A Macrophage-Pericyte Axis Directs Tissue Restoration via Amphiregulin-Induced Transforming Growth Factor Beta Activation.

The epidermal growth factor receptor ligand Amphiregulin has a well-documented role in the restoration of tissue homeostasis after injury; however, the mechanism by which Amphiregulin contributes to wound repair remains unknown. Here we show that Amphiregulin functioned by releasing bioactive transforming growth factor beta (TGF-ß) from latent complexes via integrin-αV activation. Using acute injury models in two different tissues, we found that by inducing TGF-ß activation on mesenchymal stromal cells (pericytes), Amphiregulin induced their differentiation into myofibroblasts, thereby selectively contributing to the restoration of vascular barrier function within injured tissue. Furthermore, we identified macrophages as a critical source of Amphiregulin, revealing a direct effector mechanism by which these cells contribute to tissue restoration after acute injury. Combined, these observations expose a so far under-appreciated mechanism of how cells of the immune system selectively control the differentiation of tissue progenitor cells during tissue repair and inflammation.

HpARI Protein Secreted by a Helminth Parasite Suppresses Interleukin-33.

Infection by helminth parasites is associated with amelioration of allergic reactivity, but mechanistic insights into this association are lacking. Products secreted by the mouse parasite Heligmosomoides polygyrus suppress type 2 (allergic) immune responses through interference in the interleukin-33 (IL-33) pathway. Here, we identified H. polygyrus Alarmin Release Inhibitor (HpARI), an IL-33-suppressive 26-kDa protein, containing three predicted complement control protein (CCP) modules. In vivo, recombinant HpARI abrogated IL-33, group 2 innate lymphoid cell (ILC2) and eosinophilic responses to Alternaria allergen administration, and diminished eosinophilic responses to Nippostrongylus brasiliensis, increasing parasite burden. HpARI bound directly to both mouse and human IL-33 (in the cytokine's activated state) and also to nuclear DNA via its N-terminal CCP module pair (CCP1/2), tethering active IL-33 within necrotic cells, preventing its release, and forestalling initiation of type 2 allergic responses. Thus, HpARI employs a novel molecular strategy to suppress type 2 immunity in both infection and allergy.

Exogenous Transforming Growth Factor-ß1 and Its Helminth-Derived Mimic Attenuate the Heart's Inflammatory Response to Ischemic Injury and Reduce Mature Scar Size.

Coronary reperfusion after acute ST-elevation myocardial infarction (STEMI) is standard therapy to salvage ischemic heart muscle. However, subsequent inflammatory responses within the infarct lead to further loss of viable myocardium. Transforming growth factor (TGF)-ß1 is a potent anti-inflammatory cytokine released in response to tissue injury. The aim of this study was to investigate the protective effects of TGF-ß1 after MI. In patients with STEMI, there was a significant correlation (P = 0.003) between higher circulating TGF-ß1 levels at 24 hours after MI and a reduction in infarct size after 3 months, suggesting a protective role of early increase in circulating TGF-ß1. A mouse model of cardiac ischemia reperfusion was used to demonstrate multiple benefits of exogenous TGF-ß1 delivered in the acute phase. It led to a significantly smaller infarct size (30% reduction, P = 0.025), reduced inflammatory infiltrate (28% reduction, P = 0.015), lower intracardiac expression of inflammatory cytokines IL-1ß and chemokine (C-C motif) ligand 2 (>50% reduction, P = 0.038 and 0.0004, respectively) at 24 hours, and reduced scar size at 4 weeks (21% reduction, P = 0.015) after reperfusion. Furthermore, a low-fibrogenic mimic of TGF-ß1, secreted by the helminth parasite Heligmosomoides polygyrus, had an almost identical protective effect on injured mouse hearts. Finally, genetic studies indicated that this benefit was mediated by TGF-ß signaling in the vascular endothelium.

IL-33: A central cytokine in helminth infections.

IL-33 is an alarmin cytokine which has been implicated in allergy, fibrosis, inflammation, tumorigenesis, metabolism, and homeostasis. However, amongst its strongest roles are in helminth infections, where IL-33 usually (but not always) is central to induction of an effective anti-parasitic immune response. In this review, we will summarise the literature around this fascinating cytokine, its activity on immune and non-immune cells, the unique (and sometimes counterintuitive) responses it induces, and how it can coordinate the immune response during infections by parasitic helminths. Finally, we will summarise some of the ways that parasites have developed to modulate the IL-33 pathway for their own benefit.

