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1.
EMBO J ; 32(23): 3055-65, 2013 Nov 27.
Article in English | MEDLINE | ID: mdl-24169568

ABSTRACT

The IFNL4 gene is a recently discovered type III interferon, which in a significant fraction of the human population harbours a frameshift mutation abolishing the IFNλ4 ORF. The expression of IFNλ4 is correlated with both poor spontaneous clearance of hepatitis C virus (HCV) and poor response to treatment with type I interferon. Here, we show that the IFNL4 gene encodes an active type III interferon, named IFNλ4, which signals through the IFNλR1 and IL-10R2 receptor chains. Recombinant IFNλ4 is antiviral against both HCV and coronaviruses at levels comparable to IFNλ3. However, the secretion of IFNλ4 is impaired compared to that of IFNλ3, and this impairment is not due to a weak signal peptide, which was previously believed. We found that IFNλ4 gets N-linked glycosylated and that this glycosylation is required for secretion. Nevertheless, this glycosylation is not required for activity. Together, these findings result in the paradox that IFNλ4 is strongly antiviral but a disadvantage during HCV infection.


Subject(s)
Antiviral Agents/pharmacology , Coronaviridae Infections/prevention & control , Hepatitis C/prevention & control , Interleukins/metabolism , Receptors, Interferon/metabolism , Receptors, Interleukin/metabolism , Amino Acid Sequence , Blotting, Western , Cell Proliferation , Cells, Cultured , Coronaviridae/pathogenicity , Coronaviridae Infections/metabolism , Coronaviridae Infections/virology , Glycosylation , Hepacivirus/pathogenicity , Hepatitis C/metabolism , Hepatitis C/virology , Humans , Immunoenzyme Techniques , Interferon-gamma/metabolism , Interleukins/chemistry , Interleukins/genetics , Molecular Sequence Data , Protein Conformation , Protein Sorting Signals/physiology , RNA, Messenger/genetics , Real-Time Polymerase Chain Reaction , Receptors, Interferon/genetics , Receptors, Interleukin/genetics , Respiratory System/cytology , Respiratory System/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Sequence Homology, Amino Acid , Virus Replication , Interferon gamma Receptor
2.
mBio ; 4(1): e00611-12, 2013 Feb 19.
Article in English | MEDLINE | ID: mdl-23422412

ABSTRACT

The recent emergence of a novel human coronavirus (HCoV-EMC) in the Middle East raised considerable concerns, as it is associated with severe acute pneumonia, renal failure, and fatal outcome and thus resembles the clinical presentation of severe acute respiratory syndrome (SARS) observed in 2002 and 2003. Like SARS-CoV, HCoV-EMC is of zoonotic origin and closely related to bat coronaviruses. The human airway epithelium (HAE) represents the entry point and primary target tissue for respiratory viruses and is highly relevant for assessing the zoonotic potential of emerging respiratory viruses, such as HCoV-EMC. Here, we show that pseudostratified HAE cultures derived from different donors are highly permissive to HCoV-EMC infection, and by using reverse transcription (RT)-PCR and RNAseq data, we experimentally determined the identity of seven HCoV-EMC subgenomic mRNAs. Although the HAE cells were readily responsive to type I and type III interferon (IFN), we observed neither a pronounced inflammatory cytokine nor any detectable IFN responses following HCoV-EMC, SARS-CoV, or HCoV-229E infection, suggesting that innate immune evasion mechanisms and putative IFN antagonists of HCoV-EMC are operational in the new host. Importantly, however, we demonstrate that both type I and type III IFN can efficiently reduce HCoV-EMC replication in HAE cultures, providing a possible treatment option in cases of suspected HCoV-EMC infection. IMPORTANCE A novel human coronavirus, HCoV-EMC, has recently been described to be associated with severe respiratory tract infection and fatalities, similar to severe acute respiratory syndrome (SARS) observed during the 2002-2003 epidemic. Closely related coronaviruses replicate in bats, suggesting that, like SARS-CoV, HCoV-EMC is of zoonotic origin. Since the animal reservoir and circumstances of zoonotic transmission are yet elusive, it is critically important to assess potential species barriers of HCoV-EMC infection. An important first barrier against invading respiratory pathogens is the epithelium, representing the entry point and primary target tissue of respiratory viruses. We show that human bronchial epithelia are highly susceptible to HCoV-EMC infection. Furthermore, HCoV-EMC, like other coronaviruses, evades innate immune recognition, reflected by the lack of interferon and minimal inflammatory cytokine expression following infection. Importantly, type I and type III interferon treatment can efficiently reduce HCoV-EMC replication in the human airway epithelium, providing a possible avenue for treatment of emerging virus infections.


