Malaria Parasite cGMP-dependent Protein Kinase Regulates Blood Stage Merozoite Secretory Organelle Discharge and Egress

Abstract

The malaria parasite replicates within an intraerythrocytic parasitophorous vacuole (PV). Eventually, in a tightly regulated process called egress, proteins of the PV and intracellular merozoite surface are modified by an essential parasite serine protease called PfSUB1, whilst the enclosing PV and erythrocyte membranes rupture, releasing merozoites to invade fresh erythrocytes. Inhibition of the Plasmodium falciparum cGMP-dependent protein kinase (PfPKG) prevents egress, but the underlying mechanism is unknown. Here we show that PfPKG activity is required for PfSUB1 discharge into the PV, as well as for release of distinct merozoite organelles called micronemes. Stimulation of PfPKG by inhibiting parasite phosphodiesterase activity induces premature PfSUB1 discharge and egress of developmentally immature, non-invasive parasites. Our findings identify the signalling pathway that regulates PfSUB1 function and egress, and raise the possibility of targeting PfPKG or parasite phosphodiesterases in therapeutic approaches to dysregulate critical protease-mediated steps in the parasite life cycle.

Author Summary

Malaria is a scourge of the developing world and many researchers are seeking new ways to treat and control the disease. Malaria is caused by a single-celled parasite that grows within red blood cells, eventually rupturing them to release invasive merozoites in a process known as egress. In earlier work we found that, just prior to egress, an enzyme called SUB1 is released from the intracellular parasites into the vacuole in which they reside. SUB1 then cleaves a number of proteins required for egress and development of invasive merozoites. The signals that control SUB1 discharge are poorly understood. In this work, we show that SUB1 release requires the activity of another parasite enzyme called protein kinase G (PKG), which is in turn activated by a small molecule called cGMP. Inhibition of PKG blocks SUB1 discharge and egress, whilst premature activation of PKG by a member of a class of compounds called phosphodiesterase inhibitors, which increase cGMP levels in the parasite, induces premature egress of mostly non-invasive merozoites. These findings increase our understanding of egress and show that both malarial PKG and parasite phosphodiesterases (which are validated drug targets in humans) are potential targets for a new class of antimalarial drugs.