Convergent evolution of a parasite-encoded complement control protein-scaffold to mimic binding of mammalian TGF-ß to its receptors, TßRI and TßRII.

The mouse intestinal helminth Heligmosomoides polygyrus modulates host immune responses by secreting a transforming growth factor (TGF)-ß mimic (TGM), to expand the population of Foxp3+ Tregs. TGM comprises five complement control protein (CCP)-like domains, designated D1-D5. Though lacking homology to TGF-ß, TGM binds directly to the TGF-ß receptors TßRI and TßRII and stimulates the differentiation of naïve T-cells into Tregs. However, the molecular determinants of binding are unclear. Here, we used surface plasmon resonance, isothermal calorimetry, NMR spectroscopy, and mutagenesis to investigate how TGM binds the TGF-ß receptors. We demonstrate that binding is modular, with D1-D2 binding to TßRI and D3 binding to TßRII. D1-D2 and D3 were further shown to compete with TGF-ß(TßRII)2 and TGF-ß for binding to TßRI and TßRII, respectively. The solution structure of TGM-D3 revealed that TGM adopts a CCP-like fold but is also modified to allow the C-terminal strand to diverge, leading to an expansion of the domain and opening potential interaction surfaces. TGM-D3 also incorporates a long structurally ordered hypervariable loop, adding further potential interaction sites. Through NMR shift perturbations and binding studies of TGM-D3 and TßRII variants, TGM-D3 was shown to occupy the same site of TßRII as bound by TGF-ß using both a novel interaction surface and the hypervariable loop. These results, together with the identification of other secreted CCP-like proteins with immunomodulatory activity in H. polygyrus, suggest that TGM is part of a larger family of evolutionarily plastic parasite effector molecules that mediate novel interactions with their host.

Metabolic regulation by prostaglandin E2 impairs lung group 2 innate lymphoid cell responses.

BACKGROUND: Group 2 innate lymphoid cells (ILC2s) play a critical role in asthma pathogenesis. Non-steroidal anti-inflammatory drug (NSAID)-exacerbated respiratory disease (NERD) is associated with reduced signaling via EP2, a receptor for prostaglandin E2 (PGE2 ). However, the respective roles for the PGE2 receptors EP2 and EP4 (both share same downstream signaling) in the regulation of lung ILC2 responses has yet been deciphered. METHODS: The roles of PGE2 receptors EP2 and EP4 on ILC2-mediated lung inflammation were investigated using genetically modified mouse lines and pharmacological approaches in IL-33-induced lung allergy model. The effects of PGE2 receptors and downstream signals on ILC2 metabolic activation and effector function were examined using in vitro cell cultures. RESULTS: Deficiency of EP2 rather than EP4 augments IL-33-induced mouse lung ILC2 responses and eosinophilic inflammation in vivo. In contrast, exogenous agonism of EP4 and EP2 or inhibition of phosphodiesterase markedly restricts IL-33-induced lung ILC2 responses. Mechanistically, PGE2 directly suppresses IL-33-dependent ILC2 activation through the EP2/EP4-cAMP pathway, which downregulates STAT5 and MYC pathway gene expression and ILC2 energy metabolism. Blocking glycolysis diminishes IL-33-dependent ILC2 responses in mice where endogenous PG synthesis or EP2 signaling is blocked but not in mice with intact PGE2 -EP2 signaling. CONCLUSION: We have defined a mechanism for optimal suppression of mouse lung ILC2 responses by endogenous PGE2 -EP2 signaling which underpins the clinical findings of defective EP2 signaling in patients with NERD. Our findings also indicate that exogenously targeting the PGE2 -EP4-cAMP and energy metabolic pathways may provide novel opportunities for treating the ILC2-initiated lung inflammation in asthma and NERD.

Suppression of airway allergic eosinophilia by Hp-TGM, a helminth mimic of TGF-ß.

