ABSTRACT
The innate immune system recognizes viral nucleic acids and stimulates cellular antiviral responses. Intracellular detection of viral RNA is mediated by the Retinoic acid inducible gene (RIG)-I Like Receptor (RLR), leading to production of type I interferon (IFN) and pro-inflammatory cytokines. Once cells are infected with a virus, RIG-I and MDA5 bind to viral RNA and undergo conformational change to transmit a signal through direct interaction with downstream CARD-containing adaptor protein, IFN-ß promoter stimulator-1 (IPS-1, also referred as MAVS/VISA/Cardif). IPS-1 is composed of N-terminal Caspase Activation and Recruitment Domain (CARD), proline-rich domain, intermediate domain, and C-terminal transmembrane (TM) domain. The TM domain of IPS-1 anchors it to the mitochondrial outer membrane. It has been hypothesized that activated RLR triggers the accumulation of IPS-1, which forms oligomer as a scaffold for downstream signal proteins. However, the exact mechanisms of IPS-1-mediated signaling remain controversial. In this study, to reveal the details of IPS-1 signaling, we used an artificial oligomerization system to induce oligomerization of IPS-1 in cells. Artificial oligomerization of IPS-1 activated antiviral signaling without a viral infection. Using this system, we investigated the domain-requirement of IPS-1 for its signaling. We discovered that artificial oligomerization of IPS-1 could overcome the requirement of CARD and the TM domain. Moreover, from deletion- and point-mutant analyses, the C-terminal Tumor necrosis factor Receptor-Associated Factor (TRAF) binding motif of IPS-1 (aa. 453-460) present in the intermediate domain is critical for downstream signal transduction. Our results suggest that IPS-1 oligomerization is essential for the formation of a multiprotein signaling complex and enables downstream activation of transcription factors, Interferon Regulatory Factor 3 (IRF3) and Nuclear Factor-κB (NF-κB), leading to type I IFN and pro-inflammatory cytokine production.
Subject(s)
Adaptor Proteins, Signal Transducing/genetics , DEAD-box RNA Helicases/genetics , Protein Interaction Domains and Motifs/genetics , Adaptor Proteins, Signal Transducing/chemistry , Adaptor Proteins, Signal Transducing/metabolism , Animals , DEAD Box Protein 58 , DEAD-box RNA Helicases/metabolism , Fibroblasts/drug effects , Fibroblasts/metabolism , Fibroblasts/virology , Gene Expression Regulation/drug effects , HEK293 Cells , HeLa Cells , Humans , Interferon Regulatory Factor-3/genetics , Interferon Regulatory Factor-3/metabolism , Interferon Type I/biosynthesis , Interferon Type I/immunology , Mice , NF-kappa B/genetics , NF-kappa B/metabolism , Newcastle disease virus/growth & development , Oligopeptides/pharmacology , Protein Multimerization/drug effects , Protein Structure, Tertiary , Receptors, Immunologic , Signal Transduction/drug effectsABSTRACT
In virus-infected cells, RIG-I-like receptor (RLR) recognizes cytoplasmic viral RNA and triggers innate immune responses including production of type I and III interferon (IFN) and the subsequent expression of IFN-inducible genes. Interferon-beta promoter stimulator 1 (IPS-1, also known as MAVS, VISA and Cardif) is a downstream molecule of RLR and is expressed on the outer membrane of mitochondria. While it is known that the location of IPS-1 is essential to its function, its underlying mechanism is unknown. Our aim in this study was to delineate the function of mitochondria so as to identify more precisely its role in innate immunity. In doing so we discovered that viral infection as well as transfection with 5'ppp-RNA resulted in the redistribution of IPS-1 to form speckle-like aggregates in cells. We further found that Mitofusin 1 (MFN1), a key regulator of mitochondrial fusion and a protein associated with IPS-1 on the outer membrane of mitochondria, positively regulates RLR-mediated innate antiviral responses. Conversely, specific knockdown of MFN1 abrogates both the virus-induced redistribution of IPS-1 and IFN production. Our study suggests that mitochondria participate in the segregation of IPS-1 through their fusion processes.
Subject(s)
Adaptor Proteins, Signal Transducing/immunology , GTP Phosphohydrolases/immunology , Membrane Transport Proteins/immunology , Mitochondrial Proteins/immunology , Adaptor Proteins, Signal Transducing/metabolism , Animals , Cell Line , Humans , Immunity, Innate , Mice , Mitochondria/immunology , Mitochondria/pathology , Mitochondrial Membrane Transport Proteins , RNA, ViralABSTRACT
OBJECTIVE: The purpose of this study was to clarify whether alcohol drinking patterns were associated with sleep quality. METHODS: A cross-sectional survey was carried out by self-administered questionnaire in 2008 among 2,118 employees aged 18 to 65 years working in local government in Toyama. After excluding those without relevant data for this study, 661 men and 618 women represented the study population. Logistic regression analysis was used to evaluate whether alcohol drinking patterns (as measured by the frequency, the amount of alcohol per day and the timing of alcohol drinking) were associated with poor sleep quality (as measured by the Pittsburgh Sleep Quality Index), after adjustment for potential confounding factors: age, family structure, work characteristics (as measured by the job-demand-control-support model, shift work and occupational class), chronic disease, body mass index, smoking status and physical activity. RESULTS: In comparison with men who did not drink, the adjusted odds ratio for poor sleep quality was 0.52 (95%confidence interval: 0.32-0.85) for those who drank alcohol once a week or more, 0.32 (0.13-0.84) for those who drank 1-3 glasses daily, 0.30 (0.13-0.70) for those who drank 7-14 glasses per week, 0.37 (0.17-0.77) for those who drank only at meals. In women, the drinking patterns were not significantly associated with sleep quality. CONCLUSIONS: The results of this study suggest that some alcohol drinking patterns may affect sleep quality among men who do not use sleeping medicine.
