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Host cell egress and invasion induce marked relocations of glycolytic enzymes in Toxoplasma gondii tachyzoites.

Pomel S, Luk FC, Beckers CJ - PLoS Pathog. (2008)

Bottom Line: Translocation of glycolytic enzymes to and from the Toxoplasma pellicle appears to occur in response to changes in extracellular [K(+)] experienced during egress and invasion, a signal that requires changes of [Ca(2+)](c) in the parasite during egress.Enzyme translocation is, however, not dependent on either F-actin or intact microtubules.We propose that this ability allows Toxoplasma to optimize ATP delivery to those cellular processes that are most critical for survival outside host cells and those required for growth and replication of intracellular parasites.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell & Developmental Biology, University of North Carolina, Chapel Hill, North Carolina, USA.

ABSTRACT
Apicomplexan parasites are dependent on an F-actin and myosin-based motility system for their invasion into and escape from animal host cells, as well as for their general motility. In Toxoplasma gondii and Plasmodium species, the actin filaments and myosin motor required for this process are located in a narrow space between the parasite plasma membrane and the underlying inner membrane complex, a set of flattened cisternae that covers most the cytoplasmic face of the plasma membrane. Here we show that the energy required for Toxoplasma motility is derived mostly, if not entirely, from glycolysis and lactic acid production. We also demonstrate that the glycolytic enzymes of Toxoplasma tachyzoites undergo a striking relocation from the parasites' cytoplasm to their pellicles upon Toxoplasma egress from host cells. Specifically, it appears that the glycolytic enzymes are translocated to the cytoplasmic face of the inner membrane complex as well as to the space between the plasma membrane and inner membrane complex. The glycolytic enzymes remain pellicle-associated during extended incubations of parasites in the extracellular milieu and do not revert to a cytoplasmic location until well after parasites have completed invasion of new host cells. Translocation of glycolytic enzymes to and from the Toxoplasma pellicle appears to occur in response to changes in extracellular [K(+)] experienced during egress and invasion, a signal that requires changes of [Ca(2+)](c) in the parasite during egress. Enzyme translocation is, however, not dependent on either F-actin or intact microtubules. Our observations indicate that Toxoplasma gondii is capable of relocating its main source of energy between its cytoplasm and pellicle in response to exit from or entry into host cells. We propose that this ability allows Toxoplasma to optimize ATP delivery to those cellular processes that are most critical for survival outside host cells and those required for growth and replication of intracellular parasites.

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Related in: MedlinePlus

Quantitative analysis of aldolase-1 distribution in Toxoplasma.Freshly egressed parasites expressing myc-aldolase 1 were allowed to attach to HFF monolayers for 5 minutes at 37°C in complete medium. Non-attached parasites were washed off and the incubation continued in complete medium at 37°C for the indicated times. (A) The distribution of aldolase-1 30 minutes and 4 hours after invasion of host cells. Overlays are shown of myc-aldolase-1 (mALD1, green), the IMC protein IMC1 (red). Overlay pictures of DIC and DAPI blue) are shown on the right. Parasite-infected cells were fixed in −20°C methanol. Images of the individual channels are shown in Figure S4. Bars = 2 µm. (B) A quantitative analysis of the results is shown. Data represent averages of 32 fields counted over 4 samples. Error bars indicate the standard deviation.
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ppat-1000188-g006: Quantitative analysis of aldolase-1 distribution in Toxoplasma.Freshly egressed parasites expressing myc-aldolase 1 were allowed to attach to HFF monolayers for 5 minutes at 37°C in complete medium. Non-attached parasites were washed off and the incubation continued in complete medium at 37°C for the indicated times. (A) The distribution of aldolase-1 30 minutes and 4 hours after invasion of host cells. Overlays are shown of myc-aldolase-1 (mALD1, green), the IMC protein IMC1 (red). Overlay pictures of DIC and DAPI blue) are shown on the right. Parasite-infected cells were fixed in −20°C methanol. Images of the individual channels are shown in Figure S4. Bars = 2 µm. (B) A quantitative analysis of the results is shown. Data represent averages of 32 fields counted over 4 samples. Error bars indicate the standard deviation.

Mentions: As Toxoplasma aldolase-1 reverts to a smooth cytoplasmic distribution at some point after host cell invasion we first tested whether this occurred during or shortly after host cell invasion, or if it occurred at a later stage. We therefore allowed freshly isolated extracellular parasites to interact with new host cells for 2 minutes and monitored the aldolase-1 distribution immediately after this incubation and after various lengths of incubation at 37°C. Extracellular and intracellular parasites could be distinguished by the inability of the latter to be recognized by an antibody to the surface antigen SAG1 in intact host cells. After short incubations, numerous parasites were captured in the process of invading host cells as judged by the fact that only a part of their surface reacted with the SAG1 antiserum (Figure 5B). In these parasites, the aldolase-1 remains located at the parasite periphery. The aldolase-1 distribution was also not affected in parasites that had just completed host cell invasion (Figure 6A). The peripheral location of the aldolase-1 was maintained in a large fraction (74±5%) of parasites at 30 minutes after invasion but decreased to less than 10% after 4 hours (Figure 6B) and was no longer detectable after parasites had undergone one round of replication (Figure 5C; Figure S4). The cytoplasmic distribution of aldolase-1 was maintained as long as the parasites remained inside host cells, irrespective of the number of parasites per vacuole or the stage of parasite replication (Figure 5C; Figure S4).


