Toxoplasma gondii infection shifts dendritic cells into an amoeboid rapid migration mode encompassing podosome dissolution, secretion of TIMP-1, and reduced proteolysis of extracellular matrix

Abstract

Dendritic cells (DCs) infected by Toxoplasma gondii rapidly acquire a hypermigratory phenotype that promotes systemic parasite dissemination by a "Trojan horse" mechanism in mice. Recent paradigms of leukocyte migration have identified the amoeboid migration mode of DCs as particularly suited for rapid locomotion in extracellular matrix and tissues. Here, we have developed a microscopy-based high-throughput approach to assess motility and matrix degradation by Toxoplasma-challenged murine and human DCs. DCs challenged with T. gondii exhibited dependency on metalloproteinase activity for hypermotility and transmigration but, strikingly, also dramatically reduced pericellular proteolysis. Toxoplasma-challenged DCs up-regulated expression and secretion of tissue inhibitor of metalloproteinases-1 (TIMP-1) and their supernatants impaired matrix degradation by naïve DCs and by-stander DCs dose dependently. Gene silencing of TIMP-1 by short hairpin RNA restored matrix degradation activity in Toxoplasma-infected DCs. Additionally, dissolution of podosome structures in parasitised DCs coincided with abrogated matrix degradation. Toxoplasma lysates inhibited pericellular proteolysis in a MyD88-dependent fashion whereas abrogated proteolysis persevered in Toxoplasma-infected MyD88-deficient DCs. This indicated that both TLR/MyD88-dependent and TLR/MyD88-independent signalling pathways mediated podosome dissolution and the abrogated matrix degradation. We report that increased TIMP-1 secretion and cytoskeletal rearrangements encompassing podosome dissolution are features of Toxoplasma-induced hypermigration of DCs with an impact on matrix degradation. Jointly, the data highlight how an obligate intracellular parasite orchestrates key regulatory cellular processes consistent with non-proteolytic amoeboid migration of the vehicle cells that facilitate its dissemination.

Convergent Met and voltage-gated Ca2+channel signaling on Ras-Erk MAPK drives migratory activation of dendritic cells parasitized byToxoplasma gondii

“…Our data show that VGCC-CaM-CaMkII-Ras signaling mediate migratory activation of DCs upstream of Erk. Previously, we showed that the onset of the hypermigratory phenotype in DCs depends on live intracellular parasites and the discharge of T. gondii secretory organelles, independently of TLR-MyD88 signaling or chemotaxis (Fuks et al, 2012; Lambert et al, 2006; Olafsson et al, 2018; Sangare et al, 2019; Weidner et al, 2013). However, TLR-MyD88 signaling and chemotactic stimuli can also activate Erk.…”

“…Specifically, in T. gondii -infected macrophages, both TLR-MyD88-dependent and a strong component of TLR-MyD88-independent Erk activation has been described (Kim et al, 2006). However, T. gondii -induced hypermigration is fully maintained in MyD88-deficient DCs (Lambert et al, 2006; Olafsson et al, 2018). Similarly, hypermigration is independent of chemotactic stimuli (but can act synergistically with chemotaxis upon stimulation) (Fuks et al, 2012; Kanatani et al, 2015; Weidner et al, 2013).…”

“…T. gondii tachyzoites of the RFP-expressing Prugniaud strain (Pru and Pru-RFP, type II) (Lambert et al, 2009) or GFP-expressing Ptg strain (type II) (Weidner et al, 2013) lines were maintained by serial 2-days passages in human foreskin fibroblast (HFF-1 SCRC-1041, American Type Culture Collection) monolayers. The hypermigratory phenotype induced by type II strains in DCs has been previously characterized (Olafsson et al, 2018).…”

“…Within minutes of active invasion by T. gondii , DCs adopt a hypermigratory phenotype that mediates rapid systemic dissemination in mice (Kanatani et al, 2017; Weidner and Barragan, 2014). We recently reported that T. gondii induces mesenchymal-to-amoeboid transition (MAT) in DCs (Olafsson et al, 2018). Hypermigratory parasitized DCs are characterized by high-velocity amoeboid locomotion (termed hypermotility), cytoskeletal reorganization with loss of podosomes, decreased pericellular proteolysis and redistribution of integrins (Kanatani et al, 2015; Olafsson et al, 2018; Weidner et al, 2013).…”

“…We recently reported that T. gondii induces mesenchymal-to-amoeboid transition (MAT) in DCs (Olafsson et al, 2018). Hypermigratory parasitized DCs are characterized by high-velocity amoeboid locomotion (termed hypermotility), cytoskeletal reorganization with loss of podosomes, decreased pericellular proteolysis and redistribution of integrins (Kanatani et al, 2015; Olafsson et al, 2018; Weidner et al, 2013). Hypermigration is triggered by non-canonical GABA A receptor-mediated (Fuks et al, 2012) activation of VGCCs, subtype Ca V 1.3 (Kanatani et al, 2017) and Timp-1-mediated Itgb1-dependent activation of focal adhesion kinase (Fak) (Olafsson et al, 2019).…”