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Phosphatidylinositol 3-monophosphate is involved in toxoplasma apicoplast biogenesis.

Tawk L, Dubremetz JF, Montcourrier P, Chicanne G, Merezegue F, Richard V, Payrastre B, Meissner M, Vial HJ, Roy C, Wengelnik K, Lebrun M - PLoS Pathog. (2011)

Bottom Line: Imaging of PI3P in T. gondii showed that the lipid was associated with the apicoplast and apicoplast protein-shuttling vesicles.These findings point to an unexpected implication for this ubiquitous lipid and open new perspectives on how nuclear encoded proteins traffic to the apicoplast.This study also highlights the possibility of developing specific pharmacological inhibitors of the parasite PI3-kinase as novel anti-apicomplexan drugs.

View Article: PubMed Central - PubMed

Affiliation: UMR 5235 CNRS, Université Montpellier 1 & 2, Montpellier, France.

ABSTRACT
Apicomplexan parasites cause devastating diseases including malaria and toxoplasmosis. They harbour a plastid-like, non-photosynthetic organelle of algal origin, the apicoplast, which fulfils critical functions for parasite survival. Because of its essential and original metabolic pathways, the apicoplast has become a target for the development of new anti-apicomplexan drugs. Here we show that the lipid phosphatidylinositol 3-monophosphate (PI3P) is involved in apicoplast biogenesis in Toxoplasma gondii. In yeast and mammalian cells, PI3P is concentrated on early endosomes and regulates trafficking of endosomal compartments. Imaging of PI3P in T. gondii showed that the lipid was associated with the apicoplast and apicoplast protein-shuttling vesicles. Interference with regular PI3P function by over-expression of a PI3P specific binding module in the parasite led to the accumulation of vesicles containing apicoplast peripheral membrane proteins around the apicoplast and, ultimately, to the loss of the organelle. Accordingly, inhibition of the PI3P-synthesising kinase interfered with apicoplast biogenesis. These findings point to an unexpected implication for this ubiquitous lipid and open new perspectives on how nuclear encoded proteins traffic to the apicoplast. This study also highlights the possibility of developing specific pharmacological inhibitors of the parasite PI3-kinase as novel anti-apicomplexan drugs.

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Related in: MedlinePlus

Electron-lucent vesicles accumulate upon ddFYVE expression and contain apicoplast peripheral membrane proteins and PI3P.Immuno-EM analysis of ddFYVE expressing parasites transiently expressing V5-FtsH1 (A, B, C) or APT1-HA (D, E) fixed 4 h (A, B, C) or 24 h (D, E) after 1 µM Shield-1 addition. The scale is the same for all five images. (A, B, C) Double immuno-EM with anti-GFP (5 nm gold, green arrows) and anti-V5 (10 nm gold, black arrows) antibodies (a: apicoplast; m: microneme). (A) shows double labelling of the apicoplast outer membranes, (B) of a small dense vesicle, (C) of an electron lucent vesicle. (D) Immuno-EM with anti-HA antibodies. (E) Double immuno-EM with anti-GFP (10 nm gold, green arrows) and anti-HA (5 nm gold, black arrows) antibodies.
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ppat-1001286-g007: Electron-lucent vesicles accumulate upon ddFYVE expression and contain apicoplast peripheral membrane proteins and PI3P.Immuno-EM analysis of ddFYVE expressing parasites transiently expressing V5-FtsH1 (A, B, C) or APT1-HA (D, E) fixed 4 h (A, B, C) or 24 h (D, E) after 1 µM Shield-1 addition. The scale is the same for all five images. (A, B, C) Double immuno-EM with anti-GFP (5 nm gold, green arrows) and anti-V5 (10 nm gold, black arrows) antibodies (a: apicoplast; m: microneme). (A) shows double labelling of the apicoplast outer membranes, (B) of a small dense vesicle, (C) of an electron lucent vesicle. (D) Immuno-EM with anti-HA antibodies. (E) Double immuno-EM with anti-GFP (10 nm gold, green arrows) and anti-HA (5 nm gold, black arrows) antibodies.

Mentions: On dividing apicoplasts, we detected an accumulation of ATrx1 at the base of the V-shaped intermediates that co-localized with ddFYVE label (Figure 6C). Double IEM analysis was then conducted on ddFYVE/FtsH1 or ddFYVE/APT1 parasites to localize outermost apicoplast proteins and PI3P simultaneously. After four hours of Shield-1 treatment, anti-V5 and anti-GFP antibodies showed the simultaneous presence of FtsH1 (black arrows) and PI3P (green arrows) on both the outermost membrane of the apicoplast and on dense-core vesicles of ca. 0.1 µm in its vicinity (Figure 7A, B, C), as in untreated control cells ([33] and data not shown) demonstrating that PI3P was present on vesicles containing outermost apicoplast membrane proteins. In addition, we also detected FtsH1 on the larger, ddFYVE positive lucent vesicles. After one day of Shield-1 treatment, most of the vesicles containing APT1 (black arrows) (Figure 6D) and FtsH1 (data not shown) were the large electron lucent vesicles, also shown to be positive for the ddFYVE marker (green arrows) by double IEM (Figure 7E).