Host cell egress and invasion induce marked relocations of glycolytic enzymes in Toxoplasma gondii tachyzoites.

Pomel S, Luk FC, Beckers CJ - PLoS Pathog. (2008)

Quantitative analysis of aldolase-1 distribution in Toxoplasma.Freshly egressed parasites expressing myc-aldolase 1 were allowed to attach to HFF monolayers for 5 minutes at 37°C in complete medium. Non-attached parasites were washed off and the incubation continued in complete medium at 37°C for the indicated times. (A) The distribution of aldolase-1 30 minutes and 4 hours after invasion of host cells. Overlays are shown of myc-aldolase-1 (mALD1, green), the IMC protein IMC1 (red). Overlay pictures of DIC and DAPI blue) are shown on the right. Parasite-infected cells were fixed in −20°C methanol. Images of the individual channels are shown in Figure S4. Bars = 2 µm. (B) A quantitative analysis of the results is shown. Data represent averages of 32 fields counted over 4 samples. Error bars indicate the standard deviation.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2563030&req=5

ppat-1000188-g006: Quantitative analysis of aldolase-1 distribution in Toxoplasma.Freshly egressed parasites expressing myc-aldolase 1 were allowed to attach to HFF monolayers for 5 minutes at 37°C in complete medium. Non-attached parasites were washed off and the incubation continued in complete medium at 37°C for the indicated times. (A) The distribution of aldolase-1 30 minutes and 4 hours after invasion of host cells. Overlays are shown of myc-aldolase-1 (mALD1, green), the IMC protein IMC1 (red). Overlay pictures of DIC and DAPI blue) are shown on the right. Parasite-infected cells were fixed in −20°C methanol. Images of the individual channels are shown in Figure S4. Bars = 2 µm. (B) A quantitative analysis of the results is shown. Data represent averages of 32 fields counted over 4 samples. Error bars indicate the standard deviation.
Mentions: As Toxoplasma aldolase-1 reverts to a smooth cytoplasmic distribution at some point after host cell invasion we first tested whether this occurred during or shortly after host cell invasion, or if it occurred at a later stage. We therefore allowed freshly isolated extracellular parasites to interact with new host cells for 2 minutes and monitored the aldolase-1 distribution immediately after this incubation and after various lengths of incubation at 37°C. Extracellular and intracellular parasites could be distinguished by the inability of the latter to be recognized by an antibody to the surface antigen SAG1 in intact host cells. After short incubations, numerous parasites were captured in the process of invading host cells as judged by the fact that only a part of their surface reacted with the SAG1 antiserum (Figure 5B). In these parasites, the aldolase-1 remains located at the parasite periphery. The aldolase-1 distribution was also not affected in parasites that had just completed host cell invasion (Figure 6A). The peripheral location of the aldolase-1 was maintained in a large fraction (74±5%) of parasites at 30 minutes after invasion but decreased to less than 10% after 4 hours (Figure 6B) and was no longer detectable after parasites had undergone one round of replication (Figure 5C; Figure S4). The cytoplasmic distribution of aldolase-1 was maintained as long as the parasites remained inside host cells, irrespective of the number of parasites per vacuole or the stage of parasite replication (Figure 5C; Figure S4).

Bottom Line: Translocation of glycolytic enzymes to and from the Toxoplasma pellicle appears to occur in response to changes in extracellular [K(+)] experienced during egress and invasion, a signal that requires changes of [Ca(2+)](c) in the parasite during egress.Enzyme translocation is, however, not dependent on either F-actin or intact microtubules.We propose that this ability allows Toxoplasma to optimize ATP delivery to those cellular processes that are most critical for survival outside host cells and those required for growth and replication of intracellular parasites.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell & Developmental Biology, University of North Carolina, Chapel Hill, North Carolina, USA.

ABSTRACT
Apicomplexan parasites are dependent on an F-actin and myosin-based motility system for their invasion into and escape from animal host cells, as well as for their general motility. In Toxoplasma gondii and Plasmodium species, the actin filaments and myosin motor required for this process are located in a narrow space between the parasite plasma membrane and the underlying inner membrane complex, a set of flattened cisternae that covers most the cytoplasmic face of the plasma membrane. Here we show that the energy required for Toxoplasma motility is derived mostly, if not entirely, from glycolysis and lactic acid production. We also demonstrate that the glycolytic enzymes of Toxoplasma tachyzoites undergo a striking relocation from the parasites' cytoplasm to their pellicles upon Toxoplasma egress from host cells. Specifically, it appears that the glycolytic enzymes are translocated to the cytoplasmic face of the inner membrane complex as well as to the space between the plasma membrane and inner membrane complex. The glycolytic enzymes remain pellicle-associated during extended incubations of parasites in the extracellular milieu and do not revert to a cytoplasmic location until well after parasites have completed invasion of new host cells. Translocation of glycolytic enzymes to and from the Toxoplasma pellicle appears to occur in response to changes in extracellular [K(+)] experienced during egress and invasion, a signal that requires changes of [Ca(2+)](c) in the parasite during egress. Enzyme translocation is, however, not dependent on either F-actin or intact microtubules. Our observations indicate that Toxoplasma gondii is capable of relocating its main source of energy between its cytoplasm and pellicle in response to exit from or entry into host cells. We propose that this ability allows Toxoplasma to optimize ATP delivery to those cellular processes that are most critical for survival outside host cells and those required for growth and replication of intracellular parasites.

Show MeSH
Related in: MedlinePlus