Role of Plasmodium berghei cGMP-dependent Protein Kinase in Late Liver Stage Development

Abstract

The liver is the first organ infected by Plasmodium sporozoites during malaria infection. In the infected hepatocytes, sporozoites undergo a complex developmental program to eventually generate hepatic merozoites that are released into the bloodstream in membrane-bound vesicles termed merosomes. Parasites blocked at an early developmental stage inside hepatocytes elicit a protective host immune response, making them attractive targets in the effort to develop a pre-erythrocytic stage vaccine. Here, we generated parasites blocked at a late developmental stage inside hepatocytes by conditionally disrupting the Plasmodium berghei cGMP-dependent protein kinase in sporozoites. Mutant sporozoites are able to invade hepatocytes and undergo intracellular development. However, they remain blocked as late liver stages that do not release merosomes into the medium. These late arrested liver stages induce protection in immunized animals. This suggests that, similar to the well studied early liver stages, late stage liver stages too can confer protection from sporozoite challenge.Malaria is among the deadliest infectious diseases in the world. It is caused by protozoan parasites of the genus Plasmodium that undergo a complex life cycle in the mammalian host and the mosquito vector. A human malaria infection begins when a Plasmodium sporozoite delivered through the bite of an infected mosquito infects a hepatocyte in the host liver. Within an intrahepatic membrane-bound vacuole the sporozoite undergoes extensive physical transformation followed by nuclear divisions, cytoplasmic segmentation, and eventually the formation of thousands of merozoites (1). Merozoites exit the infected hepatocyte by budding off in membrane-bound vesicles termed merosomes (2). Merosomes extrude from the infected hepatocyte through the endothelial cell layer and are released into the neighboring sinusoids. Thus, hepatic merozoites are delivered directly into the blood stream where they initiate invasion of erythrocytes and the symptomatic phase of a malaria infection (2). Unlike other stages of the Plasmodium life cycle, the stages that develop inside the hepatocytes, called "liver stages" (LSs), 3 are relatively poorly understood. Although the execution of the LS developmental program must require a large repertoire of molecules, only a few have been functionally identified so far (3-10). LS are of significant clinical and biological interest. Inhibiting the growth of LS could prevent the pathology associated with the erythrocytic stages of a malaria infection. The morbidity associated with Plasmodium vivax, the major human species in South America and South Asia, partly results from its ability to form dormant liver stages, termed hypnozoites, against which there are few effective treatment options (11). Reactivated hypnozoites can cause disease relapse up to a year after initial infection. Finally, LS have long been recognized to be ideal targets for developing a pre-erythrocytic stage malaria vaccine. Animals immunized with irradiated or genetica...

Sterile Protective Immunity to Malaria is Associated with a Panel of Novel P. falciparum Antigens

“…Protein kinases are also known to be involved in sporozoite infectivity (reviewed in (81)); the conditional disruption of P. berghei cGMP-dependent protein kinase in sporozoites arrested development at the late liver stage and conferred protection against wild-type sporozoites (82). In addition, P. berghei calcium dependent PK-6 mutants were less infective for hepatocytes and had a delayed onset of blood stage malaria (83).…”

Characterization of the cGMP-dependent protein kinase SmcGK1 of Schistosoma mansoni

“…cGKs participate in gliding motility of Plasmodium (Moon et al 2009), Eimeria and Toxoplasma and is involved in the host invasion of the latter two parasites (Wiersma et al 2004). Also, in Plasmodium they influence blood-stage schizogony (Taylor et al 2010), merosome egress from liver cells (Falae et al 2010), and finally influence gametogenesis (McRobert et al 2008). In eukaryotes cGKs play diverse physiological functions (Lohmann and Walter 2005), among which a cGMP/PKG pathway was hypothesized to be involved in oocyte maturation (Petr et al 2006, Zhang et al 2005.…”

A Key Role for Plasmodium Subtilisin-like SUB1 Protease in Egress of Malaria Parasites from Host Hepatocytes

“…In addition, the P. berghei cGMP-dependent protein kinase (PbPKG) has been shown to play a key role for the egress of hepatic merozoites (40). Its P. falciparum orthologue (PfPKG) has recently been shown to participate in an intracellular shift in calcium concentration that triggers the secretion of the B, PCR genotyping of the sub1 locus using the P1-P2 and P3-P2 primers ( Fig.…”