Toxoplasma gondii excreted/secreted proteases disrupt intercellular junction proteins in epithelial cell monolayers to facilitate tachyzoites paracellular migration

Abstract

Toxoplasma gondii shows high dissemination and migration properties across biological barriers infecting immunologically privileged organs. Toxoplasma uses different routes for dissemination; however, the mechanisms are not fully understood. Herein, we studied the effects of proteases present in excretion/secretion products (ESPs) of Toxoplasma on MDCK cell monolayers. Ultrastructural analysis showed that ESPs of Toxoplasma disrupt the intercellular junctions (IJ) of adjacent cells. The tight junction (TJ) proteins ZO‐1, occludin, and claudin‐1 suffered a progressive decrease in protein levels upon ESPs treatment. In addition, ESPs induced mislocalization of such TJ proteins, along with the adherent junction protein E‐cadherin, and this was prevented by pre‐treating the ESPs with protease inhibitors. Reorganisation of cytoskeleton proteins was also observed. Endocytosis inhibitors, Dyngo®‐4a and Dynasore, impeded the modifications, suggesting that TJ proteins internalisation is triggered by the ESPs proteases hence contributing to the loss of IJ. The observed disruption in TJ proteins went in line with a decrease in the transepithelial electrical resistance of the monolayers, which was significantly blocked by pre‐treating ESPs with metalloprotease and serine protease inhibitors. Moreover, exposure of cell monolayers to ESPs facilitated paracellular migration of tachyzoites. Our results demonstrate that Toxoplasma ESPs contain proteases that can disrupt the IJ of epithelial monolayers and this could facilitate the paracellular route for Toxoplasma tissue dissemination and migration.

Heligmosomoides bakeriandToxoplasma gondiico-infection leads to increased mortality associated with intestinal pathology

“…Gene expression of tight junction proteins Zo1 and Occludin , and the key regulator of inflammation Il-22 were measured in the SI (SI2 region). Epithelial barrier dysfunction is characterised by decreased levels of Zo1 and Occludin , which has been observed during Tg infection both at the gene expression (75) and protein levels (76). Transgenic animals lacking Il-22 infected with Tg have higher mortalities than their wildtype counterparts (77).…”

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“…To determine the effect of T. denticola infection on tight junctions, Madin-Darby canine kidney epithelial (MDCK) cells NBL-2, which are widely used in epithelial models to investigate tight junctions 21,26,29) , were used. The MDCK cells were obtained from the Health Research Resources Bank (Osaka, Japan) and maintained in α-minimum essential Medium (MEM) supplemented with 10% fetal bovine serum (FBS) (Gibco Life Technologies, New York, NY, USA) and antibodies at 37°C in a humidified 5% CO 2 atmosphere.…”

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“…It has also been observed that tachyzoites co-localize with the tight junction protein occludin, which appears to facilitate paracellular transmigration of the parasite ( Barragan et al., 2005 ; Weight and Carding, 2012 ). Recent data suggest that paracellular migration is facilitated by parasite secretory proteases that target tight junction proteins ZO-1, occludin, and claudin-1 ( Ramirez-Flores et al., 2020 ).…”