Role of the parafusin orthologue, PRP1, in microneme exocytosis and cell invasion in Toxoplasma gondii.
The association of PRP1, a Paramecium parafusin orthologue, with Toxoplasma gondii micronemes, now confirmed by immunoelectron microscopy, has here been studied in relation to exocytosis and cell invasion. PRP1 becomes labelled in vivo by inorganic 32P and is dephosphorylated when ethanol is used to stimulate Ca2+-dependent exocytosis of the micronemes. The ethanol Ca2+-stimulated exocytosis is accompanied by translocation of PRP1 and microneme content protein (MIC3) from the apical end of the parasite. Immunoblotting showed that PRP1 is redistributed inside the parasite, while microneme content is secreted. To study whether similar changes occur during cell invasion, quantitative microscopy was performed during secretion, invasion and exit (egress) from the host cell. Time-course experiments showed that fluorescence intensities of PRP1 and MIC3 immediately after invasion were reduced 10-fold compared to preinvasion levels, indicating that PRP1 translocation and microneme secretion accompanies invasion. MIC3 regained fluorescence intensity and apical distribution after 15 min, while PRP1 recovered after 1 h. Intensity of both proteins then increased throughout the parasite division period until host cell lysis, suggesting the need to secrete microneme proteins to egress. These studies suggest that PRP1 associated with the secretory vesicle scaffold serves an important role in Ca2+-regulated exocytosis and cell invasion.
Matthiesen
Steen H
SH
Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA.
Shenoy
Shailesh M
SM
Kim
Kami
K
Singer
Robert H
RH
Satir
Birgit H
BH
D000818:Animals; D002478:Cells, Cultured; D005089:Exocytosis / Q000502:physiology*; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism / Q000469:parasitology; D005947:Glucose / Q000031:analogs & derivatives / Q000378:metabolism; D006801:Humans; D010733:Phosphoglucomutase; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014122:Toxoplasma / Q000378:metabolism* / Q000472:pathogenicity* / Q000648:ultrastructure
D010750:Phosphoproteins; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; C488050:PGM1 protein, human; D010733:Phosphoglucomutase; D005947:Glucose