A highly sensitive FRET-based approach reveals secretion of the actin-binding protein toxofilin duringToxoplasma gondiiinfection

We have utilized a highly sensitive approach based on fluorescence resonance energy transfer (FRET) and β-lactamase (BLA) (Zlokarnik et al., 1998), which we adapted for the detection of Toxoplasma gondii secreted proteins. This assay revealed that the actin-binding protein toxofilin appears to be secreted into host cells during invasion. To determine the function of toxofilin during infection, we engineered a type I (RH strain) parasite with a targeted deletion of the toxofilin gene and compared the phenotypes of control and toxofilin knock-out (Δtxf) parasites in several in vitro assays, including invasion, growth, and gliding motility, and egress of the Δtxf parasites, as well as F-actin staining, phagocytosis and migration of cells infected with Δtxf parasites or wild-type controls. Despite its apparent secretion into host cells and its ability to bind to and modulate host actin, we observed that toxofilin does not appear to play a role in these processes, under the conditions we examined, and we report these findings here.