Artemisinin binds and inhibits the activity of plasmodium falciparum ddi1, a retroviral aspartyl protease

Abstract

Reduced sensitivity of the human malaria parasite, Plasmodium falciparum, to Artemisinin and its derivatives (ARTs) threatens the global efforts towards eliminating malaria. ARTs have been shown to cause ubiquitous cellular and genetic insults, which results in the activation of the unfolded protein response (UPR) pathways. The UPR restores protein homeostasis, which otherwise would be toxic to cellular survival. Here, we interrogated the role of DNA-damage inducible protein 1 (Pf Ddi1), a unique proteasome-interacting retropepsin in mediating the actions of the ARTs. We demonstrate that Pf Ddi1 is an active A2 family protease that hydrolyzes ubiquitinated proteasome substrates. Treatment of P. falciparum parasites with ARTs leads to the accumulation of ubiquitinated proteins in the parasites and blocks the destruction of ubiquitinated proteins by inhibiting the Pf Ddi1 protease activity. Besides, whereas the Pf Ddi1 is predominantly localized in the cytoplasm, exposure of the parasites to ARTs leads to DNA fragmentation and increased recruitment of the Pf Ddi1 into the nucleus. Furthermore, we show that Ddi1 knock-out Saccharomyces cerevisiae cells are more susceptible to ARTs and the Pf DdI1 protein robustly restores the corresponding functions in the knock-out cells. Together, these results show that ARTs act in multiple ways; by inducing DNA and protein damage and might be impairing the damage recovery by inhibiting the activity of Pf Ddi1, an essential ubiquitin-proteasome retropepsin.