Type 2-high asthma is a chronic inflammatory disease of the airways which is increasingly prevalent in countries where helminth parasite infections are rare, and characterized by T helper 2 (Th2)-dependent accumulation of eosinophils in the lungs. Regulatory cytokines such as TGF-ß can restrain inflammatory reactions, dampen allergic Th2 responses, and control eosinophil activation. The murine helminth parasite Heligmosomoides polygyrus releases a TGF-ß mimic (Hp-TGM) that replicates the biological and functional properties of TGF-ß despite bearing no structural similarity to the mammalian protein. Here, we investigated if Hp-TGM could alleviate allergic airway inflammation in mice exposed to Alternaria alternata allergen, house dust mite (HDM) extract or alum-adjuvanted ovalbumin protein (OVA). Intranasal administration of Hp-TGM during Alternaria exposure sharply reduced airway and lung tissue eosinophilia along with bronchoalveolar lavage fluid IL-5 and lung IL-33 cytokine levels at 24 h. The protective effect of Hp-TGM on airway eosinophilia was also obtained in the longer T-cell mediated models of HDM or OVA sensitisation with significant inhibition of eotaxin-1, IL-4 and IL-13 responses depending on the model and time-point. Hp-TGM was also protective when administered parenterally either when given at the time of allergic sensitisation or during airway allergen challenge. This project has taken the first steps in identifying the role of Hp-TGM in allergic asthma and highlighted its ability to control lung inflammation and allergic pathology. Future research will investigate the mode of action of Hp-TGM against airway allergic eosinophilia, and further explore its potential to be developed as a biotherapeutic in allergic asthma.

Intestinal epithelial tuft cells initiate type 2 mucosal immunity to helminth parasites.

Helminth parasitic infections are a major global health and social burden. The host defence against helminths such as Nippostrongylus brasiliensis is orchestrated by type 2 cell-mediated immunity. Induction of type 2 cytokines, including interleukins (IL) IL-4 and IL-13, induce goblet cell hyperplasia with mucus production, ultimately resulting in worm expulsion. However, the mechanisms underlying the initiation of type 2 responses remain incompletely understood. Here we show that tuft cells, a rare epithelial cell type in the steady-state intestinal epithelium, are responsible for initiating type 2 responses to parasites by a cytokine-mediated cellular relay. Tuft cells have a Th2-related gene expression signature and we demonstrate that they undergo a rapid and extensive IL-4Rα-dependent amplification following infection with helminth parasites, owing to direct differentiation of epithelial crypt progenitor cells. We find that the Pou2f3 gene is essential for tuft cell specification. Pou2f3(-/-) mice lack intestinal tuft cells and have defective mucosal type 2 responses to helminth infection; goblet cell hyperplasia is abrogated and worm expulsion is compromised. Notably, IL-4Rα signalling is sufficient to induce expansion of the tuft cell lineage, and ectopic stimulation of this signalling cascade obviates the need for tuft cells in the epithelial cell remodelling of the intestine. Moreover, tuft cells secrete IL-25, thereby regulating type 2 immune responses. Our data reveal a novel function of intestinal epithelial tuft cells and demonstrate a cellular relay required for initiating mucosal type 2 immunity to helminth infection.

The parasite cytokine mimic Hp-TGM potently replicates the regulatory effects of TGF-ß on murine CD4+ T cells.

Transforming growth factor-beta (TGF-ß) family proteins mediate many vital biological functions in growth, development and regulation of the immune system. TGF-ß itself controls immune homeostasis and inflammation, including conversion of naïve CD4+ T cells into Foxp3+ regulatory T cells (Tregs) in the presence of interleukin-2 and T-cell receptor ligands. The helminth parasite Heligmosomoides polygyrus exploits this pathway through a structurally novel TGF-ß mimic (Hp-TGM), which binds to mammalian TGF-ß receptors and induces Tregs. Here, we performed detailed comparisons of Hp-TGM with mammalian TGF-ß. Compared with TGF-ß, Hp-TGM induced greater numbers of Foxp3+ Tregs (iTregs), with more intense Foxp3 expression. Both ligands upregulated Treg functional markers CD73, CD103 and programmed death-ligand 1, but Hp-TGM induced significantly higher CD39 expression than did TGF-ß. Interestingly, in contrast to canonical TGF-ß signaling through Smad2/3, Hp-TGM stimulation was slower and more sustained. Gene expression profiles induced by TGF-ß and Hp-TGM were remarkably similar, and both types of iTregs suppressed T-cell responses in vitro and experimental autoimmune encephalomyelitis-driven inflammation in vivo. In vitro, both types of iTregs were equally stable under inflammatory conditions, but Hp-TGM-induced iTregs were more stable in vivo during dextran sodium sulfate-induced colitis, with greater retention of Foxp3 expression and lower conversion to a ROR-γt+ phenotype. Altogether, results from this study suggest that the parasite cytokine mimic, Hp-TGM, may deliver a qualitatively different signal to CD4+ T cells with downstream consequences for the long-term stability of iTregs. These data highlight the potential of Hp-TGM as a new modulator of T-cell responses in vitro and in vivo.

