Microbial virulence and inflammatory signaling in disease (VIRINFLAM)
Involvement of the NLRP3 inflammasome in bacterial toxin detection and COVID-19 evolution
The innate immune detection of pathogenic microbes is crucial to enhance the immune response. During an infection, the activation of the innate immune system is the first step to establish an adaptative immune response. A physiological immune response is defined as adapted to the microbial environment and as an effectively resolved inflammation. To do so, the host must be able to qualitatively and quantitatively detect microbes. The detection of conserved MAMPs allows the host to evaluate the number of microbes. On the other hand, the detection of virulence factors’ activity – pathogen-specific – give a qualitative view of microbes.
The discovery of inflammasomes has allowed a major advance in the understanding of the mechanisms of detection of pathogen- and danger associated molecular patterns. Inflammasomes are macromolecular complexes responsible for the activation of Caspase-1 and the maturation of IL-1ß and IL-18. Initially discovered for their involvement in autoinflammatory pathologies, there is growing evidence for the role of inflammasomes in detecting the activity of toxins and microbial effectors including those targeting Rho GTPases.
Rho GTPases are located at the crossroads of major cell signaling pathways; by regulating cell migration, phagocytosis or even gene transcription, these are major players in immunity. This makes them preferential targets for pathogenic microorganisms. Indeed, more than 30 bacterial virulence factors manipulate Rho GTPases using various strategies leading to their inhibition or activation.
The uropathogenic Escherichia coli (E. coli) is the primary cause of cystitis and pyelonephritis and in the most severe cases of bacteremia. More than a third of uropathogenic E. coli possess the CNF1 toxin, a de-amidase that activates Rho GTPases. This modification destroys the intrinsic GTPase activity stimulated by the GAPs (GTPase-activating protein) of Rho, Rac and Cdc42.
Before my arrival at the laboratory, Dr. Laurent Boyer's team showed that the CNF1 toxin induces an immune response. Indeed, during bacteremia in mice or systemic infection in drosophila, E. coli expressing CNF1 are eliminated more quickly than E. coli deleted for CNF1. At the cellular level, the activation of Rho GTPases by CNF1 is responsible for the production of pro-inflammatory cytokines. Alongside this transcriptional response, the CNF1 toxin causes IL-1β to mature in a Caspase-1 dependent fashion, suggesting the involvement of an inflammasome.
My thesis project was to identify the inflammasome involved in the detection of the CNF1 toxin and to characterize the signaling pathway leading to this activation.
We have identified the NLRP3 inflammasome as being responsible for the detection of GTPase Rac2 activation by the CNF1 toxin. We then studied the role of the kinase Pak1, a major effector of Rac2, in this signaling pathway and we were able to show that Pak1 plays a crucial role in the activation of the inflammasome NLRP3 by CNF1. Pak1 phosphorylates Thr659 of the NLRP3 receptor and this phosphorylation is critical for the recruitment of the regulatory protein Nek7 and the activation of the NLRP3 inflammasome downstream of the detection of CNF1. Finally, we were able to show the major role of the Pak1-NLRP3 pathway in the establishment of an anti-virulence immune response during mice bacteremia.
In addition, we used our expertise to study the activation of the NLRP3 inflammasome in patients infected with SARS-CoV-2. This study allowed us to establish the signature of the NLRP3 response in the circulating myeloid cells of COVID-19 patients and to use these parameters to define a score to predict the patients’ outcome.
My work on the involvement of the NLRP3 inflammasome in the detection of virulence has provided a better understanding of the mechanisms involved in an anti-infective response.
Inflammasome – Rho-GTPases – Virulence factors – COVID-19
Dr Patrick AUBERGER
Dr Thomas HENRY
Dr Nicolas MANEL
PhD supervisor :
Dr Laurent BOYER