Date Published: May 29, 2014
Publisher: Public Library of Science
Author(s): Aline Dumas, Nathalie Amiable, Juan Pablo de Rivero Vaccari, Jae Jin Chae, Robert W. Keane, Steve Lacroix, Luc Vallières, Dana J. Philpott.
Microbial agents can aggravate inflammatory diseases, such as multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE). An example is pertussis toxin (PTX), a bacterial virulence factor commonly used as an adjuvant to promote EAE, but whose mechanism of action is unclear. We have reported that PTX triggers an IL-6-mediated signaling cascade that increases the number of leukocytes that patrol the vasculature by crawling on its luminal surface. In the present study, we examined this response in mice lacking either TLR4 or inflammasome components and using enzymatically active and inactive forms of PTX. Our results indicate that PTX, through its ADP-ribosyltransferase activity, induces two series of events upstream of IL-6: 1) the activation of TLR4 signaling in myeloid cells, leading to pro-IL-1β synthesis; and 2) the formation of a pyrin-dependent inflammasome that cleaves pro-IL-1β into its active form. In turn, IL-1β stimulates nearby stromal cells to secrete IL-6, which is known to induce vascular changes required for leukocyte adhesion. Without pyrin, PTX does not induce neutrophil adhesion to cerebral capillaries and is less effective at inducing EAE in transgenic mice with encephalitogenic T lymphocytes. This study identifies the first microbial molecule that activates pyrin, a mechanism by which infections may influence MS and a potential therapeutic target for immune disorders.
MS is a T lymphocyte-mediated autoimmune demyelinating disease of the central nervous system (CNS), whose development is dictated by complex interactions between genetic and environmental factors , . Among the latter are microbes and their toxins. Although the search for a causative microbial agent has proved fruitless so far, many epidemiological studies have shown associations between infections and MS. For example, the risk of developing MS is at least doubled in individuals with a clinical history of infectious mononucleosis –, while the risk of MS exacerbation is higher after common infections of the upper respiratory tract , –.
This study provides answers to the long-lasting question of how PTX produces an adjuvant effect that promotes EAE. First, contrary to the prevailing concept , our results suggest that PTX does not act directly via membrane-bound receptors such as TLR4, but rather by catalyzing the ADP-ribosylation of G-protein αi subunits, a phenomenon that requires the internalization and disassembling of PTX to release its A protomer in the cytoplasm. Indeed, mutant PTX, containing only 2 amino acid substitutions in one of its 6 subunits, was unable to activate IL-1β transcription (Fig. 4), which was dependent on TLR4 (Fig. 3). The mutations affect the enzymatic activity of the A-protomer, but not the antigenic structure of the whole toxin . The protein binding properties of mutant PTX are thus likely to be comparable to that of wild-type PTX. Therefore, we propose that PTX causes perturbations in G protein-coupled receptor signaling (e.g., cAMP-dependent pathways), leading to the release of an endogenous danger signal called alarmin that activates TLR4 on myeloid cells and drives IL-1β transcription. Several TLR4-interacting alarmins have been identified so far, including HMGB1, S100A8/A9 and heat-shock proteins . Additional work will be needed to further clarify the mechanism by which TLR4 is activated in response to PTX. Meanwhile, we speculate that genetic defects or other factors that may disrupt G protein-coupled receptor signaling in the manner of PTX might contribute to MS and MS-like diseases.