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Trafficking of plasmepsin II to the food vacuole of the malaria parasite Plasmodium falciparum.

Klemba M, Beatty W, Gluzman I, Goldberg DE - J. Cell Biol. (2004)

Bottom Line: A family of aspartic proteases, the plasmepsins (PMs), plays a key role in the degradation of hemoglobin in the Plasmodium falciparum food vacuole.To study the trafficking of proPM II, we have modified the chromosomal PM II gene in P. falciparum to encode a proPM II-GFP chimera.Our data support a model whereby proPM II is transported through the secretory system to cytostomal vacuoles and then is carried along with its substrate hemoglobin to the food vacuole where it is proteolytically processed to mature PM II.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Microbiology, Washington University School of Medicine, 660 S. Euclid Ave., Box 8230, St. Louis, MO 63110, USA.

ABSTRACT
A family of aspartic proteases, the plasmepsins (PMs), plays a key role in the degradation of hemoglobin in the Plasmodium falciparum food vacuole. To study the trafficking of proPM II, we have modified the chromosomal PM II gene in P. falciparum to encode a proPM II-GFP chimera. By taking advantage of green fluorescent protein fluorescence in live parasites, the ultrastructural resolution of immunoelectron microscopy, and inhibitors of trafficking and PM maturation, we have investigated the biosynthetic path leading to mature PM II in the food vacuole. Our data support a model whereby proPM II is transported through the secretory system to cytostomal vacuoles and then is carried along with its substrate hemoglobin to the food vacuole where it is proteolytically processed to mature PM II.

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GFP fluorescence in live B7 parasites. (A) A trophozoite exhibiting GFP fluorescence in the food vacuole (arrowhead) and in a perinuclear ring (arrow). Fluorescence from the nuclear stain Hoechst 33342 is pseudocolored red. (B) A trophozoite with a fluorescent food vacuole (arrowhead) and a bright fluorescent spot (arrow) that lies at the periphery of the parasite. (C) A mature parasite displaying a large fluorescent food vacuole. Note the absence of fluorescence outside of the food vacuole. Bar, 2 μm.
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fig4: GFP fluorescence in live B7 parasites. (A) A trophozoite exhibiting GFP fluorescence in the food vacuole (arrowhead) and in a perinuclear ring (arrow). Fluorescence from the nuclear stain Hoechst 33342 is pseudocolored red. (B) A trophozoite with a fluorescent food vacuole (arrowhead) and a bright fluorescent spot (arrow) that lies at the periphery of the parasite. (C) A mature parasite displaying a large fluorescent food vacuole. Note the absence of fluorescence outside of the food vacuole. Bar, 2 μm.

Mentions: The distribution of GFP in live B7 parasites was followed over the course of the ∼44 h intraerythrocytic asexual reproductive cycle by epifluorescence microscopy. The food vacuole of trophozoites and schizonts contained abundant GFP (Fig. 4), which indicates that it is transported along with PM II to this organelle. In some parasites, particularly younger trophozoites, GFP fluorescence was observed in a perinuclear ring-like structure adjacent to the food vacuole (Fig. 4 A). The perinuclear distribution of GFP suggested that it might be within the nuclear envelope. Further analysis of this compartment in the presence of BFA is described in the next section. Also in trophozoites, some GFP fluorescence was concentrated in a small number of spots or foci (2.4 ± 1.2 foci per trophozoite, range 0–5, n = 35) that often resided at the periphery of the parasite (Fig. 4 B). In parasites undergoing nuclear division (schizonts), fluorescence was limited to the food vacuole (Fig. 4 C). No fluorescence was observed in association with individual merozoites in segmented schizonts or in ring-stage parasites. Similar GFP distributions were observed with the cloned parasite lines C9 and F4 (unpublished data).


Trafficking of plasmepsin II to the food vacuole of the malaria parasite Plasmodium falciparum.

Klemba M, Beatty W, Gluzman I, Goldberg DE - J. Cell Biol. (2004)

GFP fluorescence in live B7 parasites. (A) A trophozoite exhibiting GFP fluorescence in the food vacuole (arrowhead) and in a perinuclear ring (arrow). Fluorescence from the nuclear stain Hoechst 33342 is pseudocolored red. (B) A trophozoite with a fluorescent food vacuole (arrowhead) and a bright fluorescent spot (arrow) that lies at the periphery of the parasite. (C) A mature parasite displaying a large fluorescent food vacuole. Note the absence of fluorescence outside of the food vacuole. Bar, 2 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171955&req=5

fig4: GFP fluorescence in live B7 parasites. (A) A trophozoite exhibiting GFP fluorescence in the food vacuole (arrowhead) and in a perinuclear ring (arrow). Fluorescence from the nuclear stain Hoechst 33342 is pseudocolored red. (B) A trophozoite with a fluorescent food vacuole (arrowhead) and a bright fluorescent spot (arrow) that lies at the periphery of the parasite. (C) A mature parasite displaying a large fluorescent food vacuole. Note the absence of fluorescence outside of the food vacuole. Bar, 2 μm.
Mentions: The distribution of GFP in live B7 parasites was followed over the course of the ∼44 h intraerythrocytic asexual reproductive cycle by epifluorescence microscopy. The food vacuole of trophozoites and schizonts contained abundant GFP (Fig. 4), which indicates that it is transported along with PM II to this organelle. In some parasites, particularly younger trophozoites, GFP fluorescence was observed in a perinuclear ring-like structure adjacent to the food vacuole (Fig. 4 A). The perinuclear distribution of GFP suggested that it might be within the nuclear envelope. Further analysis of this compartment in the presence of BFA is described in the next section. Also in trophozoites, some GFP fluorescence was concentrated in a small number of spots or foci (2.4 ± 1.2 foci per trophozoite, range 0–5, n = 35) that often resided at the periphery of the parasite (Fig. 4 B). In parasites undergoing nuclear division (schizonts), fluorescence was limited to the food vacuole (Fig. 4 C). No fluorescence was observed in association with individual merozoites in segmented schizonts or in ring-stage parasites. Similar GFP distributions were observed with the cloned parasite lines C9 and F4 (unpublished data).

Bottom Line: A family of aspartic proteases, the plasmepsins (PMs), plays a key role in the degradation of hemoglobin in the Plasmodium falciparum food vacuole.To study the trafficking of proPM II, we have modified the chromosomal PM II gene in P. falciparum to encode a proPM II-GFP chimera.Our data support a model whereby proPM II is transported through the secretory system to cytostomal vacuoles and then is carried along with its substrate hemoglobin to the food vacuole where it is proteolytically processed to mature PM II.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Microbiology, Washington University School of Medicine, 660 S. Euclid Ave., Box 8230, St. Louis, MO 63110, USA.

ABSTRACT
A family of aspartic proteases, the plasmepsins (PMs), plays a key role in the degradation of hemoglobin in the Plasmodium falciparum food vacuole. To study the trafficking of proPM II, we have modified the chromosomal PM II gene in P. falciparum to encode a proPM II-GFP chimera. By taking advantage of green fluorescent protein fluorescence in live parasites, the ultrastructural resolution of immunoelectron microscopy, and inhibitors of trafficking and PM maturation, we have investigated the biosynthetic path leading to mature PM II in the food vacuole. Our data support a model whereby proPM II is transported through the secretory system to cytostomal vacuoles and then is carried along with its substrate hemoglobin to the food vacuole where it is proteolytically processed to mature PM II.

Show MeSH
Related in: MedlinePlus