The TLR3/IRF1/Type III IFN Axis Facilitates Antiviral Responses against Enterovirus Infections in the Intestine

Abstract

Enteroviruses infect gastrointestinal epithelium cells, cause multiple human diseases, and present public health risks worldwide. However, the mechanisms underlying host immune responses in intestinal mucosa against the early enterovirus infections remain elusive. Here, we showed that human enteroviruses including enterovirus 71 (EV71), coxsackievirus B3 (CVB3), and poliovirus 1 (PV1) predominantly induce type III interferons (IFN-λ1 and IFN-λ2/3), rather than type I interferons (IFN-α and IFN-β), in cultured human normal and cancerous intestine epithelial cells (IECs), mouse intestine tissues, and human clinical intestine specimens. Mechanistic studies demonstrated that IFN-λ production is induced upon enterovirus infection through the Toll-like receptor 3/interferon regulatory factor 1 (TLR3/IRF1) signaling pathway in IECs. In turn, the supplementation of IFN-λ subsequently induces intrinsically antiviral responses against enterovirus replication. Notably, intraperitoneal injection in neonatal C57BL/6J mice with mouse recombinant IFN-λ2 protein represses EV71 replication and protects mice from viral lethal effects. Altogether, these results revealed a distinct mechanism by which the host elicited immune responses against enterovirus infections in intestine through activating the TLR3/IRF1/type III IFN axis. The new findings would provide an antiviral strategy for the prevention and treatment of enterovirus infections and associated diseases. IMPORTANCE Enterovirus infections are significant sources of human diseases and public health risks worldwide, but little is known about the mechanism of innate immune response in host intestine epithelial surface during the viral replication. We reported the epithelial immune response in cultured human normal and cancerous cells (IECs), mouse tissues, and human clinical intestine specimens following infection with enterovirus 71. The results mechanistically revealed type III interferons (IFN-λ1 and IFN-λ2/3), rather than type I interferons (IFN-α and IFN-β), as the dominant production through TLR3/IRF1 signaling upon multiple human enterovirus infection, including enterovirus 71 (EV71), coxsackievirus B3 (CVB3), and poliovirus 1 (PV1). IFN-λ subsequently induced antiviral activity against enterovirus replication in vitro and in vivo. These studies uncovered the role of the novel process of type III IFN production involved in the TLR3/IRF1 pathway in host intestine upon enterovirus infection, which highlighted a regulatory manner of antiviral defense in intestine during enterovirus infection.

Inhibition of Type III Interferon Expression in Intestinal Epithelial Cells—A Strategy Used by Coxsackie B Virus to Evade the Host’s Innate Immune Response at the Primary Site of Infection?

“…Intracellular receptors such as MDA5 and RIG-I may be key in the detection of cytosolic dsRNA including poly(I:C) delivered via transfection. Other studies have suggested that IECs, including HT-29 and CaCo-2 cells can respond to longer poly(I:C) exposure than those used in the present study (6 h) [56]. Based on the aforementioned and our present observations it is possible to hypothesize that IECs are slightly more inert to exposure with PAMPs and less likely to respond to dsRNA (e.g., poly (I:C)) to avoid the continual induction of an inflammatory state in the gut.…”

Rotavirus NSP1 Inhibits Type I and Type III Interferon Induction

“…Indeed, porcine epidemic diarrhea virus (PEDV) which also replicates in IECs, has been shown to inhibit IRF-1-dependent type III IFN production [ 52 ]. Furthermore, a recent report demonstrated a role for IRF-1 in the upregulation of type III IFNs in human IECs in response to enterovirus infection, confirming the relevance of this transcription factor in the cells that support RV replication [ 53 ].…”

Comparative analysis of public RNA-sequencing data from human intestinal enteroid (HIEs) infected with enteric RNA viruses identifies universal and virus-specific epithelial responses

“…This highlights a caveat of transcriptomic analysis, whereby the findings are influenced by the depth of sequencing required to pick up, in this case, these specific IFN genes. For example, mammalian reovirus infected T84 intestinal cells and enterovirus 71-infected HT29 cells express lower levels of IFNβ transcripts compared to IFNλ [21, 22]. As such the level of IFNα and/or IFNβ production in the HIEs could have been below the limit of detection of RNA-sequencing.…”

“…While TLR1, 2 and 3 were detectable across all datasets, they were only differentially expressed in response to norovirus infection. TLR3 is primarily present in the endosomal compartment and senses double-stranded (ds) RNA [22, 23]. DsRNA is a RNA virus replication intermediate present during infection with all three viruses.…”

“…Thus, TLR3 could conceivably function in the antiviral response to all three viruses. However, TLR3 may only be differentially expressed in the norovirus-infected HIEs dataset due to the timing of sample collection (48 hours post-infection) since an amplification of the baseline expression level of TLR3 during viral infection of intestinal epithelial cells is part of the ISG response [22]. It is also conceivable that TLR3 might play a more unique role in the epithelial response to human norovirus rather than astrovirus and rotavirus.…”