanistic insights into Plasmodium eIF2alpha dephosphorylation by UIS2 during the erythrocytic stage

UIS2 is a promising therapeutic target for malaria due to its essential role in the transmission and infectivity of Plasmodium to the host hepatocytes. In Plasmodium, UIS2 acts as a phosphatase toward phosphorylated eIF2alpha at Ser59, regulating translation initiation. However, its role during the erythrocytic stage, which causes clinical symptoms, remains unclear, and its protein structure for elucidating the dephosphorylation mechanism is unavailable. In this study, we analyzed the Plasmodium phenotype screening database and found that UIS2-deficient Plasmodium fails to proliferate during the erythrocytic stages. Single-cell transcriptomic data from the Malaria Cell Atlas revealed UIS2 expression during both hepatic and erythrocytic stages, with significant upregulation in trophozoites and late schizonts, suggesting UIS2s essential role in erythrocytic stage development. Structural analyses using AlphaFold modeling demonstrated that the UIS2 phosphatase domain (UIS2-PD) shares homology with human purple acid phosphatase, featuring conserved catalytic site residues. We observed that the N-terminal domain of UIS2 interacts with the S1 domain loop of a phospho-mimic mutant (Ser59Asp) eIF2alpha via electrostatic interactions. This protein interaction is distal to the phosphatase domain, indicating a unique substrate recruitment mechanism distinct from mammalian serine/threonine phosphatase PP1. Additionally, we identified a negatively charged cavity in UIS2-PD capable of binding the Ser59-containing loop in eIF2alpha for substrate recognition. Molecular docking studies showed that the phosphatase inhibitor salubrinal binds to this negatively charged cavity, potentially obstructing enzyme-substrate interactions. These structural insights into UIS2 and its interaction with eIF2alpha provide promising avenues for developing novel antimalarial drugs. Significance StatementMalaria poses a significant global health challenge, particularly with rising resistance to artemisinin therapies. This study reveals the critical role of UIS2 in Plasmodium proliferation during the symptomatic blood stage through phenotype screening and single-cell transcriptomics. We characterized the catalytic site within UIS2-PD and discovered a surface cavity for recognizing phosphorylated eIF2alpha by UIS2-PD, which can be inhibited by small molecules. Our findings identify UIS2 as a promising target for new antimalarial drugs, addressing both blood-stage parasites to alleviate clinical symptoms and liver-stage parasites for prophylactic treatment. This dual targeting strategy could prevent malaria onset post-exposure and reduce the overall parasite burden. The protein interaction between UIS2 and eIF2alpha is chemically targetable, facilitating the development of effective UIS2 inhibitors.