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Interferon-Induced Protein 44 Interacts with Cellular FK506-Binding Protein 5, Negatively Regulates Host Antiviral Responses, and Supports Virus Replication.

ABSTRACT: Using multiple viral systems, and performing silencing approaches, overexpression approaches, and experiments in knockout cells, we report, for the first time, that interferon (IFN)-induced protein 44 (IFI44) positively affects virus production and negatively modulates innate immune responses induced after viral infections. Moreover, IFI44 is able to rescue poly(I·C)- and IFN-mediated inhibition of virus growth. Furthermore, we report a novel interaction of IFI44 with the cellular factor FK506-binding protein 5 (FKBP5), which binds to cellular kinases such as the inhibitor of nuclear factor kappa B (I?B) kinases (IKK?, IKK?, and IKK?). Importantly, in the presence of FKBP5, IFI44 decreases the ability of IKK? to phosphorylate I?B? and the ability of IKK? to phosphorylate interferon regulatory factor 3 (IRF-3), providing a novel mechanism for the function of IFI44 in negatively modulating IFN responses. Remarkably, these new IFI44 functions may have implications for diseases associated with excessive immune signaling and for controlling virus infections mediated by IFN responses.IMPORTANCE Innate immune responses mediated by IFN and inflammatory cytokines are critical for controlling virus replication. Nevertheless, exacerbated innate immune responses could be detrimental for the host and feedback mechanisms are needed to maintain the cellular homeostasis. In this work, we describe a completely novel function for IFI44 in negatively modulating the innate immune responses induced after viral infections. We show that decreasing IFI44 expression by using small interfering RNAs (siRNAs) or by generating knockout (KO) cells impairs virus production and increases the levels of IFN responses. Moreover, we report a novel interaction of IFI44 with the cellular protein FKBP5, which in turn interacts with kinases essential for type I and III IFN induction and signaling, such as the inhibitor of nuclear factor kappa B (I?B) kinases IKK?, IKK?, and IKK?. Our data indicate that binding of IFI44 to FKBP5 decreased the phosphorylation of IRF-3 and I?B? mediated by IKK? and IKK?, respectively, providing a likely explanation for the function of IFI44 in negatively modulating IFN responses. These results provide new insights into the induction of innate immune responses and suggest that IFI44 is a new potential antiviral target for reducing virus replication.

SUBMITTER: DeDiego ML

PROVIDER: S-EPMC6712396 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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Interferon-Induced Protein 44 Interacts with Cellular FK506-Binding Protein 5, Negatively Regulates Host Antiviral Responses, and Supports Virus Replication.

DeDiego Marta L ML Nogales Aitor A Martinez-Sobrido Luis L Topham David J DJ

mBio 20190827 4

Using multiple viral systems, and performing silencing approaches, overexpression approaches, and experiments in knockout cells, we report, for the first time, that interferon (IFN)-induced protein 44 (IFI44) positively affects virus production and negatively modulates innate immune responses induced after viral infections. Moreover, IFI44 is able to rescue poly(I·C)- and IFN-mediated inhibition of virus growth. Furthermore, we report a novel interaction of IFI44 with the cellular factor FK506-b ...[more]

PMID: 31455651

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Project description:Cellular intrinsic immunity, mediated by the expression of an array of interferon-stimulated antiviral genes, is a vital part of host defense. We have previously used a bioinformatic screen to identify two interferon-stimulated genes (ISG) with poorly characterized function, interferon-induced protein 44 (IFI44) and interferon-induced protein 44-like (IFI44L), as potentially being important in respiratory syncytial virus (RSV) infection. Using overexpression systems, CRISPR-Cas9-mediated knockout, and a knockout mouse model, we investigated the antiviral capability of these genes in the control of RSV replication. Overexpression of IFI44 or IFI44L was sufficient to restrict RSV infection at an early time postinfection. Knocking out these genes in mammalian airway epithelial cells increased levels of infection. Both genes express antiproliferative factors that have no effect on RSV attachment but reduce RSV replication in a minigenome assay. The loss of Ifi44 was associated with a more severe infection phenotype in a mouse model of infection. These studies demonstrate a function for IFI44 and IFI44L in controlling RSV infection.IMPORTANCE RSV infects all children under 2 years of age, but only a subset of children get severe disease. We hypothesize that susceptibility to severe RSV necessitating hospitalization in children without predefined risk factors is, in part, mediated at the antiviral gene level. However, there is a large array of antiviral genes, particularly in the ISG family, the mechanism of which is poorly understood. Having previously identified IFI44 and IFI44L as possible genes of interest in a bioinformatic screen, we dissected the function of these two genes in the control of RSV. Through a range of overexpression and knockout studies, we show that the genes are antiviral and antiproliferative. This study is important because IFI44 and IFI44L are upregulated after a wide range of viral infections, and IFI44L can serve as a diagnostic biomarker of viral infection.

Project description:Nonstructural protein 3A of foot-and-mouth disease virus (FMDV) is a partially conserved protein of 153 amino acids that is in most FMDVs examined to date, and it plays important roles in virus replication, virulence, and host range. To better understand the role of 3A during FMDV infection, we used coimmunoprecipitation followed by mass spectrometry to identify host proteins that interact with 3A in FMDV-infected cells. Here, we report that cellular vimentin is a host binding partner for 3A. The 3A-vimentin interaction was further confirmed by coimmunoprecipitation, glutathione S-transferase (GST) pull down, and immunofluorescence assays. Alanine-scanning mutagenesis indicated that amino acid residues 15 to 21 at the N-terminal region of the FMDV 3A are responsible for the interaction between 3A and vimentin. Using reverse genetics, we demonstrate that mutations in 3A that disrupt the interaction between 3A and vimentin are also critical for virus growth. Overexpression of vimentin significantly suppressed the replication of FMDV, whereas knockdown of vimentin significantly enhanced FMDV replication. However, chemical disruption of the vimentin network by acrylamide resulted in a significant decrease in viral yield, suggesting that an intact vimentin network is needed for FMDV replication. These results indicate that vimentin interacts with FMDV 3A and negatively regulates FMDV replication and that the vimentin-3A interaction is essential for FMDV replication. This study provides information that should be helpful for understanding the molecular mechanism of FMDV replication.IMPORTANCE Foot-and-mouth disease virus (FMDV) nonstructural protein 3A plays important roles in virus replication, host range, and virulence. To further understand the role of 3A during FMDV infection, identification of host cell factors that interact with FMDV 3A is needed. Here, we found that vimentin is a direct binding partner of FMDV 3A, and manipulation of vimentin has a negative effect on virus replication. We also demonstrated that amino acid residues 15 to 21 at the N-terminal region of the FMDV 3A are responsible for the interaction between 3A and vimentin and that the 3A-vimentin interaction is critical for viral replication since the full-length cDNA clone harboring mutations in 3A, which were disrupt 3A-vimentin reactivity, could not produce viable virus progeny. This study provides information that not only provides us a better understanding of the mechanism of FMDV replication but also helps in the development of novel antiviral strategies in the future.

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Dataset Information

Interferon-Induced Protein 44 Interacts with Cellular FK506-Binding Protein 5, Negatively Regulates Host Antiviral Responses, and Supports Virus Replication.

Publications

Interferon-Induced Protein 44 Interacts with Cellular FK506-Binding Protein 5, Negatively Regulates Host Antiviral Responses, and Supports Virus Replication.

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