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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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Distribution of aldolase-1 in extracellular, invading, and intracellular Toxoplasma.(A–D) Immunofluorescence microscopy was used to determine the distribution of myc-tagged aldolase-1 (mALD1, green) in (A, D) extracellular parasites, (B) invading parasites, and (C) intracellular parasites immediately after invasion or after 1, 2, 3, and 5 rounds of replication. Panel D demonstrates the distribution of aldolase-1 in extracellular parasites caught in the process of endodyogeny. Note the selective association of the enzyme with only the IMC of the mother parasite and not that of the immature daughter cells. All panels show parasites and parasite-infected cells fixed in −20°C methanol. In panels A and D, parasites were counterstained with antibodies to the membrane skeleton protein IMC1 (red). In panel B, parasites expressing myc-aldolase-1 were allowed to interact with HFF cells for 2 minutes at 37°C, followed by the decoration of extracellular parts of the parasites with anti-SAG1 antiserum (red), which was in turn followed by cell fixation and permeabilization and staining with anti-myc monoclonal antibody (green). Parasite nuclei were visualized using DAPI (blue). Bars = 2 µm.
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ppat-1000188-g005: Distribution of aldolase-1 in extracellular, invading, and intracellular Toxoplasma.(A–D) Immunofluorescence microscopy was used to determine the distribution of myc-tagged aldolase-1 (mALD1, green) in (A, D) extracellular parasites, (B) invading parasites, and (C) intracellular parasites immediately after invasion or after 1, 2, 3, and 5 rounds of replication. Panel D demonstrates the distribution of aldolase-1 in extracellular parasites caught in the process of endodyogeny. Note the selective association of the enzyme with only the IMC of the mother parasite and not that of the immature daughter cells. All panels show parasites and parasite-infected cells fixed in −20°C methanol. In panels A and D, parasites were counterstained with antibodies to the membrane skeleton protein IMC1 (red). In panel B, parasites expressing myc-aldolase-1 were allowed to interact with HFF cells for 2 minutes at 37°C, followed by the decoration of extracellular parts of the parasites with anti-SAG1 antiserum (red), which was in turn followed by cell fixation and permeabilization and staining with anti-myc monoclonal antibody (green). Parasite nuclei were visualized using DAPI (blue). Bars = 2 µm.

Mentions: To further assure ourselves that we were observing a relocation of aldolase-1, we repeated the experiments in Figure 3A with parasites expressing myc-tagged aldolase-1. As can be seen in Figure 5A and C, myc-aldolase-1 displays a similar change in distribution between intracellular and extracellular parasites. It is interesting to note that we did not observe an overlap between the aldolase-1 or myc-aldolase-1 signal with that of the microneme protein MIC2 (Figure S3) which stands in contrast to the previous observations [1].


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

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

Distribution of aldolase-1 in extracellular, invading, and intracellular Toxoplasma.(A–D) Immunofluorescence microscopy was used to determine the distribution of myc-tagged aldolase-1 (mALD1, green) in (A, D) extracellular parasites, (B) invading parasites, and (C) intracellular parasites immediately after invasion or after 1, 2, 3, and 5 rounds of replication. Panel D demonstrates the distribution of aldolase-1 in extracellular parasites caught in the process of endodyogeny. Note the selective association of the enzyme with only the IMC of the mother parasite and not that of the immature daughter cells. All panels show parasites and parasite-infected cells fixed in −20°C methanol. In panels A and D, parasites were counterstained with antibodies to the membrane skeleton protein IMC1 (red). In panel B, parasites expressing myc-aldolase-1 were allowed to interact with HFF cells for 2 minutes at 37°C, followed by the decoration of extracellular parts of the parasites with anti-SAG1 antiserum (red), which was in turn followed by cell fixation and permeabilization and staining with anti-myc monoclonal antibody (green). Parasite nuclei were visualized using DAPI (blue). Bars = 2 µm.
© Copyright Policy
Related In: Results  -  Collection

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

ppat-1000188-g005: Distribution of aldolase-1 in extracellular, invading, and intracellular Toxoplasma.(A–D) Immunofluorescence microscopy was used to determine the distribution of myc-tagged aldolase-1 (mALD1, green) in (A, D) extracellular parasites, (B) invading parasites, and (C) intracellular parasites immediately after invasion or after 1, 2, 3, and 5 rounds of replication. Panel D demonstrates the distribution of aldolase-1 in extracellular parasites caught in the process of endodyogeny. Note the selective association of the enzyme with only the IMC of the mother parasite and not that of the immature daughter cells. All panels show parasites and parasite-infected cells fixed in −20°C methanol. In panels A and D, parasites were counterstained with antibodies to the membrane skeleton protein IMC1 (red). In panel B, parasites expressing myc-aldolase-1 were allowed to interact with HFF cells for 2 minutes at 37°C, followed by the decoration of extracellular parts of the parasites with anti-SAG1 antiserum (red), which was in turn followed by cell fixation and permeabilization and staining with anti-myc monoclonal antibody (green). Parasite nuclei were visualized using DAPI (blue). Bars = 2 µm.
Mentions: To further assure ourselves that we were observing a relocation of aldolase-1, we repeated the experiments in Figure 3A with parasites expressing myc-tagged aldolase-1. As can be seen in Figure 5A and C, myc-aldolase-1 displays a similar change in distribution between intracellular and extracellular parasites. It is interesting to note that we did not observe an overlap between the aldolase-1 or myc-aldolase-1 signal with that of the microneme protein MIC2 (Figure S3) which stands in contrast to the previous observations [1].

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