Plasmodium falciparum dipeptidyl aminopeptidase 3 activity is important for efficient erythrocyte invasion.

Parasite egress from infected erythrocytes and invasion of new red blood cells are essential for the exponential asexual replication of the malaria parasite resulting in malaria pathology. These two tightly coordinated processes take place in less than a minute and are regulated and mediated by proteases. The putative cysteine protease dipeptidyl aminopeptidase 3 (DPAP3) was previously suggested to play an essential role as a regulator of parasite egress, but little is known about its biological function. In this study, we demonstrate that DPAP3 has proteolytic activity, but contrary to previously studies DPAPs, removal of its internal prodomain is not required for activation. By combining super resolution microscopy with time-lapse fluorescence microscopy and immuno electron microscopy, we show that P. falciparum DPAP3 localizes to merozoite apical organelles from which it is secreted immediately before parasite egress. Using a conditional knock out approach coupled to complementation studies with wild type or mutant DPAP3, we show that DPAP3 activity is important for parasite replication and is critical for efficient RBC invasion. Finally, we demonstrate that DPAP3 does not play a significant role in parasite egress, and that the block in egress phenotype previously reported for DPAP3 inhibitors is due to off target or toxicity effects.