Abstract
The vast majority of malaria mortality is attributed to one parasite species: Plasmodium falciparum. Asexual replication of the parasite within the red blood cell is responsible for the pathology of the disease. In Plasmodium, the endoplasmic reticulum (ER) is a central hub for protein folding and trafficking as well as stress response pathways. In this study, we tested the role of an uncharacterised ER protein, PfGRP170, in regulating these key functions by generating conditional mutants. Our data show that PfGRP170 localises to the ER and is essential for asexual growth, specifically required for proper development of schizonts. PfGRP170 is essential for surviving heat shock, suggesting a critical role in cellular stress response. The data demonstrate that PfGRP170 interacts with the Plasmodium orthologue of the ER chaperone, BiP. Finally, we found that loss of PfGRP170 function leads to the activation of the Plasmodium eIF2α kinase, PK4, suggesting a specific role for this protein in this parasite stress response pathway.
Highlights
Malaria is a deadly parasitic disease that causes over 212 million cases and nearly 430,000 deaths each year, primarily in children under the age of five[1]
Unlike Plasmodium falciparum, Yeast null for GRP170 are viable due to the upregulation of Sil[1], another nucleotide exchange factor, that usually plays a role in the IRE1 stress response pathway[51]
The Plasmodium genome does not encode Sil[1] and IRE1, which aligns with the observed essentiality of PfGRP170 during the blood stages
Summary
Malaria is a deadly parasitic disease that causes over 212 million cases and nearly 430,000 deaths each year, primarily in children under the age of five[1]. Upon removal TMP, the PfGRP170-DDD parasites die 38-44 hours post invasion (Figure 2E) and it is likely that the essential function of PfGRP170 is linked to proteins expressed during these late stages of the asexual life-cycle. This approach has been used successfully in Plasmodium to demonstrate interaction of exported proteins[50] We performed this assay using anti-GFP and anti-BiP antibodies and observed a positive signal at all life cycle stages (Figure 5C). As a negative control we probed with an antibody against the ER localized protease PMV and despite the co-localization of these two proteins in the ER, we did not see a positive PLA signal, suggesting distinct sub-organellar localizations (Figure 5D) Together, these results demonstrate that PfGRP170 and PfBiP interact during all stages of the asexual life cycle of P. falciparum. This experiment showed no changes in levels of EIF2-α regardless of the presence of TMP or the PK4 inhibitor. 304 305
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