Elucidating Pathways of Toxoplasma gondii Infection in the Gastrointestinal Tract: Involvement of the Tight Junction Protein Occludin

Toxoplasma gondii (T. gondii) is an obligate intracellular parasite, infecting one third of the world’s population through the consumption of contaminated undercooked meat, water and soil. The parasite’s primary route of infection is via the small intestine, although the routes of entry remain unclear. Using epithelial cells derived from the small intestine we set out to determine if the parasite transmigrates epithelial cells via the paracellular route by interacting with and/or disrupting tight junction complexes.
In vitro invasion assays revealed that T. gondii infects and transmigrates through polarized epithelial cell monolayers without altering barrier integrity. However, during invasion Toxoplasma co-localized with occludin, which was subsequently redistributed from tight junction complexes to intracellular sites. Reduction of occludin expression reduced the ability of Toxoplasma to penetrate epithelial cells, consistent with the involvement of occludin in parasite transmigration. Furthermore, in vitro binding assays using recombinant fragments of occludin confirmed the ability of the parasite to interact with occludin and in particular with the extracellular loops. Following immunoprecipitation of occludin from infected cell lysates, we have identified potential candidate occludin-binding partners from T. gondii, by mass spectrometry. Together, we provide evidence that T. gondii can interact with occludin to help facilitate its infection of epithelial cells.