A highly sensitive FRET-based approach reveals secretion of the actin-binding protein toxofilin during Toxoplasma gondii infection
Article first published online: 2 SEP 2009
© 2009 Blackwell Publishing Ltd
Volume 12, Issue 1, pages 55–66, January 2010
How to Cite
Lodoen, M. B., Gerke, C. and Boothroyd, J. C. (2010), A highly sensitive FRET-based approach reveals secretion of the actin-binding protein toxofilin during Toxoplasma gondii infection. Cellular Microbiology, 12: 55–66. doi: 10.1111/j.1462-5822.2009.01378.x
- Issue published online: 27 DEC 2009
- Article first published online: 2 SEP 2009
- Received 21 February, 2009; revised 19 August, 2009; accepted 24 August, 2009.
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 (Δtxf) parasites in several in vitro assays, including invasion, growth, 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.