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Formation of ER-lumenal intermediates during export of Plasmodium proteins containing transmembrane-like hydrophobic
Yvette S Levray1, Bianca Bana2, Sarah J Tarr1,2
1Institute of Structural and Molecular Biology, University College London, London, United Kingdom.
Abstract:
During the blood stage of a malaria infection, malaria parasites export both soluble and membrane proteins into the erythrocytes in which they reside. Exported proteins are trafficked via the parasite endoplasmic reticulum and secretory pathway, before being exported across the parasitophorous vacuole membrane into the erythrocyte. Transport across the parasitophorous vacuole membrane requires protein unfolding, and in the case of membrane proteins, extraction from the parasite plasma membrane. We show that trafficking of the exported Plasmodium protein, Pf332, differs from that of canonical eukaryotic soluble-secreted and transmembrane proteins. Pf332 is initially ER-targeted by an internal hydrophobic sequence that unlike a signal peptide, is not proteolytically removed, and unlike a transmembrane segment, does not span the ER membrane. Rather, both termini of the hydrophobic sequence enter the ER lumen and the ER-lumenal species is a productive intermediate for protein export. Furthermore, we show in intact cells, that two other exported membrane proteins, SBP1 and MAHRP2, assume a lumenal topology within the parasite secretory pathway. Although the addition of a C-terminal ER-retention sequence, recognised by the lumenal domain of the KDEL receptor, does not completely block export of SBP1 and MAHRP2, it does enhance their retention in the parasite ER. This indicates that a sub-population of each protein adopts an ER-lumenal state that is an intermediate in the export process. Overall, this suggests that although many exported proteins traverse the parasite secretory pathway as typical soluble or membrane proteins, some exported proteins that are ER-targeted by a transmembrane segment-like, internal, non-cleaved hydrophobic segment, do not integrate into the ER membrane, and form an ER-lumenal species that is a productive export intermediate. This represents a novel means, not seen in typical membrane proteins found in model systems, by which exported transmembrane-like proteins can be targeted and trafficked within the lumen of the secretory pathway.
Insights
Malaria parasites export proteins using a novel pathway. Some exported proteins, like Pf332, use an internal sequence to enter the endoplasmic reticulum lumen as an export intermediate.
Area of Science:
- Cell Biology
- Parasitology
- Molecular Biology
Background:
- Malaria parasites export proteins into host erythrocytes during blood-stage infection.
- Protein export involves the parasite's endoplasmic reticulum (ER) and secretory pathway.
- Exported proteins cross the parasitophorous vacuole membrane, often requiring unfolding or membrane extraction.
Purpose of the Study:
- To investigate the unique trafficking pathway of the exported Plasmodium protein, Pf332.
- To determine if other exported membrane proteins utilize similar non-canonical ER targeting and trafficking mechanisms.
- To elucidate novel mechanisms of protein export in malaria parasites.
Main Methods:
- Analysis of Pf332 trafficking using its internal hydrophobic sequence for ER targeting.
- Investigation of the topology of exported membrane proteins SBP1 and MAHRP2 within the parasite secretory pathway.
- Functional studies involving the addition of an ER-retention sequence to SBP1 and MAHRP2.
Main Results:
- Pf332 utilizes an internal hydrophobic sequence for ER targeting, which is not cleaved and does not span the ER membrane.
- Both termini of Pf332's hydrophobic sequence enter the ER lumen, forming a lumenal intermediate for export.
- SBP1 and MAHRP2 also adopt lumenal topologies in the secretory pathway, with some ER retention observed upon addition of a KDEL sequence.
Conclusions:
- Some malaria parasite exported proteins, particularly those with internal hydrophobic targeting sequences, do not integrate into the ER membrane.
- These proteins form ER-lumenal species that serve as productive intermediates for export.
- This represents a novel protein trafficking mechanism distinct from canonical eukaryotic pathways for transmembrane proteins.
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