The Host Endocytic Pathway is Essential for Plasmodium berghei Late Liver Stage Development

Abstract

The obligate intracellular liver stage of the Plasmodium parasite represents a bottleneck in the parasite life cycle and remains a promising target for therapeutic intervention. During this stage, parasites undergo dramatic morphological changes and achieve one of the fastest replication rates among eukaryotic species. Nevertheless, relatively little is known about the parasite interactions with the host hepatocyte. Using immunofluorescence, live cell imaging and electron microscopy, we show that Plasmodium berghei parasites are surrounded by vesicles from the host late endocytic pathway. We found that these vesicles are acidic and contain the membrane markers Rab7a, CD63 and LAMP1. When host cell vesicle acidification was disrupted using ammonium chloride or Concanamycin A during the late liver stage of infection, parasite survival was not affected, but schizont size was significantly decreased. Furthermore, when the host cell endocytic pathway was loaded with BSA-gold, gold particles were found within the parasite cytoplasm, showing the transport of material from the host endocytic pathway toward the parasite interior. These observations reveal a novel Plasmodium-host interaction and suggest that vesicles from the host endolysosomal pathway could represent an important source of nutrients exploited by the fastgrowing late liver stage parasites.

Long-term live imaging reveals cytosolic immune responses of host hepatocytes againstPlasmodiuminfection and parasite escape mechanisms

“…It is not known why the PV of successfully developing parasites does not acidify, despite fusion of the PVM with lysosomes, as shown in this and a previous study. 20 One possibility is that the PVM contains outward-directed proton pumps. However, this may not be necessary considering that the PVM contains pores for the easy uptake of nutrients of up to 855 Da, 39 and that proton gradients may not be able to form in such an environment because they need tight membranes without pores.…”

“…19 One study suggested that lysosomes fuse with the PVM in P. berghei-infected hepatocytes. 20 We therefore sought to determine i) the dynamics of lysosome fusion in relation to LC3 recruitment to the PVM in infected cells, ii) whether the pH drops in the PV, and iii) whether the lysosomal protease CTSD (cathepsin D) is recruited to the PV.…”

Plasmodium berghei EXP-1 interacts with host Apolipoprotein H during Plasmodium liver-stage development

“…A total of 50,000 and 25,000 Huh7 cells grown on coverslips or eight-well Lab-Tek chamber slides (Nunc), respectively, were seeded 1 d before infection with 10,000 Pb sporozoites and fixed at the indicated time points (10,15,18,21,24, and 48 hpi) after sporozoite addition. Cells grown on coverslips were fixed with 4% (vol/vol) paraformaldehyde for 15 min, washed three times with PBS, permeabilized in 0.2% (vol/vol) saponin or Triton in PBS for 20 min, blocked for 1 h with 0.1% (vol/vol) Triton in 1% (wt/vol) BSA in PBS, and incubated with the appropriate primary antibodies in blocking buffer for 1 h at room temperature or overnight at 4°C.…”

“…However, little is known about intrahepatic Plasmodium stages with regard to interactions between the parasite and the host hepatocyte and their potential for nutrient uptake and/or exchange. Small molecules (up to 800 Da) can cross the PVM freely via specialized transport channels (14), whereas larger molecules might reach the parasite via association and possibly fusion of late endosomes, lysosomes, or amphisomes with the PVM (15)(16)(17)(18)(19).…”

The role of nanogold in human tropical diseases: research, detection and therapy

“…Recently, Lopes da Silva et al [56] used bovine serum albumin (BSA) linked to 10 nm AuNP in order to follow their position in the parasite-host cell system, in the liver stage, a crucial phase of the Plasmodium development. The authors showed that when BSA-Au are loaded into the host cell endocytic pathway, the AuNP are later found within the parasite cytoplasm, showing the transport of materials from the host endocytic pathway towards the parasite interior, revealing a possible form of the parasite to obtain nutrients.…”