Induction of stable human FOXP3+ Tregs by a parasite-derived TGF-ß mimic.

Immune homeostasis in the intestine is tightly controlled by FOXP3+ regulatory T cells (Tregs), defects of which are linked to the development of chronic conditions, such as inflammatory bowel disease (IBD). As a mechanism of immune evasion, several species of intestinal parasites boost Treg activity. The parasite Heligmosomoides polygyrus is known to secrete a molecule (Hp-TGM) that mimics the ability of TGF-ß to induce FOXP3 expression in CD4+ T cells. The study aimed to investigate whether Hp-TGM could induce human FOXP3+ Tregs as a potential therapeutic approach for inflammatory diseases. CD4+ T cells from healthy volunteers were expanded in the presence of Hp-TGM or TGF-ß. Treg induction was measured by flow cytometric detection of FOXP3 and other Treg markers, such as CD25 and CTLA-4. Epigenetic changes were detected using ChIP-Seq and pyrosequencing of FOXP3. Treg phenotype stability was assessed following inflammatory cytokine challenge and Treg function was evaluated by cellular co-culture suppression assays and cytometric bead arrays for secreted cytokines. Hp-TGM efficiently induced FOXP3 expression (> 60%), in addition to CD25 and CTLA-4, and caused epigenetic modification of the FOXP3 locus to a greater extent than TGF-ß. Hp-TGM-induced Tregs had superior suppressive function compared with TGF-ß-induced Tregs, and retained their phenotype following exposure to inflammatory cytokines. Furthermore, Hp-TGM induced a Treg-like phenotype in in vivo differentiated Th1 and Th17 cells, indicating its potential to re-program memory cells to enhance immune tolerance. These data indicate Hp-TGM has potential to be used to generate stable human FOXP3+ Tregs to treat IBD and other inflammatory diseases.

TGF-ß in tolerance, development and regulation of immunity.

The TGF-ß superfamily is an ancient metazoan protein class which cuts across cell and tissue differentiation, developmental biology and immunology. Its many members are regulated at multiple levels from intricate control of gene transcription, post-translational processing and activation, and signaling through overlapping receptor structures and downstream intracellular messengers. We have been interested in TGF-ß homologues firstly as key players in the induction of immunological tolerance, the topic so closely associated with Ray Owen. Secondly, our interests in how parasites may manipulate the immune system of their host has also brought us to study the TGF-ß pathway in infections with longlived, essentially tolerogenic, helminth parasites. Finally, within the spectrum of mammalian TGF-ß proteins is an exquisitely tightly-regulated gene, anti-Müllerian hormone (AMH), whose role in sex determination underpins the phenotype of freemartin calves that formed the focus of Ray's seminal work on immunological tolerance.

Helminth protein enhances wound healing by inhibiting fibrosis and promoting tissue regeneration.

Skin wound healing due to full thickness wounds typically results in fibrosis and scarring, where parenchyma tissue is replaced with connective tissue. A major advance in wound healing research would be to instead promote tissue regeneration. Helminth parasites express excretory/secretory (ES) molecules, which can modulate mammalian host responses. One recently discovered ES protein, TGF-ß mimic (TGM), binds the TGF-ß receptor, though likely has other activities. Here, we demonstrate that topical administration of TGM under a Tegaderm bandage enhanced wound healing and tissue regeneration in an in vivo wound biopsy model. Increased restoration of normal tissue structure in the wound beds of TGM-treated mice was observed during mid- to late-stage wound healing. Both accelerated re-epithelialization and hair follicle regeneration were observed. Further analysis showed differential expansion of myeloid populations at different wound healing stages, suggesting recruitment and reprogramming of specific macrophage subsets. This study indicates a role for TGM as a potential therapeutic option for enhanced wound healing.

Tetraspanins on the surface of Schistosoma mansoni are protective antigens against schistosomiasis.

