Virus-infected cells secrete a broad range of interferons (IFN) which confer resistance to yet uninfected cells by triggering the synthesis of antiviral factors. The relative contributions of the various IFN subtypes to innate immunity against virus infections remain elusive. IFN-␣, IFN-␤, and other type I IFN molecules signal through a common, universally expressed cell surface receptor, whereas type III IFN (IFN-) uses a distinct cell-typespecific receptor complex for signaling. Using mice lacking functional receptors for type I IFN, type III IFN, or both, we found that IFN-plays an important role in the defense against several human pathogens that infect the respiratory tract, such as influenza A virus, influenza B virus, respiratory syncytial virus, human metapneumovirus, and severe acute respiratory syndrome (SARS) coronavirus. These viruses were more pathogenic and replicated to higher titers in the lungs of mice lacking both IFN receptors than in mice with single IFN receptor defects. In contrast, Lassa fever virus, which infects via the respiratory tract but primarily replicates in the liver, was not influenced by the IFN-receptor defect. Careful analysis revealed that expression of functional IFN-receptor complexes in the lung and intestinal tract is restricted to epithelial cells and a few other, undefined cell types. Interestingly, we found that SARS coronavirus was present in feces from infected mice lacking receptors for both type I and type III IFN but not in those from mice lacking single receptors, supporting the view that IFN-contributes to the control of viral infections in epithelial cells of both respiratory and gastrointestinal tracts.The interferon (IFN) system represents a major element of the innate immune response against viral infections (10,13,14). Virus-induced IFN is a complex mixture of biologically active molecules, which includes type I and type III IFN. Type I IFN consists of 14 different IFN-␣ subtypes in the mouse as well as IFN-␤, IFN-, IFN-ε, and limitin, which all signal through the same universally expressed cell surface receptor complex (IFNAR) (30). Type III IFN includes IFN-1, IFN-2, and IFN-3 (21, 28), of which only the latter two are encoded by genes that are expressed in the mouse (22). Type III IFN uses a distinct receptor complex (IL28R) for signaling (21, 28), which appears to be expressed on only a few cell types, including epithelial cells (29). Binding of type I IFN and type III IFN to their cognate receptor complexes triggers signaling cascades that result in the activation of a large number of genes, many of which encode antiviral proteins (10, 32). Type I IFN and type III IFN trigger highly similar gene expression profiles in responsive cells, suggesting that both IFN types might serve similar functions. However, it has to date been largely unclear to which extent IFN-might contribute to innate immunity.Using knockout mouse strains that lack receptors for type I IFN (IFNAR1 0/0 ), type III IFN (IL28R␣ 0/0 ), or both (IFNAR1 0/0 IL28R␣ 0/0 ), we have recently ...

BackgroundHeterologous gene expression is well established for various prokaryotic model systems. However, low yield, incorrect folding and instability still impede the production of soluble, bioactive proteins. To improve protein production with the Gram-positive host Bacillus subtilis, a secretory expression system was designed that enhances translocation, folding and stability of heterologous proteins, and simplifies purification. Based on the theta-replication plasmid pHT01, a B. subtilis secretory expression vector was constructed that encodes a fusion protein consisting of a signal peptide and a StrepII-tag linked to a SUMO-tag serving as a folding catalyst. The gene of a protein of interest can be translationally fused to the SUMO cassette and an additional 6xHis-tag encoding region. In order to maximize secretory expression of the construct by fitting the signal peptide to the StrepII-SUMO part of the fusion protein, a B. subtilis signal-peptide library was screened with the Escherichia coli alkaline phosphatase PhoA as a reporter.ResultsThe YoaW signal peptide-encoding region (SPyoaW) was identified with highest secretory expression capacity in context with the StrepII-SUMO-tag fusion in a B. subtilis eightfold extracellular protease deletion strain. PhoA activity and fusion protein production was elevated by a factor of approximately five when compared to an α-amylase (AmyQ) signal peptide construct. Replacement of PhoA with a single-chain variable fragment antibody specific for GFP or the B. amyloliquefaciens RNase barnase, respectively, resulted in a similar enhancement of secretory expression, demonstrating universality of the YoaW signal peptide-StrepII-SUMO encoding cassette for secretory expression in B. subtilis. Optimisation of codon usage and culture conditions further increased GFP-specific scFv fusion-protein production, and a simple affinity purification strategy from culture supernatant with removal of the StrepII-SUMO-tag by SenP-processing yielded 4 mg of pure, soluble and active GFP-specific scFv from 1 l of culture under standard laboratory conditions.ConclusionsThe new expression system employing a YoaW signal peptide-StrepII-SUMO fusion will simplify secretory protein production and purification with B. subtilis. It can obviate the need for time consuming individual signal-peptide fitting to maximize yield for many different heterologous proteins of interest.Electronic supplementary materialThe online version of this article (10.1186/s12934-019-1078-0) contains supplementary material, which is available to authorized users.

publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active. Interferon LambdaSS7-1 Differential negative regulation of type I and type III interferons underlies extended antiviral effects of interferon-lambdas

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