Active-Cargo Biophysics in Immune Cells Hijacked by Toxoplasma gondii

• Applicants: Professor Dr. Jörg Renkawitz; Professor Dr. Benedikt Sabass
• Subject Area: Biophysics ; Parasitology and Biology of Tropical Infectious Disease Pathogens ; Statistical Physics, Nonlinear Dynamics, Complex Systems, Soft and Fluid Matter, Biological Physics ; Cell Biology
• Term: since 2021
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 492014049

The protozoan parasite Toxoplasma gondii (T.gondii) causes toxoplasmosis in humans and disseminates rapidly throughout a host body. As a strategy for immune response evasion and to achieve rapid traversal of large distances between different organs, T.gondii hijacks migrating immune cells, taking advantage of their abilities to cross tissue barriers. Moreover, T.gondii replicates inside intracellular parasitophorous vacuoles (PV), thus turning immune cells into “Trojan horses”. While T.gondii biology is a well-established field, key biophysical questions regarding the trojan-horse mechanism are currently almost completely unresolved. During the past two years of the 1st funding period, we focused on the question of how host-cell parasite interactions enable the Trojan horses to pass through microenvironmental barriers. Experiments revealed that infected immune cells can migrate through extraordinarily tight constrictions in spite of a large, bulky parasite “cargo” since they up-regulate generated forces. Strikingly, cells orchestrate the relative position of their main cargoes, namely the PV and the nucleus, depending on the geometry and size of the constrictions. Experiments and computer simulations together reveal a microtubule-independent mechanism by which the larger cargo is placed at the front of the cell, allowing effective squeezing through narrow pores. We also discovered that pore passage is predicated on unusual PV mechanics, which is very stiff, yet extremely deformable and resilient. Anatomically, the PV is a multi-layered structure where a host microtubule network surrounds a delimiting membrane that contains the replicating parasites. Intravacuolar parasites are connected by posterior membrane tubes containing a delicate F-actin network, where dedicated T. gondii myosins possibly act as molecular motors. This network is essential for PV organization, mechanics, and communication of parasites but its architecture and biophysics are largely unknown. Moreover, preliminary data indicates that PV organization possibly affects the path-decision making of navigating immune cells. We also have indications that mechanical forces on the hijacked cells stimulate parasite egress under certain conditions.For the 2nd funding period, we therefore propose to investigate the basic anatomy and biophysics of the PV. We will test the "active-cargo" hypothesis that an active PV cytoskeleton determines its mechanical properties, determine how the cargo determines path decisions in defined environments, and determine when and how external forces induce cargo release from the Trojan horse. Combining our expertise in biology and biophysics, we will use advanced microscopy (dSTORM), custom-made microchannels and 3D matrices, parasite mutants, mechanical measurements (TFM, AFM), and simulate the PV architecture in locomoting cells. Overall, the work will elucidate biophysical design principles and fundamental parasitic strategies based on Trojan horses.

• DFG Programme: Priority Programmes
• Subproject of: SPP 2332: Physics of Parasitism