Schistosomes are blood-dwelling flukes that infect 200 million people worldwide and are responsible for hundreds of thousands of deaths annually. Using a signal sequence trap, we cloned from Schistosoma mansoni two cDNAs, Sm-tsp-1 and Sm-tsp-2, encoding the tetraspanin (TSP) integral membrane proteins TSP-1 and TSP-2. We raised antibodies to recombinant TSP fusion proteins and showed that both proteins are exposed on the surface of S. mansoni. Recombinant TSP-2, but not TSP-1, is strongly recognized by IgG1 and IgG3 (but not IgE) from naturally resistant individuals but is not recognized by IgG from chronically infected or unexposed individuals. Vaccination of mice with the recombinant proteins followed by challenge infection with S. mansoni resulted in reductions of 57% and 64% (TSP-2) and 34% and 52% (TSP-1) for mean adult worm burdens and liver egg burdens, respectively, over two independent trials. Fecal egg counts were reduced by 65-69% in both test groups. TSP-2 in particular provided protection in excess of the 40% benchmark set by the World Health Organization for progression of schistosome vaccine antigens into clinical trials. When coupled with its selective recognition by naturally resistant people, TSP-2 seems to be an effective vaccine antigen against S. mansoni.

Protection from T cell-dependent colitis by the helminth-derived immunomodulatory mimic of transforming growth factor-ß, Hp-TGM.

In animal models of inflammatory colitis, pathology can be ameliorated by several intestinal helminth parasites, including the mouse nematode Heligmosomoides polygyrus. To identify parasite products that may exert anti-inflammatory effects in vivo, we tested H. polygyrus excretory-secretory (HES) products, as well as a recombinantly expressed parasite protein, transforming growth factor mimic (TGM), that functionally mimics the mammalian immunomodulatory cytokine TGF-ß. HES and TGM showed a degree of protection in dextran sodium sulphate-induced colitis, with a reduction in inflammatory cytokines, but did not fully block the development of pathology. HES also showed little benefit in a similar acute trinitrobenzene sulphonic acid-induced model. However, in a T cell transfer-mediated model with recombination activation gene (RAG)-deficient mice, HES-reduced disease scores if administered throughout the first 2 or 4 weeks following transfer but was less effective if treatment was delayed until 14 days after T cell transfer. Recombinant TGM similarly dampened colitis in RAG-deficient recipients of effector T cells, and was effective even if introduced only once symptoms had begun to be manifest. These results are a promising indication that TGM may replicate, and even surpass, the modulatory properties of native parasite HES.

The helminth TGF-ß mimic TGM4 is a modular ligand that binds CD44, CD49d and TGF-ß receptors to preferentially target myeloid cells.

The murine helminth parasite Heligmosomoides polygyrus expresses a family of modular proteins which, replicating the functional activity of the immunomodulatory cytokine TGF-ß, have been named TGM (TGF-ß Μimic). Multiple domains bind to different receptors, including TGF-ß receptors TßRI (ALK5) and TßRII through domains 1-3, and prototypic family member TGM1 binds the cell surface co-receptor CD44 through domains 4-5. This allows TGM1 to induce T lymphocyte Foxp3 expression, characteristic of regulatory (Treg) cells, and to activate a range of TGF-ß-responsive cell types. In contrast, a related protein, TGM4, targets a much more restricted cell repertoire, primarily acting on myeloid cells, with less potent effects on T cells and lacking activity on other TGF-ß-responsive cell types. TGM4 binds avidly to myeloid cells by flow cytometry, and can outcompete TGM1 for cell binding. Analysis of receptor binding in comparison to TGM1 reveals a 10-fold higher affinity than TGM1 for TGFßR-I (TßRI), but a 100-fold lower affinity for TßRII through Domain 3. Consequently, TGM4 is more dependent on co-receptor binding; in addition to CD44, TGM4 also engages CD49d (Itga4) through Domains 1-3, as well as CD206 and Neuropilin-1 through Domains 4 and 5. TGM4 was found to effectively modulate macrophage populations, inhibiting lipopolysaccharide-driven inflammatory cytokine production and boosting interleukin (IL)-4-stimulated responses such as Arginase-1 in vitro and in vivo. These results reveal that the modular nature of TGMs has allowed the fine tuning of the binding affinities of the TßR- and co-receptor binding domains to establish cell specificity for TGF-ß signalling in a manner that cannot be attained by the mammalian cytokine.

The IL-25-dependent tuft cell circuit driven by intestinal helminths requires macrophage migration inhibitory factor (MIF).

