Intestinal epithelial cells (IECs) are constantly exposed to commensal flora and pathogen challenges. How IECs regulate their innate immune response to maintain gut homeostasis remains unclear. Interferons (IFNs) are cytokines produced during infections. While type I IFN receptors are ubiquitously expressed, type III IFN receptors are expressed only on epithelial cells. This epithelium specificity strongly suggests exclusive functions at epithelial surfaces, but the relative roles of type I and III IFNs in the establishment of an antiviral innate immune response in human IECs are not clearly defined. Here, we used mini-gut organoids to define the functions of types I and III IFNs to protect the human gut against viral infection. We show that primary non-transformed human IECs, upon viral challenge, upregulate the expression of both type I and type III IFNs at the transcriptional level but only secrete type III IFN in the supernatant. However, human IECs respond to both type I and type III IFNs by producing IFN-stimulated genes that in turn induce an antiviral state. Using genetic ablation of either type I or type III IFN receptors, we show that either IFN can independently restrict virus infection in human IECs. Importantly, we report, for the first time, differences in the mechanisms by which each IFN establishes the antiviral state. Contrary to type I IFN, the antiviral activity induced by type III IFN is strongly dependent on the mitogen-activated protein kinases signaling pathway, suggesting a pathway used by type III IFNs that non-redundantly contributes to the antiviral state. In conclusion, we demonstrate that human intestinal epithelial cells specifically regulate their innate immune response favoring type III IFN-mediated signaling, which allows for efficient protection against pathogens without producing excessive inflammation. Our results strongly suggest that type III IFN constitutes the frontline of antiviral response in the human gut. We propose that mucosal surfaces, particularly the gastrointestinal tract, have evolved to favor type III IFN-mediated response to pathogen infections as it allows for spatial segregation of signaling and moderate production of inflammatory signals which we propose are key to maintain gut homeostasis.

Ebola virus (EBOV) causes Filoviruses cause a severe hemorrhagic fever in humans and nonhuman primates with a mortality in humans of up to 90% (1). Filoviruses include five species, Zaire ebolavirus, Bundibugyo ebolavirus, Sudan ebolavirus, Taï Forest ebolavirus, and Reston ebolavirus, belonging to the genus Ebolavirus, and a single species, Marburg marburgvirus, belonging to the genus Marburgvirus (2). Filovirus outbreaks occur in Central Africa regularly; in 2012, four filovirus outbreaks occurred in Uganda and the Democratic Republic of the Congo (3), and a new outbreak with a previously unknown clade of Ebola virus (EBOV), which belongs to the species Zaire ebolavirus, started in Guinea in the winter of 2013-2014 (4) and spread to Liberia, Sierra Leone, Nigeria, Senegal, Mali, Spain, the United States, and the United Kingdom (5). Because of the extremely high mortality of the disease caused by filoviruses and the lack of approved vaccine and treatments, work with these viruses is performed under the biosafety level 4 (BSL-4) biocontainment.EBOV causes a severe immunosuppression in both humans and nonhuman primate models, characterized by a deficient T cell response, lymphopenia, and T cell apoptosis, despite the lack of infection of T cells (6)(7)(8)(9)(10)(11)(12). On the other hand, dendritic cells (DC), which are the most effective antigen-presenting cells and are infected by EBOV, do not undergo normal maturation despite the infection (13-15). Because DC play a key role in initiation of the adaptive immune response by processing viral antigens and presenting them to naive lymphocytes, the deficient T cell response may be linked to the deficient and/or aberrant stimulation of T cells by the infected DC.EBOV has two proteins, VP35 and VP24, which antagonize the

T1/3IFN response capacity appears strongly developmentally constrained at birth. Infants in whom this negative regulation is strongest manifest increased risk for severe respiratory tract infections during infancy and subsequent persistent wheeze.