Research Article: Stable Translocation Intermediates Jam Global Protein Export in Plasmodium falciparum Parasites and Link the PTEX Component EXP2 with Translocation Activity

Date Published: May 11, 2016

Publisher: Public Library of Science

Author(s): Paolo Mesén-Ramírez, Ferdinand Reinsch, Alexandra Blancke Soares, Bärbel Bergmann, Ann-Katrin Ullrich, Stefan Tenzer, Tobias Spielmann, Dominique Soldati-Favre.

http://doi.org/10.1371/journal.ppat.1005618

Abstract

Protein export is central for the survival and virulence of intracellular P. falciparum blood stage parasites. To reach the host cell, exported proteins cross the parasite plasma membrane (PPM) and the parasite-enclosing parasitophorous vacuole membrane (PVM), a process that requires unfolding, suggestive of protein translocation. Components of a proposed translocon at the PVM termed PTEX are essential in this phase of export but translocation activity has not been shown for the complex and questions have been raised about its proposed membrane pore component EXP2 for which no functional data is available in P. falciparum. It is also unclear how PTEX mediates trafficking of both, soluble as well as transmembrane proteins. Taking advantage of conditionally foldable domains, we here dissected the translocation events in the parasite periphery, showing that two successive translocation steps are needed for the export of transmembrane proteins, one at the PPM and one at the PVM. Our data provide evidence that, depending on the length of the C-terminus of the exported substrate, these steps occur by transient interaction of the PPM and PVM translocon, similar to the situation for protein transport across the mitochondrial membranes. Remarkably, we obtained constructs of exported proteins that remained arrested in the process of being translocated across the PVM. This clogged the translocation pore, prevented the export of all types of exported proteins and, as a result, inhibited parasite growth. The substrates stuck in translocation were found in a complex with the proposed PTEX membrane pore component EXP2, suggesting a role of this protein in translocation. These data for the first time provide evidence for EXP2 to be part of a translocating entity, suggesting that PTEX has translocation activity and provide a mechanistic framework for the transport of soluble as well as transmembrane proteins from the parasite boundary into the host cell.

Partial Text

Five species of Plasmodium parasites cause human malaria. Of these P. falciparum is responsible for the majority of the over 500’000 annually recorded malaria deaths [1]. The pathology of malaria is caused by the continuous propagation of the parasite within red blood cells (RBCs). In this phase P. falciparum parasites modify extensively the host RBC by exporting hundreds of different proteins into the infected cell. These modifications include host cell surface changes that cause the sequestration of infected RBCs (iRBCs) in the vasculature, a phenomenon considered to be a major contributor to parasite virulence [2]. Other changes are required for nutrient acquisition, to adjust RBC rigidity and to facilitate protein trafficking in the host cell [3]. Protein export is therefore central for blood stage development and malaria pathology.

Here we for the first time obtained intermediates of exported proteins inducibly and stably arrested during translocation into the host cell. Intriguingly, these translocation intermediates prevented the transport of all known types of exported proteins, demonstrating that the actual translocation is a point of convergence for all exported proteins and a single kind of protein-conducting channel mediates export. Our data further support a two-step translocation process for exported TM proteins which are first extracted out of the PPM and then translocated into the host cell in a second unfolding-dependent process at the PVM.

 

Source:

http://doi.org/10.1371/journal.ppat.1005618

 

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