Selective autophagy and related mechanisms can act as variable defense mechanisms against pathogens and can therefore be considered as intracellular immune responses. When in hepatocytes, Plasmodium parasites reside in a parasitophorous vacuole (PV) and the PV membrane (PVM) is the main contact site between host cell and parasite. Early in infection, the PVM is directly labeled with host cell autophagy proteins LC3B and p62 (nucleoporin 62). We investigated the recruitment of different selective autophagy receptors and could show that mainly p62 and NBR1 (neighbour of BRCA1 gene 1) and to a lesser extent NDP52 (nuclear dot protein 52) associate with the PVM. To investigate the recruitment of these receptors to the PVM in Plasmodium-infected cells, we generated LC3B knock out HeLa cells. In these cell lines, autophagosome formation and autophagic flux are not different to those in WT cells. Unexpectedly, p62 and NBR1 recruitment to the PVM was strongly impaired in LC3B-negative host cells, suggesting that LC3B recruits both receptors to the PVM of Plasmodium parasites. We also noticed that LC3B recruited ubiquitin to the PVM. This indicates that, in comparison to classical selective autophagy, in P. berghei-infected cells the order of membrane labeling with autophagy proteins appears to be inverted from canonical ubiquitin-receptor-LC3B recruitment to LC3B-receptor and possibly ubiquitin.

Plasmodium parasites have a complex life cycle alternating between a mosquito and a vertebrate host. Following the bite of an Anopheles female mosquito, Plasmodium sporozoites are transmitted from the skin to the liver; their first place of replication within the host. Successfully invaded sporozoites undergo a massive replication and growth involving asynchronous DNA replication and division that results in the generation of tens of thousands or even hundreds of thousands of merozoites depending on the Plasmodium species. The generation of a high number of daughter parasites requires biogenesis and segregation of organelles to finally reach a relatively synchronous cytokinesis event. At the end of liver stage (LS) development, merozoites are packed into merosomes and released into the bloodstream. They are then liberated and infect red blood cells to again produce merozoites by schizogony for the erythrocytic stage of the life cycle. Although parasite LS and asexual blood stage (ABS) differ in many respects, important similarities exist between the two. This review focuses on the cell division of Plasmodium parasite LS in comparison with other life cycle stages especially the parasite blood stage.

The protozoan parasite Plasmodium, causative agent of malaria, invades hepatocytes by invaginating the host cell plasma membrane and forming a parasitophorous vacuole membrane (PVM). Surrounded by this PVM, the parasite undergoes extensive replication. Parasites inside a PVM provoke the Plasmodium-associated autophagyrelated (PAAR) response. This is characterised by a long-lasting association of the autophagy marker protein LC3 with the PVM, which is not preceded by phosphatidylinositol 3-phosphate (PI3P)-labelling. Prior to productive invasion, sporozoites transmigrate several cells and here we describe that a proportion of traversing sporozoites become trapped in a transient traversal vacuole, provoking a host cell response that clearly differs from the PAAR response. These trapped sporozoites provoke PI3P-labelling of the surrounding vacuolar membrane immediately after cell entry, followed by transient LC3-labelling and elimination of the parasite by lysosomal acidification. Our data suggest that this PI3P response is not only restricted to sporozoites trapped during transmigration but also affects invaded parasites residing in a compromised vacuole. Thus, host cells can employ a pathway distinct from the previously described PAAR response to efficiently recognise and eliminate Plasmodium parasites.

The protozoan parasite Plasmodium, causative agent of malaria, initially invades and develops in hepatocytes where it resides in a parasitophorous vacuole (PV). A single invaded parasite develops into thousands of daughter parasites. Survival of the host cell is crucial for successful completion of liver stage development. Nuclear factor erythroid-derived 2-related factor 2 (NRF2) is a transcription factor known to induce transcription of cytoprotective genes when activated. Here we show that NRF2 is activated in Plasmodium berghei-infected hepatocytes. We observed that this NRF2 activation depends on PV membrane resident p62 recruiting KEAP1, the negative regulator of NRF2. Disrupting the NRF2 gene results in reduced parasite survival, indicating that NRF2 signaling is an important event for parasite development in hepatocytes. Together, our observations uncovered a novel mechanism of how Plasmodium parasites ensure host cell survival during liver stage development.

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Immunofluorescence image of Plasmodium berghei sporozoites (red) in HeLa cells 1.5 hours post infection. The parasitophorous vacuole membrane (PVM) that surrounds the parasites was stained with antibodies against the PVM‐resident protein UIS4 (grey). DNA was stained with DAPI (blue). Imaging was performed on a Leica SP8 confocal microscope. For further details, readers are referred to the article by Bindschedler et al. on https://doi.org/10.1111/cmi.13271 of this issue.