Assessment of biological role and insight into druggability of the Plasmodium falciparum protease plasmepsin V

Abstract

Upon infection of a red blood cell (RBC), the malaria parasite Plasmodium falciparum drastically remodels its host by exporting hundreds of proteins into the RBC cytosol. This program of protein export is essential for parasite survival, hence there is interest in export-related proteins as potential drug targets. One proposed target is plasmepsin V (PMV), an aspartic protease that cleaves export-destined proteins in the parasite ER at a motif called the Plasmodium export element (PEXEL). This cleavage is essential for effector export across the vacuolar membrane. Despite long-standing interest in PMV, functional studies have been hindered by the failure of current technologies to produce a regulatable lethal depletion of PMV. To overcome this technical barrier, we designed a facile system for stringent post-transcriptional regulation, allowing a tightly controlled, tunable knockdown of PMV. Under maximal knockdown conditions, parasite growth was arrested, validating PMV as essential for parasite survival in RBCs. We found that PMV levels had to be dramatically depleted to affect parasite growth, suggesting that the parasite maintains this enzyme in substantial excess. This has important implications for antimalarial development. Additionally, we found that PMV-depleted parasites arrest immediately after invasion of the host cell, suggesting that PMV has an unappreciated role in early development that is distinct from its previously reported role in protein export in later-stage parasites.\n\nImportanceMalaria is endemic to large swaths of the developing world, causing nearly 500,000 deaths each year. While infection can be treated with antimalarial drugs, resistance continues to emerge to frontline antimalarials, spurring calls for new drugs and targets to feed the drug development pipeline. One proposed target is the aspartic protease plasmepsin V (PMV) that processes exported proteins, enabling the export program that remodels the host cell. This work uses facile genetic tools to produce lethal depletion of PMV, validating it as a drug target and showing that PMV is made in substantial excess in blood-stage parasites. Unexpectedly, PMV depletion leads to parasite death immediately after invasion of RBCs, distinct from other disruptions of the export pathway. This suggests that PMV inhibitors could lead to relatively rapid parasite death, and that PMV has additional unexplored role(s) during RBC infection.