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The cytoplasmic domain of the Plasmodium falciparum ligand EBA-175 is essential for invasion but not protein trafficking.

Gilberger TW, Thompson JK, Reed MB, Good RT, Cowman AF - J. Cell Biol. (2003)

Bottom Line: The invasion of host cells by the malaria parasite Plasmodium falciparum requires specific protein-protein interactions between parasite and host receptors and an intracellular translocation machinery to power the process.In this report, we show that the cytoplasmic domain of EBA-175 encodes crucial information for its role in merozoite invasion, and that trafficking of this protein is independent of this domain.These results show that the parasite uses the same components of its cellular machinery for invasion regardless of the host cell type and invasive form.

View Article: PubMed Central - PubMed

Affiliation: The Walter and Eliza Hall Institute of Medical Research, Melbourne 3050, Australia.

ABSTRACT
The invasion of host cells by the malaria parasite Plasmodium falciparum requires specific protein-protein interactions between parasite and host receptors and an intracellular translocation machinery to power the process. The transmembrane erythrocyte binding protein-175 (EBA-175) and thrombospondin-related anonymous protein (TRAP) play central roles in this process. EBA-175 binds to glycophorin A on human erythrocytes during the invasion process, linking the parasite to the surface of the host cell. In this report, we show that the cytoplasmic domain of EBA-175 encodes crucial information for its role in merozoite invasion, and that trafficking of this protein is independent of this domain. Further, we show that the cytoplasmic domain of TRAP, a protein that is not expressed in merozoites but is essential for invasion of liver cells by the sporozoite stage, can substitute for the cytoplasmic domain of EBA-175. These results show that the parasite uses the same components of its cellular machinery for invasion regardless of the host cell type and invasive form.

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The deletion of the cytoplasmic domain of EBA-175 leads to a switch in the invasion phenotype. The TRAP cytoplasmic tail can reconstitute the function of the same region in EBA-175 for merozoite invasion. The ability to invade neuraminidase-treated erythrocytes is indicative of a switch in invasion to sialic acid independence and loss of EBA-175 function and this loss of function was confirmed directly by measurement of the ability to invade chymotrypsin-treated erythrocytes. RBCs were treated with neuraminidase (A) or chymotrypsin (B) as described in Materials and methods before testing in merozoite invasion assays. Figures shown are the percentage of invasion compared with untreated erythrocytes. Error bars correspond to standard deviation. The data obtained with neuraminidase-treated erythrocytes were from four independent experiments for W2mef, W2mef3′R and W2mefΔtail but only two independent experiments for W2mefTRAP. All experiments were performed in triplicate. The data obtained in panel B with chymotrypsin-treated erythrocytes represent one experiment done in triplicate. A second independent experiment has been performed and the results obtained are essentially the same as described here.
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fig4: The deletion of the cytoplasmic domain of EBA-175 leads to a switch in the invasion phenotype. The TRAP cytoplasmic tail can reconstitute the function of the same region in EBA-175 for merozoite invasion. The ability to invade neuraminidase-treated erythrocytes is indicative of a switch in invasion to sialic acid independence and loss of EBA-175 function and this loss of function was confirmed directly by measurement of the ability to invade chymotrypsin-treated erythrocytes. RBCs were treated with neuraminidase (A) or chymotrypsin (B) as described in Materials and methods before testing in merozoite invasion assays. Figures shown are the percentage of invasion compared with untreated erythrocytes. Error bars correspond to standard deviation. The data obtained with neuraminidase-treated erythrocytes were from four independent experiments for W2mef, W2mef3′R and W2mefΔtail but only two independent experiments for W2mefTRAP. All experiments were performed in triplicate. The data obtained in panel B with chymotrypsin-treated erythrocytes represent one experiment done in triplicate. A second independent experiment has been performed and the results obtained are essentially the same as described here.

Mentions: Immunolocalization of EBA-175 mutant proteins in transgenic parasites. The names on the left refer to the W2mef parasite lines expressing mutant EBA-175 proteins. The structure of EBA-175 and mutant proteins are schematically shown. Free merozoites were incubated with anti-EBA175 and anti-EBA181, followed by FITC-labeled anti–mouse and rhodamine-labeled anti–rabbit antibodies. To precisely visualize the localization of mutant EBA-175 with respect to the microneme protein EBA-181, the two fluorescence photomicrographs were merged. The function of the mutant EBA-175 was measured in each parasite line as shown in Fig. 4 A. + refers to absence of a switch in invasion demonstrating EBA-175 function is retained, whereas − signifies a switch in invasion phenotype and loss of function for EBA-175.


