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Proteomic profiling of Plasmodium sporozoite maturation identifies new proteins essential for parasite development and infectivity.

Lasonder E, Janse CJ, van Gemert GJ, Mair GR, Vermunt AM, Douradinha BG, van Noort V, Huynen MA, Luty AJ, Kroeze H, Khan SM, Sauerwein RW, Waters AP, Mann M, Stunnenberg HG - PLoS Pathog. (2008)

Bottom Line: The different sporozoite proteomes contain a large number of stage specific proteins whose annotation suggest an involvement in sporozoite maturation, motility, infection of the human host and associated metabolic adjustments.Analyses of proteins identified in the P. falciparum sporozoite proteomes by orthologous gene disruption in the rodent malaria parasite, P. berghei, revealed three previously uncharacterized Plasmodium proteins that appear to be essential for sporozoite development at distinct points of maturation in the mosquito.This study sheds light on the development and maturation of the malaria parasite in an Anopheles mosquito and also identifies proteins that may be essential for sporozoite infectivity to humans.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, NCMLS, Radboud University Nijmegen, Nijmegen, The Netherlands.

ABSTRACT
Plasmodium falciparum sporozoites that develop and mature inside an Anopheles mosquito initiate a malaria infection in humans. Here we report the first proteomic comparison of different parasite stages from the mosquito -- early and late oocysts containing midgut sporozoites, and the mature, infectious salivary gland sporozoites. Despite the morphological similarity between midgut and salivary gland sporozoites, their proteomes are markedly different, in agreement with their increase in hepatocyte infectivity. The different sporozoite proteomes contain a large number of stage specific proteins whose annotation suggest an involvement in sporozoite maturation, motility, infection of the human host and associated metabolic adjustments. Analyses of proteins identified in the P. falciparum sporozoite proteomes by orthologous gene disruption in the rodent malaria parasite, P. berghei, revealed three previously uncharacterized Plasmodium proteins that appear to be essential for sporozoite development at distinct points of maturation in the mosquito. This study sheds light on the development and maturation of the malaria parasite in an Anopheles mosquito and also identifies proteins that may be essential for sporozoite infectivity to humans.

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Distribution of identified P. falciparum proteins over different life-cycle stages.(A) Venn diagram depicting the distribution of detected P. falciparum proteins over three different mosquito life-cycle stages (oocysts, oocyst-derived sporozoites and salivary gland sporozoites). Numbers represent the number of proteins, that are either shared between 2 or 3 stages (overlapping areas) or that are detected in a single stage. (B) Comparison of the expression of P. falciparum proteins detected in the three mosquito stage proteomes to the blood stage proteomes described previously [15]. (C) The percentage of proteins exclusively detected in only one proteome out of 6 different life cycle stage proteomes, i.e. ASX - asexual blood stages; GCT – gametocytes; GAM – gametes; OOC – oocysts; ODS - oocyst-derived sporozoites; SGS - salivary gland sporozoites.
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ppat-1000195-g001: Distribution of identified P. falciparum proteins over different life-cycle stages.(A) Venn diagram depicting the distribution of detected P. falciparum proteins over three different mosquito life-cycle stages (oocysts, oocyst-derived sporozoites and salivary gland sporozoites). Numbers represent the number of proteins, that are either shared between 2 or 3 stages (overlapping areas) or that are detected in a single stage. (B) Comparison of the expression of P. falciparum proteins detected in the three mosquito stage proteomes to the blood stage proteomes described previously [15]. (C) The percentage of proteins exclusively detected in only one proteome out of 6 different life cycle stage proteomes, i.e. ASX - asexual blood stages; GCT – gametocytes; GAM – gametes; OOC – oocysts; ODS - oocyst-derived sporozoites; SGS - salivary gland sporozoites.

