Overlapping and distinct roles of CDPK family members in the pre-erythrocytic cycle of the rodent malaria parasite, Plasmodium berghei

Invasion of, development within, and exit from hepatocytes by Plasmodium is essential for the parasite to establish the malaria-causing erythrocytic cycle. Identification of signaling pathways that operate during the pre-erythrocytic cycle provides insights into a critical stage of infection and potential targets for chemoprevention of disease. Calcium Dependent Protein Kinases (CDPK) represent a kinase family that is present in Plasmodium but absent in mammals. We demonstrate that P. berghei homologs of CDPK1, CDPK4 and CDPK5 play overlapping but distinct roles in sporozoite invasion and parasite egress from hepatocytes. All three kinases are expressed in sporozoites. All three are required for optimal motility of sporozoites and consequently their invasion of hepatocytes. Increased cGMP compensates for the functional loss of CDPK1 and CDPK5 during sporozoite invasion but cannot overcome CDPK4’s loss. CDPK1 and CDPK5 expression is downregulated after sporozoite invasion. CDPK5 reappears in a subset of late stage liver stages and is present in all merosomes. Chemical inhibition of CDPK4 and depletion of CDPK5 in liver stages suggests that these kinases play a role in the formation and/or release of merosomes from mature liver stages. Furthermore, depletion of CDPK5 in merosomes significantly delays a merosome-initiated erythrocytic cycle without affecting the infectivity of hepatic merozoites. These data suggest that CDPK5 is required for the release of hepatic merozoites from merosomes. Our work provides the first evidence that sporozoite invasion requires CDPK1 and CDPK5 and that the release of hepatic merozoites is a regulated process.SignificanceThe malaria-parasite Plasmodium begins its mammalian cycle by infecting hepatocytes in the liver. A single parasite differentiates into tens of thousands of hepatic merozoites which exit the host cell in vesicles called merosomes. Hepatic merozoites initiate the first round of erythrocytic infection that leads to disease symptoms. We show that optimal invasion of liver cells by Plasmodium requires the action of three closely-related parasite kinases, CDPK1, 4 and 5. Loss of any of the three enzymes in the parasite significantly reduces infection of liver cells. CDPK5 is also required for the release of hepatic merozoites from merosomes and therefore for initiating the erythrocytic cycle. A better understanding of how these kinases function could lead to drugs that prevent malaria.