Subject(s)
Coronaviridae/physiology , Coronaviridae/pathogenicity , Epithelial Cells/virology , Virus Replication , Animals , Coronaviridae/growth & development , Coronaviridae/isolation & purification , Coronavirus Infections/virology , Cytokines/biosynthesis , Humans , Immune Evasion , Middle East , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , RNA, Viral/biosynthesis , RNA, Viral/genetics , Respiratory Tract Infections/virology , Reverse Transcriptase Polymerase Chain Reaction , Sequence Analysis, RNA , Zoonoses/virology
3.
J Biol Chem ; 287(12): 9454-60, 2012 Mar 16.
Article in English | MEDLINE | ID: mdl-22235133

ABSTRACT

IL-21 is a class I cytokine that exerts pleiotropic effects on both innate and adaptive immune responses. It signals through a heterodimeric receptor complex consisting of the IL-21 receptor (IL-21R) and the common γ-chain. A hallmark of the class I cytokine receptors is the class I cytokine receptor signature motif (WSXWS). The exact role of this motif has not been determined yet; however, it has been implicated in diverse functions, including ligand binding, receptor internalization, proper folding, and export, as well as signal transduction. Furthermore, the WXXW motif is known to be a consensus sequence for C-mannosylation. Here, we present the crystal structure of IL-21 bound to IL-21R and reveal that the WSXWS motif of IL-21R is C-mannosylated at the first tryptophan. We furthermore demonstrate that a sugar chain bridges the two fibronectin domains that constitute the extracellular domain of IL-21R and anchors at the WSXWS motif through an extensive hydrogen bonding network, including mannosylation. The glycan thus transforms the V-shaped receptor into an A-frame. This finding offers a novel structural explanation of the role of the class I cytokine signature motif.


Subject(s)
Interleukins/chemistry , Interleukins/metabolism , Receptors, Interleukin-21/chemistry , Receptors, Interleukin-21/metabolism , Amino Acid Motifs , Amino Acid Sequence , Binding Sites , Crystallography, X-Ray , Glycosylation , Humans , Interleukins/genetics , Mannose/metabolism , Models, Molecular , Molecular Sequence Data , Protein Binding , Protein Structure, Secondary , Receptors, Interleukin-21/genetics
4.
J Virol ; 84(3): 1414-22, 2010 Feb.
Article in English | MEDLINE | ID: mdl-19939920

ABSTRACT

In less than 3 months after the first cases of swine origin 2009 influenza A (H1N1) virus infections were reported from Mexico, WHO declared a pandemic. The pandemic virus is antigenically distinct from seasonal influenza viruses, and the majority of human population lacks immunity against this virus. We have studied the activation of innate immune responses in pandemic virus-infected human monocyte-derived dendritic cells (DC) and macrophages. Pandemic A/Finland/553/2009 virus, representing a typical North American/European lineage virus, replicated very well in these cells. The pandemic virus, as well as the seasonal A/Brisbane/59/07 (H1N1) and A/New Caledonia/20/99 (H1N1) viruses, induced type I (alpha/beta interferon [IFN-alpha/beta]) and type III (IFN-lambda1 to -lambda3) IFN, CXCL10, and tumor necrosis factor alpha (TNF-alpha) gene expression weakly in DCs. Mouse-adapted A/WSN/33 (H1N1) and human A/Udorn/72 (H3N2) viruses, instead, induced efficiently the expression of antiviral and proinflammatory genes. Both IFN-alpha and IFN-beta inhibited the replication of the pandemic (H1N1) virus. The potential of IFN-lambda3 to inhibit viral replication was lower than that of type I IFNs. However, the pandemic virus was more sensitive to the antiviral IFN-lambda3 than the seasonal A/Brisbane/59/07 (H1N1) virus. The present study demonstrates that the novel pandemic (H1N1) influenza A virus can readily replicate in human primary DCs and macrophages and efficiently avoid the activation of innate antiviral responses. It is, however, highly sensitive to the antiviral actions of IFNs, which may provide us an additional means to treat severe cases of infection especially if significant drug resistance emerges.