Macrophage migration inhibitory factor (MIF) is a key innate immune mediator with chemokine- and cytokine-like properties in the inflammatory pathway. While its actions on macrophages are well-studied, its effects on other cell types are less understood. Here we report that MIF is required for expansion of intestinal tuft cells during infection with the helminth Nippostrongylus brasiliensis. MIF-deficient mice show defective innate responses following infection, lacking intestinal epithelial tuft cell hyperplasia or upregulation of goblet cell RELMß, and fail to expand eosinophil, type 2 innate lymphoid cell (ILC2) and macrophage (M2) populations. Similar effects were observed in MIF-sufficient wild-type mice given the MIF inhibitor 4-IPP. MIF had no direct effect on epithelial cells in organoid cultures, and MIF-deficient intestinal stem cells could generate tuft cells in vitro in the presence of type 2 cytokines. In vivo the lack of MIF could be fully compensated by administration of IL-25, restoring tuft cell differentiation and goblet cell expression of RELM-ß, demonstrating its requirement upstream of the ILC2-tuft cell circuit. Both ILC2s and macrophages expressed the MIF receptor CXCR4, indicating that MIF may act as an essential co-factor on both cell types to activate responses to IL-25 in helminth infection.

Characterisation of the secreted apyrase family of Heligmosomoides polygyrus.

Apyrases are a recurrent feature of secretomes from numerous species of parasitic nematodes. Here we characterise the five apyrases secreted by Heligmosomoides polygyrus, a natural parasite of mice and a widely used laboratory model for intestinal nematode infection. All five enzymes are closely related to soluble calcium-activated nucleotidases described in a variety of organisms, and distinct from the CD39 family of ecto-nucleotidases. Expression is maximal in adult worms and restricted to adults and L4s. Recombinant apyrases were produced and purified from Pichia pastoris. The five enzymes showed very similar biochemical properties, with strict calcium dependence and a broad substrate specificity, catalysing the hydrolysis of all nucleoside tri- and diphosphates, with no activity against nucleoside monophosphates. Natural infection of mice provoked very low antibodies to any enzyme, but immunisation with an apyrase cocktail showed partial protection against reinfection, with reduced egg output and parasite recovery. The most likely role for nematode secreted apyrases is hydrolysis of extracellular ATP, which acts as an alarmin for cellular release of IL-33 and initiation of type 2 immunity.

Oral delivery of a functional algal-expressed TGF-ß mimic halts colitis in a murine DSS model.

Inflammatory bowel disease (IBD) is a set of immunological disorders which can generate chronic pain and fatigue associated with the inflammatory symptoms. The treatment of IBD remains a significant hurdle with current therapies being only partially effective or having significant side effects, suggesting that new therapies that elicit different modes of action and delivery strategies are required. TGM1 is a TGF-ß mimic that was discovered from the intestinal helminth parasite Heligmosomoides polygyrus and is thought to be produced by the parasite to suppress the intestinal inflammation response to help evade host immunity, making it an ideal candidate to be developed as a novel anti-inflammatory bio-therapeutic. Here we utilized the expression system of the edible green algae Chlamydomonas reinhardtii in order to recombinantly produce active TGM1 in a form that could be ingested. C. reinhardtii robustly expressed TGM1, and the resultant recombinant protein is biologically active as measured by regulatory T cell induction. When delivered orally to mice, the algal expressed TGM1 is able to ameliorate weight loss, lymphadenopathy, and disease symptoms in a mouse model of DSS-induced colitis, demonstrating the potential of this biologic as a novel treatment of IBD.

Prostaglandin E2 promotes intestinal inflammation via inhibiting microbiota-dependent regulatory T cells.

The gut microbiota fundamentally regulates intestinal homeostasis and disease partially through mechanisms that involve modulation of regulatory T cells (Tregs), yet how the microbiota-Treg cross-talk is physiologically controlled is incompletely defined. Here, we report that prostaglandin E2 (PGE2), a well-known mediator of inflammation, inhibits mucosal Tregs in a manner depending on the gut microbiota. PGE2 through its receptor EP4 diminishes Treg-favorable commensal microbiota. Transfer of the gut microbiota that was modified by PGE2-EP4 signaling modulates mucosal Treg responses and exacerbates intestinal inflammation. Mechanistically, PGE2-modified microbiota regulates intestinal mononuclear phagocytes and type I interferon signaling. Depletion of mononuclear phagocytes or deficiency of type I interferon receptor diminishes PGE2-dependent Treg inhibition. Together, our findings provide emergent evidence that PGE2-mediated disruption of microbiota-Treg communication fosters intestinal inflammation.