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Plasmodium falciparum proteins involved in cytoadherence of infected erythrocytes to chemokine CX3CL1.

ABSTRACT: Malaria caused by Plasmodium falciparum is associated with cytoadherence of infected red blood cells (iRBC) to endothelial cells. Numerous host molecules have been involved in cytoadherence, including the adhesive chemokine CX3CL1. Most of the identified parasite ligands are from the multigenic and hypervariable Plasmodium falciparum Erythrocyte Membrane Protein 1 (PfEMP1) family which makes them poor targets for the development of a broadly protective vaccine. Using proteomics, we have identified two 25-kDa parasite proteins with adhesive properties for CX3CL1, called CBP for CX3CL1 Binding Proteins. CBPs are coded by single-copy genes with little polymorphic variation and no homology with other P. falciparum gene products. Specific antibodies raised against epitopes from the predicted extracellular domains of each CBP efficiently stain the surface of RBC infected with trophozoites or schizonts, which is a strong indication of CBP expression at the surface of iRBC. These anti-CBP antibodies partially neutralize iRBC adherence to CX3CL1. This adherence is similarly inhibited in the presence of peptides from the CBP extracellular domains, while irrelevant peptides had no such effect. CBP1 and CBP2 are new P. falciparum ligands for the human chemokine CX3CL1. The identification of this non-polymorphic P. falciparum factors provides a new avenue for innovative vaccination approaches.

SUBMITTER: Hermand P

PROVIDER: S-EPMC5031962 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundPrevious comparative proteomic analysis on Plasmodium falciparum isolates of different adhesion properties suggested that protein phosphorylation varies between isolates with different cytoadherence properties. But the extent and dynamic changes in phosphorylation have not been systematically studied. As a baseline for these future studies, this paper examined changes in the phosphoproteome of parasitized red blood cells (pRBC).MethodsMetabolic labelling with [35S] methionine on pRBC and 2D gel electrophoresis (2-DE) has previously been used to show the expression of parasite proteins and changes in protein iso-electric point (PI). 2-DE of different parasite strains was combined with immunoblotting using monoclonal antibodies specifically to phosphorylated serine/threonine and tyrosine, to obtain the phosphorylation profiles throughout the erythrocytic lifecycle. Affinity chromatography was used to purify/enrich phosphorylated proteins and these proteins from mature trophozoite stages which were identified using high-accuracy mass spectrometry and MASCOT search.Results2D-immunoblots showed that P. falciparum infection greatly increased phosphorylation of a set of proteins in pRBC, the dominant size classes for phosphorylated tyrosine proteins were 95, 60, 50 and 30 kDa and for phosphorylated serine/threonine were 120, 95, 60, 50, 43, 40 and 30 kDa. The most abundant molecules from 2D-gel mapping of phosphorylated proteins in ItG infected RBCs were identified by MALDI-TOF. A proteomic overview of phosphorylated proteins in pRBC was achieved by using complementary phosphorylated protein enrichment techniques combined with nano-flow LC/MS/MS analysis and MASCOT MS/MS ions search with phosphorylation as variable modifications. The definite phosphoproteins of pRBC are reported and discussed.ConclusionProtein phosphorylation is a major process in P. falciparum-parasitized erythrocytes. Preliminary screens identified 170 P. falciparum proteins and 77 human proteins as phosphorylated protein in pRBC, while only 48 human proteins were identified in the corresponding fractions from uninfected RBC. Refinement of the search to include significant ion scores indicating a specific phospho-peptide identified 21 P. falciparum proteins and 14 human proteins from pRBC, 13 host proteins were identified from normal RBC. The results achieved by complementary techniques consistently reflect a reliable proteomic overview of pRBC.

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Plasmodium falciparum proteins involved in cytoadherence of infected erythrocytes to chemokine CX3CL1.

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