Subject(s)
Antiviral Agents/pharmacology , Cytokines/biosynthesis , Dendritic Cells/metabolism , Influenza A Virus, H1N1 Subtype/physiology , Influenza, Human/epidemiology , Interferon-alpha/pharmacology , Interferon-beta/pharmacology , Macrophages/metabolism , Cells, Cultured , Enzyme Inhibitors/pharmacology , Humans , Influenza A Virus, H1N1 Subtype/drug effects , Influenza, Human/virology , Neuraminidase/antagonists & inhibitors , Reverse Transcriptase Polymerase Chain Reaction , Virus Replication
5.
Pharmaceuticals (Basel) ; 3(4): 795-809, 2010 Mar 25.
Article in English | MEDLINE | ID: mdl-27713280

ABSTRACT

Interferon lambda (IFN-λ) is a member of the class II cytokine family, and like the other members of this family, they are small helical proteins. Since their discovery significant efforts have been made to determine their role in innate and adaptive immunity. Their strong antiviral activity, both in vitro and in vivo, has firmly established their interferon status. However, in contrast to type I interferon, only a very limited subset of cells/tissues responds to interferon lambda. In addition to inducing an antiviral state in responsive cells, recent data suggest that IFN-l plays a role in shaping the adaptive immune response. However, the data is not in complete agreement regarding the effect of IFN-λ on the adaptive immune system. Recently IFN-l has entered clinical trials against hepatitis C Virus and IFN-l is a promising future therapeutic, against different viruses replicating in responsive tissues, like that of the airway epithelia. In this review we describe the knowledge acquired during the past six years about the structure and function of interferon lambda.

6.
J Biol Chem ; 284(31): 20869-75, 2009 Jul 31.
Article in English | MEDLINE | ID: mdl-19457860

ABSTRACT

Interferon-lambda (IFN-lambda) is an antiviral cytokine that signals through a distinct receptor complex, composed of the IFN-lambdaR1 and interleukin-10R2 (IL-10R2) receptor chains. We have determined the crystal structure of human IFN-lambda3 and characterized the interaction with its receptor complex through structure-based site-directed mutagenesis. The ability of IFN-lambda3 mutants to signal was determined by measuring the antiviral activity and induced STAT2 phosphorylation. In conclusion, our data show that, although IFN-lambda is functionally an interferon, it is clearly structurally related to members of the IL-10 family. In particular, we found an interesting similarity between IFN-lambda and IL-22, and we suggest that IFN-lambda and IL-22 possess parallel functions, protecting epithelial tissue against viral and bacterial infections, respectively.


Subject(s)
Interleukin-10/chemistry , Interleukins/chemistry , Interleukins/metabolism , Antiviral Agents/metabolism , Binding Sites , Cell Line, Tumor , Crystallography, X-Ray , Humans , Interferons , Interleukin-10 Receptor beta Subunit/metabolism , Models, Molecular , Mutant Proteins/chemistry , Mutant Proteins/metabolism , Phosphorylation , Protein Binding , Protein Structure, Secondary , STAT2 Transcription Factor/metabolism , Interleukin-22
7.
J Virol ; 81(14): 7749-58, 2007 Jul.
Article in English | MEDLINE | ID: mdl-17507495

ABSTRACT

Type III interferon (IFN) is a novel member of the interferon family. Type III IFN utilizes a receptor complex different from that of type I IFN, but both types of IFN induce STAT1, STAT2, and STAT3 activation. Here we describe a detailed comparison of signal transduction initiated by type I and type III IFN. Gene expression array analysis showed that IFN types I and III induced a similar subset of genes. In particular, no genes were induced uniquely by type III IFN. Next, we used chromatin immunoprecipitation (ChIP) analysis to investigate the promoter activation by types I and III IFN. The ChIP assays demonstrated that stimulation of cells with both type I and type III IFN resulted in the recruitment of ISGF3 transcription factor components to the promoter region of responsive genes and in an increase of polymerase II loading and histone acetylation. Whereas IFN type I signaling was observed for a broad spectrum of cell lines, type III IFN signaling was more restricted. The lack of IFN type III signaling was correlated with a low expression of the IL28Ra component of the IFN type III receptor, and IL28Ra overexpression was sufficient to restore IFN type III signaling. We also tested the activation of mitogen-activated protein (MAP) kinases by type III IFN and found that type III IFN relies strongly upon both p38 and JNK MAP kinases for gene induction.


Subject(s)
Interferons/physiology , Janus Kinases/metabolism , Mitogen-Activated Protein Kinases/metabolism , Signal Transduction , Cell Line , Chromatin Immunoprecipitation , Humans , STAT Transcription Factors
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