Mentions: Protein samples derived from infected mosquito midguts and salivary glands were analyzed by nano–liquid chromatography tandem mass spectrometry (nLC-MS/MS) essentially as previously described [15]. The MS/MS spectra were searched against a combined database of all possible predicted tryptic peptides derived from all P. falciparum, human, and mosquito (Anopheles gambiae) proteins. The proteomic analysis of P. falciparum oocysts, oocyst-derived sporozoites, and salivary gland sporozoites resulted in a total of 4611 unique peptides mapping to 728 non redundant P. falciparum proteins; they are distributed over the three stages with 127, 450 and 477, respectively and depicted as a Venn diagram in Figure 1A. Identified tryptic peptides and corresponding Plasmodium proteins of the mosquito stages are provided as supplementary material (Table S1). In our previous analysis of infected human red blood cells we identified 741 asexual blood stage parasite proteins from a mixture of schizonts and trophozoites and an additional 931 gametocyte and 645 gamete proteins [15]. Merging these datasets with the proteomes of the mosquito stages resulted in the identification of 250 Plasmodium proteins (Table S1) that are specifically detected in mosquito stages and 809 proteins that are expressed only in the blood stages (Figure 1B). However, it is important to note that due to the incomplete nature of all proteome datasets, absence of proteins from one dataset may also be due to the limits of detection and not the actual absence of expression. Parasite samples derived from infected mosquitoes were considerably contaminated with mosquito proteins with total parasite protein fractions of 35% for ODS, 31% for SGS and for OOC only 11% of the sequenced proteins were parasite in origin. Therefore this relatively high degree of contamination resulted in overall lower numbers of proteins compared to our previous Plasmodium infected blood stage proteome study. In particular, only 127 P. falciparum proteins in a pool of 987 mosquito proteins were identified for the oocyst sample that presumably represents the more abundantly expressed parasite proteins. Therefore, further analysis of the identified proteins and additional functional analyses are mainly focused on the proteins identified in the ODS and SGS. In total, we analyzed six different stages of Plasmodium (both from this study and our previous work) and have identified a total of 1543 Plasmodium proteins. The proportion of ‘stage specific’ proteins in the different life cycle stages ranged from 12% (gametes) to 28% and the stage specificity of proteins in the mosquito stages ranged between 15–24% (Figure 1C).


Proteomic profiling of Plasmodium sporozoite maturation identifies new proteins essential for parasite development and infectivity.

Lasonder E, Janse CJ, van Gemert GJ, Mair GR, Vermunt AM, Douradinha BG, van Noort V, Huynen MA, Luty AJ, Kroeze H, Khan SM, Sauerwein RW, Waters AP, Mann M, Stunnenberg HG - PLoS Pathog. (2008)

Distribution of identified P. falciparum proteins over different life-cycle stages.(A) Venn diagram depicting the distribution of detected P. falciparum proteins over three different mosquito life-cycle stages (oocysts, oocyst-derived sporozoites and salivary gland sporozoites). Numbers represent the number of proteins, that are either shared between 2 or 3 stages (overlapping areas) or that are detected in a single stage. (B) Comparison of the expression of P. falciparum proteins detected in the three mosquito stage proteomes to the blood stage proteomes described previously [15]. (C) The percentage of proteins exclusively detected in only one proteome out of 6 different life cycle stage proteomes, i.e. ASX - asexual blood stages; GCT – gametocytes; GAM – gametes; OOC – oocysts; ODS - oocyst-derived sporozoites; SGS - salivary gland sporozoites.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2570797&req=5

