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

PLoS Pathog. 2016 May 11;12(5):e1005618. doi: 10.1371/journal.ppat.1005618. eCollection 2016 May.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Blotting, Western
  • Erythrocytes / parasitology
  • Fluorescent Antibody Technique
  • Humans
  • Immunoprecipitation
  • Malaria, Falciparum / metabolism*
  • Plasmodium falciparum / metabolism*
  • Protein Transport / physiology*
  • Protozoan Proteins / metabolism*

Substances

  • Protozoan Proteins

Grants and funding

This work was funded by DFG grant SP1209/1-2 and SP1209/1-3. PMR gratefully acknowledges funding by the DAAD. FR and ABS were supported by the GRK1459. TS and ST acknowledge funding by the DFG priority programme SPP1580: Intracellular compartments as places of pathogen-host-interactions. The publication costs of this article were in part funded by the Open Access fund of the Leibniz Association. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.