Open AccessA highly sensitive FRET‐based approach reveals secretion of the actin‐binding protein toxofilin during Toxoplasma gondii infection
Author(s) -
Lodoen Melissa B.,
Gerke Christiane,
Boothroyd John C.
Publication year - 2010
Publication title -
cellular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.542
H-Index - 138
eISSN - 1462-5822
pISSN - 1462-5814
DOI - 10.1111/j.1462-5822.2009.01378.x
Subject(s) - toxoplasma gondii , biology , förster resonance energy transfer , secretion , actin , virology , microbiology and biotechnology , computational biology , immunology , fluorescence , antibody , biochemistry , physics , quantum mechanics
Summary We have utilized a highly sensitive approach based on fluorescence resonance energy transfer (FRET) and β‐lactamase (BLA), 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 knockout (Δ txf ) parasites in several in vitro assays, including invasion, growth, gliding motility, and egress of the Δ txf parasites, as well as F‐actin staining, phagocytosis and migration of cells infected with Δ txf 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.