Spatial organization of protein export in malaria parasite blood stages

Abstract

Plasmodium falciparum, which causes malaria, extensively remodels its human host cells, particularly erythrocytes. Remodelling is essential for parasite survival by helping to avoid host immunity and assisting in the uptake of plasma nutrients to fuel rapid growth. Host cell renovation is carried out by hundreds of parasite effector proteins that are exported into the erythrocyte across an enveloping parasitophorous vacuole membrane (PVM). The Plasmodium translocon for exported (PTEX) proteins is thought to span the PVM and provide a channel that unfolds and extrudes proteins across the PVM into the erythrocyte. We show that exported reporter proteins containing mouse dihydrofolate reductase domains that inducibly resist unfolding become trapped at the parasite surface partly colocalizing with PTEX. When cargo is trapped, loop-like extensions appear at the PVM containing both trapped cargo and PTEX protein EXP2, but not additional components HSP101 and PTEX150. Following removal of the block-inducing compound, export of reporter proteins only partly recovers possibly because much of the trapped cargo is spatially segregated in the loop regions away from PTEX. This suggests that parasites have the means to isolate unfoldable cargo proteins from PTEX-containing export zones to avert disruption of protein export that would reduce parasite growth.

EXP1 is required for organisation of EXP2 in the intraerythrocytic malaria parasite vacuole

“…These sites may correspond with the membrane abnormalities observed by transmission electron microscopy (Figure 5d). Similar loop‐like structures containing EXP2 but not other PTEX components have been observed to form in parasites expressing conditionally unfoldable exported cargo, suggesting they may represent a generalised response to perturbations in the PV (Charnaud, Jonsdottir, et al, ). EXP2 forms a dual functional pore in the PVM that is required for small molecule transport and effector protein translocation (Garten et al, ; Ho et al, ).…”

“…have been observed to form in parasites expressing conditionally unfoldable exported cargo, suggesting they may represent a generalised response to perturbations in the PV (Charnaud, Jonsdottir, et al, 2018). EXP2 forms a dual functional pore in the PVM that is required for small molecule transport and effector protein translocation Ho et al, 2018).…”

“…One of the most striking features of the PV is the intimate proximity of the PVM and PPM, which is maintained until a very late stage of parasite development when PVM rounding occurs just prior to egress (Glushakova et al, 2018;Sherling & van Ooij, 2016). Additional lateral organisation of this compartment is suggested by the formation of distinct oligomeric arrays of EXP1 and ETRAMPs in the PVM (Spielmann, Gardiner, Beck, Trenholme, & Kemp, 2006) and by the nonuniform distribution of PTEX components as well as PV-targeted exported and nonexported fluorescent fusion reporter proteins, which have been shown to display a punctate distribution in the PV described as a "necklace of beads" (Adisa et al, 2003;Bullen et al, 2012;Charnaud, Jonsdottir, et al, 2018;de Koning-Ward et al, 2009;Riglar et al, 2013;Wickham et al, 2001). Visualised by immunofluorescence, this punctate arrangement is most prominent in the early ring stage of parasite development and resolves into a more homogenous distribution in the trophozoite and schizont stage, particularly for EXP2 (Charnaud, Jonsdottir, et al, 2018).…”

Illuminating how malaria parasites export proteins into host erythrocytes

“…However, under particular conditions, such as during heat shock, cargo may be more prone to aggregation and presumably the parasite would have to prevent a build‐up of cargo clogging the translocon by regulating protein export. Interestingly, loop‐shaped structures that comprise EXP2 but not HSP101 or PTEX150 have been observed at the PVM when cargo is induced to become resistant to unfolding through its fusion to mouse dihydrofolate reductase domain (Charnaud et al, ). This is unlikely to be the form of EXP2 that creates the nutrient channels in the PVM (Garten et al, ) because the EXP2 in this case colocalises with unfolded cargo.…”

“…This is unlikely to be the form of EXP2 that creates the nutrient channels in the PVM (Garten et al, ) because the EXP2 in this case colocalises with unfolded cargo. Instead, it has been postulated that the formation of these loop regions allows the spatial segregation of cargo that cannot be unfolded, which would otherwise clog the translocons and impact protein export and parasite growth (Charnaud et al, ). Membranous extensions of the PVM have also been observed when EXP2 and HSP101 are perturbed, suggesting that a reduction of normal protein export may produce these features perhaps due to a build‐up of exported proteins in the PV (Garten et al, ).…”