Project

MALARIA; PLASMODIUM FALCIPARUM; PFEMP1; trafficking; exported proteins; protein interaction; MAHRP1; MAHRP2; Tethers; virulence factor; protein trafficking,; interactions

Lead
Lay summary
Plasmodium falciparum, the causative agent of malaria tropica infects human red blood cells. This process is unique because the erythrocyte is a highly specialized cell lacking organelles to support growing and dividing parasites. All metabolic pathways and developmental requirements must be installed by the parasite during the 48 h cycle within a red blood cell. In addition, the parasite substantially modifies the host cell resulting in an adhesive cell and thus leading to substantial morbidity. These cell modifications include the transport of the major virulence factor to the surface of the infected cell. However, invading such a highly specialized cell is rare in biology and requires the interplay of a specific and unique set of parasite proteins to refurbish these cells. In this project, we aim to elucidate the interaction and function of proteins which are exported early during the life cycle from the parasite into the host cell. For this we will use recombinant transfection technology, proteomic analysis, and other molecular techniques to identify and understand essential processes in the parasite's quest to survive in the human host cell. Only the understanding of these processes might enable us to design innovative intervention strategies against this deadly disease.

Direct link to Lay Summary

Last update: 21.02.2013

Active participation

Number	Title	Start	Funding scheme

Background and Hypothesis:Plasmodium falciparum, the causative agent of malaria tropica invades erythrocytes and this stage is the only stage responsible for pathology. Invasion into an empty cell such as the erythrocyte requires unique processes immediately after invasion. These processes involve a complete restructuring of the cell, establishment of a protein translation and trafficking machinery to subsequently deposit adhesion conferring ligands onto the surface of the erythrocyte. We are interested in the interplay of parasite derived proteins which occur early in the erythrocyte life cycle and which are translocated into the cytosol of the infected erythrocyte. We and others have previously shown that some of these proteins are essential in the trafficking of the virulence determinants and that some of these proteins are not amendable to gene knock out approaches, suggesting their necessity for the survival of the parasite. In this project we would like to elucidate the protein-protein interactions of these early exported proteins. Objectives and approach:The overall objective is to elucidate the function and the identification of interaction partners of a group of previously identified PEXEL-negative exported proteins in P. falciparum.In detail we want toI) further develop our existing transfection platform to generate tagged versions of identified proteins to elucidate their localization and to identify interaction partners using pull down experiments and immuno-precipitations.II) generate knock out parasite lines for phenotypical analysis, or if impossible to establish conditional knock down parasite lines using the FKBP-DD approach.III) establish a recombinant screening system for interaction partners using a membrane bound split ubiquitin system in yeast or using a classical yeast two hybrid system for soluble proteins.IV) establish the proteome of tethers by differential ultracentrifugation and mass-spectrometry.V) analyse changes in the proteomic pattern upon over-expression or deletion of genes both for protein expression levels but also for post-translational protein modifications applying whole proteome fluorescent 2D-DIGE (differential gel electrophoresis) with subsequent mass-spectrometrical analysis. Expected outcome:We this project we hope to understand early events essential for parasite survival upon invasion into its host cell. This will not only deepen our understanding of the biology of P. falciparum but might provide new targets for innovative interventions.