Toxofilin, a novel actin-binding protein from Toxoplasma gondii, sequesters actin monomers and caps actin filaments.
Toxoplasma gondii relies on its actin cytoskeleton to glide and enter its host cell. However, T. gondii tachyzoites are known to display a strikingly low amount of actin filaments, which suggests that sequestration of actin monomers could play a key role in parasite actin dynamics. We isolated a 27-kDa tachyzoite protein on the basis of its ability to bind muscle G-actin and demonstrated that it interacts with parasite G-actin. Cloning and sequence analysis of the gene coding for this protein, which we named Toxofilin, showed that it is a novel actin-binding protein. In in vitro assays, Toxofilin not only bound to G-actin and inhibited actin polymerization as an actin-sequestering protein but also slowed down F-actin disassembly through a filament end capping activity. In addition, when green fluorescent protein-tagged Toxofilin was overexpressed in mammalian nonmuscle cells, the dynamics of actin stress fibers was drastically impaired, whereas green fluorescent protein-Toxofilin copurified with G-actin. Finally, in motile parasites, during gliding or host cell entry, Toxofilin was localized in the entire cytoplasm, including the rear end of the parasite, whereas in intracellular tachyzoites, especially before they exit from the parasitophorous vacuole of their host cell, Toxofilin was found to be restricted to the apical end.
Poupel
O
O
Laboratoire de Biochimie et Biologie Moléculaire des Insectes, Unité de Recherche Associée, Centre National de la Recherche Scientifique 1960, 75724 Paris Cedex 15, France.
Boleti
H
H
Axisa
S
S
Couture-Tosi
E
E
Tardieux
I
I
D051344:Actin Capping Proteins; D000199:Actins / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D003600:Cytosol / Q000378:metabolism; D016054:DNA, Protozoan; D006367:HeLa Cells; D006801:Humans; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014122:Toxoplasma / Q000235:genetics / Q000378:metabolism*
D051344:Actin Capping Proteins; D000199:Actins; D016054:DNA, Protozoan; D008840:Microfilament Proteins; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; C406004:toxofilin protein, Toxoplasma gondii