Voltage-dependent calcium channel signaling mediates GABAA receptor-induced migratory activation of dendritic cells infected by Toxoplasma gondii

Abstract

The obligate intracellular parasite Toxoplasma gondii exploits cells of the immune system to disseminate. Upon T. gondii-infection, γ–aminobutyric acid (GABA)/GABAA receptor signaling triggers a hypermigratory phenotype in dendritic cells (DCs) by unknown signal transduction pathways. Here, we demonstrate that calcium (Ca2+) signaling in DCs is indispensable for T. gondii-induced DC hypermotility and transmigration in vitro. We report that activation of GABAA receptors by GABA induces transient Ca2+ entry in DCs. Murine bone marrow-derived DCs preferentially expressed the L-type voltage-dependent Ca2+ channel (VDCC) subtype Cav1.3. Silencing of Cav1.3 by short hairpin RNA or selective pharmacological antagonism of VDCCs abolished the Toxoplasma-induced hypermigratory phenotype. In a mouse model of toxoplasmosis, VDCC inhibition of adoptively transferred Toxoplasma-infected DCs delayed the appearance of cell-associated parasites in the blood circulation and reduced parasite dissemination to target organs. The present data establish that T. gondii-induced hypermigration of DCs requires signaling via VDCCs and that Ca2+ acts as a second messenger to GABAergic signaling via the VDCC Cav1.3. The findings define a novel motility-related signaling axis in DCs and unveil that interneurons and DCs share common GABAergic motogenic pathways. T. gondii employs GABAergic non-canonical pathways to induce host cell migration and facilitate dissemination.

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

“…Importantly, hypermotility was abrogated by Ras antagonism (Fig. 2D, E), in line with VGCC inhibition, Ca V 1.3 gene silencing (Kanatani et al, 2017) and Mek inhibition (Fig. 1C; Fig.…”

“…Based on concepts in neuronal cells showing that Erk is activated downstream of VGCC signaling (Kotturi et al, 2003) and because VGCCs are implicated in T. gondii -mediated hypermotility (Kanatani et al, 2017), we hypothesized that VGCC-Ras-Raf-Mek signaling mediated Erk phosphorylation in T. gondii -infected DCs. To this end, we analyzed the phosphorylation of Erk in T. gondii -infected DCs upon treatment with the VGCC inhibitors nifedipine (targeting L-type VGCCs) and CPCPT (a selective inhibitor of VGCC subtype Ca V 1.3).…”

“…Paradoxically, T. gondii exploits the inherent migratory ability of DCs for dissemination via a ‘Trojan horse’ mechanism (Courret et al, 2006; Lambert et al, 2006; Lambert et al, 2009; Sangare et al, 2019). 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). 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). However, the signaling elements downstream of VGCC and Itgb1-Fak signaling, and their respective contribution to the hypermigratory phenotype of DCs, have remained unknown.…”

The hitchhiker's guide to parasite dissemination

“…T. gondii infection induces an in vitro hypermotility phenotype in DCs that includes enhanced migration across plastic transwell membranes and endothelial monolayers, increased crawling velocity and displacement on two-dimensional surfaces (Lambert et al, 2006), and enhanced migration through threedimensional collagen matrix (Kanatani, Uhlen, & Barragan, 2015). The hypermotility of infected DCs correlates to rapid cytoskeletal changes including integrin redistribution (Weidner et al, 2013), requires GABA receptor signalling to a Cav1.3 voltage-dependent calcium channel (Bhandage & Barragan, 2019;Fuks et al, 2012;Kanatani et al, 2017), and can be induced by heterologous expression of a 14-3-3 protein (Weidner et al, 2016). DCs are not typically found in the blood in large numbers and thus not surprisingly do not account for a meaningful portion of parasitised leukocytes in the blood during murine T. gondii infections (Courret et al, 2006).…”