ppat-1000195-g001: Distribution of identified P. falciparum proteins over different life-cycle stages.(A) Venn diagram depicting the distribution of detected P. falciparum proteins over three different mosquito life-cycle stages (oocysts, oocyst-derived sporozoites and salivary gland sporozoites). Numbers represent the number of proteins, that are either shared between 2 or 3 stages (overlapping areas) or that are detected in a single stage. (B) Comparison of the expression of P. falciparum proteins detected in the three mosquito stage proteomes to the blood stage proteomes described previously [15]. (C) The percentage of proteins exclusively detected in only one proteome out of 6 different life cycle stage proteomes, i.e. ASX - asexual blood stages; GCT – gametocytes; GAM – gametes; OOC – oocysts; ODS - oocyst-derived sporozoites; SGS - salivary gland sporozoites.
Mentions: Protein samples derived from infected mosquito midguts and salivary glands were analyzed by nano–liquid chromatography tandem mass spectrometry (nLC-MS/MS) essentially as previously described [15]. The MS/MS spectra were searched against a combined database of all possible predicted tryptic peptides derived from all P. falciparum, human, and mosquito (Anopheles gambiae) proteins. The proteomic analysis of P. falciparum oocysts, oocyst-derived sporozoites, and salivary gland sporozoites resulted in a total of 4611 unique peptides mapping to 728 non redundant P. falciparum proteins; they are distributed over the three stages with 127, 450 and 477, respectively and depicted as a Venn diagram in Figure 1A. Identified tryptic peptides and corresponding Plasmodium proteins of the mosquito stages are provided as supplementary material (Table S1). In our previous analysis of infected human red blood cells we identified 741 asexual blood stage parasite proteins from a mixture of schizonts and trophozoites and an additional 931 gametocyte and 645 gamete proteins [15]. Merging these datasets with the proteomes of the mosquito stages resulted in the identification of 250 Plasmodium proteins (Table S1) that are specifically detected in mosquito stages and 809 proteins that are expressed only in the blood stages (Figure 1B). However, it is important to note that due to the incomplete nature of all proteome datasets, absence of proteins from one dataset may also be due to the limits of detection and not the actual absence of expression. Parasite samples derived from infected mosquitoes were considerably contaminated with mosquito proteins with total parasite protein fractions of 35% for ODS, 31% for SGS and for OOC only 11% of the sequenced proteins were parasite in origin. Therefore this relatively high degree of contamination resulted in overall lower numbers of proteins compared to our previous Plasmodium infected blood stage proteome study. In particular, only 127 P. falciparum proteins in a pool of 987 mosquito proteins were identified for the oocyst sample that presumably represents the more abundantly expressed parasite proteins. Therefore, further analysis of the identified proteins and additional functional analyses are mainly focused on the proteins identified in the ODS and SGS. In total, we analyzed six different stages of Plasmodium (both from this study and our previous work) and have identified a total of 1543 Plasmodium proteins. The proportion of ‘stage specific’ proteins in the different life cycle stages ranged from 12% (gametes) to 28% and the stage specificity of proteins in the mosquito stages ranged between 15–24% (Figure 1C).

Bottom Line: The different sporozoite proteomes contain a large number of stage specific proteins whose annotation suggest an involvement in sporozoite maturation, motility, infection of the human host and associated metabolic adjustments.Analyses of proteins identified in the P. falciparum sporozoite proteomes by orthologous gene disruption in the rodent malaria parasite, P. berghei, revealed three previously uncharacterized Plasmodium proteins that appear to be essential for sporozoite development at distinct points of maturation in the mosquito.This study sheds light on the development and maturation of the malaria parasite in an Anopheles mosquito and also identifies proteins that may be essential for sporozoite infectivity to humans.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, NCMLS, Radboud University Nijmegen, Nijmegen, The Netherlands.

ABSTRACT
Plasmodium falciparum sporozoites that develop and mature inside an Anopheles mosquito initiate a malaria infection in humans. Here we report the first proteomic comparison of different parasite stages from the mosquito -- early and late oocysts containing midgut sporozoites, and the mature, infectious salivary gland sporozoites. Despite the morphological similarity between midgut and salivary gland sporozoites, their proteomes are markedly different, in agreement with their increase in hepatocyte infectivity. The different sporozoite proteomes contain a large number of stage specific proteins whose annotation suggest an involvement in sporozoite maturation, motility, infection of the human host and associated metabolic adjustments. Analyses of proteins identified in the P. falciparum sporozoite proteomes by orthologous gene disruption in the rodent malaria parasite, P. berghei, revealed three previously uncharacterized Plasmodium proteins that appear to be essential for sporozoite development at distinct points of maturation in the mosquito. This study sheds light on the development and maturation of the malaria parasite in an Anopheles mosquito and also identifies proteins that may be essential for sporozoite infectivity to humans.

Show MeSH
Related in: MedlinePlus