The cytoplasmic domain of the Plasmodium falciparum ligand EBA-175 is essential for invasion but not protein trafficking.

Gilberger TW, Thompson JK, Reed MB, Good RT, Cowman AF - J. Cell Biol. (2003)

The deletion of the cytoplasmic domain of EBA-175 leads to a switch in the invasion phenotype. The TRAP cytoplasmic tail can reconstitute the function of the same region in EBA-175 for merozoite invasion. The ability to invade neuraminidase-treated erythrocytes is indicative of a switch in invasion to sialic acid independence and loss of EBA-175 function and this loss of function was confirmed directly by measurement of the ability to invade chymotrypsin-treated erythrocytes. RBCs were treated with neuraminidase (A) or chymotrypsin (B) as described in Materials and methods before testing in merozoite invasion assays. Figures shown are the percentage of invasion compared with untreated erythrocytes. Error bars correspond to standard deviation. The data obtained with neuraminidase-treated erythrocytes were from four independent experiments for W2mef, W2mef3′R and W2mefΔtail but only two independent experiments for W2mefTRAP. All experiments were performed in triplicate. The data obtained in panel B with chymotrypsin-treated erythrocytes represent one experiment done in triplicate. A second independent experiment has been performed and the results obtained are essentially the same as described here.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172798&req=5

fig4: The deletion of the cytoplasmic domain of EBA-175 leads to a switch in the invasion phenotype. The TRAP cytoplasmic tail can reconstitute the function of the same region in EBA-175 for merozoite invasion. The ability to invade neuraminidase-treated erythrocytes is indicative of a switch in invasion to sialic acid independence and loss of EBA-175 function and this loss of function was confirmed directly by measurement of the ability to invade chymotrypsin-treated erythrocytes. RBCs were treated with neuraminidase (A) or chymotrypsin (B) as described in Materials and methods before testing in merozoite invasion assays. Figures shown are the percentage of invasion compared with untreated erythrocytes. Error bars correspond to standard deviation. The data obtained with neuraminidase-treated erythrocytes were from four independent experiments for W2mef, W2mef3′R and W2mefΔtail but only two independent experiments for W2mefTRAP. All experiments were performed in triplicate. The data obtained in panel B with chymotrypsin-treated erythrocytes represent one experiment done in triplicate. A second independent experiment has been performed and the results obtained are essentially the same as described here.
Mentions: Immunolocalization of EBA-175 mutant proteins in transgenic parasites. The names on the left refer to the W2mef parasite lines expressing mutant EBA-175 proteins. The structure of EBA-175 and mutant proteins are schematically shown. Free merozoites were incubated with anti-EBA175 and anti-EBA181, followed by FITC-labeled anti–mouse and rhodamine-labeled anti–rabbit antibodies. To precisely visualize the localization of mutant EBA-175 with respect to the microneme protein EBA-181, the two fluorescence photomicrographs were merged. The function of the mutant EBA-175 was measured in each parasite line as shown in Fig. 4 A. + refers to absence of a switch in invasion demonstrating EBA-175 function is retained, whereas − signifies a switch in invasion phenotype and loss of function for EBA-175.

Bottom Line: The invasion of host cells by the malaria parasite Plasmodium falciparum requires specific protein-protein interactions between parasite and host receptors and an intracellular translocation machinery to power the process.In this report, we show that the cytoplasmic domain of EBA-175 encodes crucial information for its role in merozoite invasion, and that trafficking of this protein is independent of this domain.These results show that the parasite uses the same components of its cellular machinery for invasion regardless of the host cell type and invasive form.

View Article: PubMed Central - PubMed

Affiliation: The Walter and Eliza Hall Institute of Medical Research, Melbourne 3050, Australia.

ABSTRACT
The invasion of host cells by the malaria parasite Plasmodium falciparum requires specific protein-protein interactions between parasite and host receptors and an intracellular translocation machinery to power the process. The transmembrane erythrocyte binding protein-175 (EBA-175) and thrombospondin-related anonymous protein (TRAP) play central roles in this process. EBA-175 binds to glycophorin A on human erythrocytes during the invasion process, linking the parasite to the surface of the host cell. In this report, we show that the cytoplasmic domain of EBA-175 encodes crucial information for its role in merozoite invasion, and that trafficking of this protein is independent of this domain. Further, we show that the cytoplasmic domain of TRAP, a protein that is not expressed in merozoites but is essential for invasion of liver cells by the sporozoite stage, can substitute for the cytoplasmic domain of EBA-175. These results show that the parasite uses the same components of its cellular machinery for invasion regardless of the host cell type and invasive form.

Show MeSH
Related in: MedlinePlus