Phosphatidylinositol 3-monophosphate is involved in toxoplasma apicoplast biogenesis.

Tawk L, Dubremetz JF, Montcourrier P, Chicanne G, Merezegue F, Richard V, Payrastre B, Meissner M, Vial HJ, Roy C, Wengelnik K, Lebrun M - PLoS Pathog. (2011)

Electron-lucent vesicles accumulate upon ddFYVE expression and contain apicoplast peripheral membrane proteins and PI3P.Immuno-EM analysis of ddFYVE expressing parasites transiently expressing V5-FtsH1 (A, B, C) or APT1-HA (D, E) fixed 4 h (A, B, C) or 24 h (D, E) after 1 µM Shield-1 addition. The scale is the same for all five images. (A, B, C) Double immuno-EM with anti-GFP (5 nm gold, green arrows) and anti-V5 (10 nm gold, black arrows) antibodies (a: apicoplast; m: microneme). (A) shows double labelling of the apicoplast outer membranes, (B) of a small dense vesicle, (C) of an electron lucent vesicle. (D) Immuno-EM with anti-HA antibodies. (E) Double immuno-EM with anti-GFP (10 nm gold, green arrows) and anti-HA (5 nm gold, black arrows) antibodies.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3040667&req=5

ppat-1001286-g007: Electron-lucent vesicles accumulate upon ddFYVE expression and contain apicoplast peripheral membrane proteins and PI3P.Immuno-EM analysis of ddFYVE expressing parasites transiently expressing V5-FtsH1 (A, B, C) or APT1-HA (D, E) fixed 4 h (A, B, C) or 24 h (D, E) after 1 µM Shield-1 addition. The scale is the same for all five images. (A, B, C) Double immuno-EM with anti-GFP (5 nm gold, green arrows) and anti-V5 (10 nm gold, black arrows) antibodies (a: apicoplast; m: microneme). (A) shows double labelling of the apicoplast outer membranes, (B) of a small dense vesicle, (C) of an electron lucent vesicle. (D) Immuno-EM with anti-HA antibodies. (E) Double immuno-EM with anti-GFP (10 nm gold, green arrows) and anti-HA (5 nm gold, black arrows) antibodies.
Mentions: On dividing apicoplasts, we detected an accumulation of ATrx1 at the base of the V-shaped intermediates that co-localized with ddFYVE label (Figure 6C). Double IEM analysis was then conducted on ddFYVE/FtsH1 or ddFYVE/APT1 parasites to localize outermost apicoplast proteins and PI3P simultaneously. After four hours of Shield-1 treatment, anti-V5 and anti-GFP antibodies showed the simultaneous presence of FtsH1 (black arrows) and PI3P (green arrows) on both the outermost membrane of the apicoplast and on dense-core vesicles of ca. 0.1 µm in its vicinity (Figure 7A, B, C), as in untreated control cells ([33] and data not shown) demonstrating that PI3P was present on vesicles containing outermost apicoplast membrane proteins. In addition, we also detected FtsH1 on the larger, ddFYVE positive lucent vesicles. After one day of Shield-1 treatment, most of the vesicles containing APT1 (black arrows) (Figure 6D) and FtsH1 (data not shown) were the large electron lucent vesicles, also shown to be positive for the ddFYVE marker (green arrows) by double IEM (Figure 7E).

Bottom Line: Imaging of PI3P in T. gondii showed that the lipid was associated with the apicoplast and apicoplast protein-shuttling vesicles.These findings point to an unexpected implication for this ubiquitous lipid and open new perspectives on how nuclear encoded proteins traffic to the apicoplast.This study also highlights the possibility of developing specific pharmacological inhibitors of the parasite PI3-kinase as novel anti-apicomplexan drugs.

View Article: PubMed Central - PubMed

Affiliation: UMR 5235 CNRS, Université Montpellier 1 & 2, Montpellier, France.

ABSTRACT
Apicomplexan parasites cause devastating diseases including malaria and toxoplasmosis. They harbour a plastid-like, non-photosynthetic organelle of algal origin, the apicoplast, which fulfils critical functions for parasite survival. Because of its essential and original metabolic pathways, the apicoplast has become a target for the development of new anti-apicomplexan drugs. Here we show that the lipid phosphatidylinositol 3-monophosphate (PI3P) is involved in apicoplast biogenesis in Toxoplasma gondii. In yeast and mammalian cells, PI3P is concentrated on early endosomes and regulates trafficking of endosomal compartments. Imaging of PI3P in T. gondii showed that the lipid was associated with the apicoplast and apicoplast protein-shuttling vesicles. Interference with regular PI3P function by over-expression of a PI3P specific binding module in the parasite led to the accumulation of vesicles containing apicoplast peripheral membrane proteins around the apicoplast and, ultimately, to the loss of the organelle. Accordingly, inhibition of the PI3P-synthesising kinase interfered with apicoplast biogenesis. These findings point to an unexpected implication for this ubiquitous lipid and open new perspectives on how nuclear encoded proteins traffic to the apicoplast. This study also highlights the possibility of developing specific pharmacological inhibitors of the parasite PI3-kinase as novel anti-apicomplexan drugs.

Show MeSH
Related in: MedlinePlus