Research activity
Toxoplasma gondii is responsible for toxoplasmosis, a disease which can be fatal for the fetus of infected mothers and for immunocompromised patients. This parasite belongs to the Apicomplexa family which contains other parasites of medical importance such as Plasmodium (responsible for malaria) or Cryptosporidium (cryptosporidiosis). The pathogenicity of these parasites is based on their ability to divide in order to proliferate in the organism. They have therefore invented simplified division patterns in order to multiply efficiently and quickly while producing a large number of parasites. The mechanisms allowing the control and coordination of these modes of division are therefore essential to their ability to proliferate and therefore to the survival of these parasites in humans. We aim to expand our understanding on how these parasites are able to evolve flexible modes of division allowing them to proliferate in a large number of different organisms. We study the proteins that control and coordinate the division of these parasites, using T. gondii as a model Apicomplexa. We are particularly interested in transcription factors of the ApiAP2 family which coordinate specific expression profiles during the cell cycle. In addition, we also study the centrosome, which serves as a platform coordinating the cell cycle. These studies could lead us to better understand the mechanisms of division in this family of parasites in order to develop new molecules aiming at controlling their proliferation.
Toxoplasma gondii effectors and modulation of the host immune responses
Despite being critical for the establishment of infection, the mechanisms governing exchanges between the parasite and its host remain poorly understood. Specifically, dense granule (GRA) proteins, which are released into the vacuolar space and host cytosol, act as key effectors that modulate the host cell immune defenses. These GRA proteins are also essential for the establishment of chronic toxoplasmosis, as they facilitate cyst formation in neurons. By employing reverse genetics, proteomics, live imaging, and high-resolution microscopy, our aim is to uncover and characterize novel secretory pathways that regulate parasite effector release and influence parasite virulence.
On the host side, we focus on how infection alters the immune response during both the acute and cerebral latent phases of the disease. In particular, we investigate how splenic dendritic cells respond to different parasite strains and how this affects antigen presentation and the development of long-term protective immunity. In the brain, we examine the impact of latent infection on immune cell signatures and how infection-induced chronic inflammation may alter neuronal function.
Plasmodium has an elaborate and fascinating life cycle aimed at adapting and surviving in different hosts and tissues. It is transmitted back and forth between a vertebrate (intermediate host) and an Anopheles mosquito (definitive host), where a bottleneck phase is rapidly followed by a period of replication, leading to the generation of thousands of progenies. During its intraerythrocytic cell development, it undergoes closed mitosis for its replication. This process, called schizogony, involves asynchronous DNA replication events, results in the formation of a multinucleated cell (schizont), and is followed by budding and cytokinesis. As expected, the regulation of such a replicative cycle differs from that of the eukaryotic cell cycle. In our team, we have investigated the role of the PP1 phosphatase and its regulators in Plasmodium and T. gondii. In eukaryotes, Inhibitor 2 (I2) and SDS22, two regulatory subunits of PP1, are involved in key steps of cell division, in particular in mitotic entry, chromosome condensation and cytokinesis. Our previous work showed that although I2 and LRR1 (SDS22 homolog in Plasmodium) interact with and regulate PP1 in both Plasmodium and T. gondii, they exhibit specificities in structure and phosphorylation pattern, the latter being known to be involved in their regulation. These data strongly suggest that in these parasites both proteins form holoenzymes that may have functions different from those described in eukaryotes, especially during parasite replication. Using reverse genetics, our aim is to explore the biological functions of I2 and LRR1 and the importance of their phosho-regulation to unravel the contribution of these complexes in the regulation of parasite proliferation.
Discovery of new antimalarial molecules through molecular and functional characterization of phosphatases specific to Plasmodium falciparum
Plasmodium falciparum (Pf), the deadliest agent of malaria, has developed resistance to almost all chemotherapeutics. It is necessary to understand the biology of this parasite in order to develop new drugs. In Pf, extensive research has now been started to study the Pf kinome and to examine whether targeting kinases could represent an effective mean for the treatment of the infection. However, the study of Pf phosphatases is still under-investigated. Amino acid sequence comparative analyses of Pf and Plasmodium berghei (Pb), a rodent malaria species, revealed that several of them are Plasmodium specific. Among these phosphatases, three were also suggested to be essential for blood stage parasites development of Pf. The present project is focused on the molecular and functional characterization of one of them and on the validation of this specific phosphatase as a new potential target for malaria. The gene has been cloned, annotated and expressed as a recombinant protein and its phosphatase activity has been demonstrated in vitro. Functional characterization in vivo was explored by conditional gene knock-out studies as well as by generating knock-in parasite lines to follow their trafficking during the parasite lifecycle (in Pf and Pb). Finally, we solved in silico the 3D structure of catalytic site of this phosphatase by homology modelling and identified a new set of potential specific inhibitors. A first series of pharmacomodulations allowed us to discover 13 new hits with IC50 ≤ 100 nM, including 3 with IC50 ≤ 50 nM. Cytotoxicity studies showed low cell toxicity on human HFF (Human Foreskin Fibroblasts) and HepG2 (human hepatocytes) cells with a selectivity index >100. Currently we are pursuing the pharmacomodulation of these hits, the molecule-target interaction and the in vivo efficacy in an animal model of malaria. Our project is supported by Inserm Transfert and SATT Nord.
