Conserved induction of distinct antiviral signalling kinetics by primate interferon lambda 4 proteins

Interferon lambdas (IFNλ) (also known as type III IFNs) are critical cytokines that combat infection predominantly at barrier tissues, such as the lung, liver, and gastrointestinal tract. Humans have four IFNλs (1–4), where IFNλ1–3 show ~80%–95% homology, and IFNλ4 is the most divergent displaying only ~30% sequence identity. Variants in IFNλ4 in humans are associated with the outcome of infection, such as with hepatitis C virus. However, how IFNλ4 variants impact cytokine signalling in other tissues and how well this is conserved is largely unknown. In this study, we address whether differences in antiviral signalling exist between IFNλ4 variants in human hepatocyte and intestinal cells, comparing them to IFNλ3. We demonstrate that compared to IFNλ3, wild-type human IFNλ4 induces a signalling response with distinct magnitudes and kinetics, which is modified by naturally occurring variants P70S and K154E in both cell types. IFNλ4’s distinct antiviral response was more rapid yet transient compared to IFNλ1 and 3. Additionally, divergent antiviral kinetics were also observed using non-human primate IFNλs and cell lines. Furthermore, an IFNλ4-like receptor-interacting interface failed to alter IFNλ1’s kinetics. Together, our data provide further evidence that major functional differences exist within the IFNλ gene family. These results highlight the possible tissue specialisation of IFNλs and encourage further investigation of the divergent, non-redundant activities of IFNλ4 and other IFNλs

Investigation of the crosstalk of different interferon lambdas and lambda-related cytokines in the JAK/STAT pathway

During my PhD, I firstly explored the crosstalk of type I and III interferons (IFNs) signaling. Intestinal epithelial cells (IECs) are primarily responsive to enteric viruses in human intestine. The virus infection results in the induction of both type I and type III IFNs. Subsequently, the IFNs induce a series of antiviral molecules to prevent IECs from viral propagation. Currently, whether there is a crosstalk between these two cytokine pathways remains unsolved. Using either type I or type III receptor-deficient human intestinal epithelial cells, the results showed that the two cytokine pathways are interconnected at the level of ISG induction and potent of antiviral activity. Moreover, in human IECs, type I IFN receptor upregulates type III IFN siganling whereas type III IFN downregulates type I IFN signaling. These findings indicate that human intestinal cells are preferentially protected by type III IFN signaling. Subsequently, I investigated how the newly discovered IFN-λ4 and its variants impact cytokine signaling and the conservation between them. Human IFN-λ4 is very divergent and only shares about 30% homology with IFN-λ1-3. Interestingly, IFN-λ4 variants are related to the outcome of HCV infection in humans. In this study, I determine whether human IFN-λ4 and its variants have differences in antiviral signalling compared to IFN-λ3. My results demonstrate that human IFN-λ4 and its variants P70S and K154E induce a distinct magnitude and kinetics of ISG production in human hepatocyte and intestinal cells. In addition, antiviral response induced by IFN-λ4 is faster yet transient compared to IFN-λ3. Furthermore, the distinct antiviral potency was also found in non-human primate IFN-λs and cell lines. Modifications in IFN-λ1 receptor-interacting interface do not alter its kinetic profile. Together, the results emphasis the possibility of IFN-λs in tissue specialisation. My third project is to explore the interaction between interleukin-22 and IFN-λ. In this project, I employed human cell lines and human intestinal organoids to reveal several important findings about the interaction. I found that co-treatment of IL-22 and IFN-λ can induce more p-STAT1 than individual treatment of IL-22 or IFN-λ, but IL-22 cannot enhance IFN-λ-induced ISGs and antiviral activity in vitro. Using RNA-seq, I found signaling induced by IL-22 and IFN-λ are relatively independent even though they share a receptor and activate the same JAK/STAT pathways. Subsequently, I applied human intestinal organoids to validate the cell proliferation function induced by IL-22 which is found in the RNA-seq. I demonstrate IL-22 can promote the proliferation and regeneration of human intestinal organoids. Notably, IL-22 can promote stem cell proliferation marked by the increase in OLFM4 expression in the organoids. Subsequent smRNAFish in OLFM4 confirms the result in organoids. These results indicate a new finding concerning IL-22 in human intestinal cells and intestinal organoids

Genetic regulation of OAS1 nonsense-mediated decay underlies association with COVID-19 hospitalization in patients of European and African ancestries

The chr12q24.13 locus encoding OAS1-OAS3 antiviral proteins has been associated with coronavirus disease 2019 (COVID-19) susceptibility. Here, we report genetic, functional and clinical insights into this locus in relation to COVID-19 severity. In our analysis of patients of European (n = 2,249) and African (n = 835) ancestries with hospitalized versus nonhospitalized COVID-19, the risk of hospitalized disease was associated with a common OAS1 haplotype, which was also associated with reduced severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) clearance in a clinical trial with pegIFN-lambda 1. Bioinformatic analyses and in vitro studies reveal the functional contribution of two associated OAS1 exonic variants comprising the risk haplotype. Derived human-specific alleles rs10774671-A and rs1131454-A decrease OAS1 protein abundance through allele-specific regulation of splicing and nonsense-mediated decay (NMD). We conclude that decreased OAS1 expression due to a common haplotype contributes to COVID-19 severity. Our results provide insight into molecular mechanisms through which early treatment with interferons could accelerate SARS-CoV-2 clearance and mitigate against severe COVID-19.N