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Virus-infection or 5'ppp-RNA activates antiviral signal through redistribution of IPS-1 mediated by MFN1.

Onoguchi K, Onomoto K, Takamatsu S, Jogi M, Takemura A, Morimoto S, Julkunen I, Namiki H, Yoneyama M, Fujita T - PLoS Pathog. (2010)

Bottom Line: 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.Our study suggests that mitochondria participate in the segregation of IPS-1 through their fusion processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics, Institute for Virus Research, Kyoto University, Kyoto, Japan.

ABSTRACT
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.

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Redistribution of IPS-1 in SeV-infected cells.A, The IPS-1-HeLa clone #2 was mock-treated or infected with SeV for 12 h and stained with MitoTracker (Mitochondria) and anti-FLAG antibody (FLAG-IPS-1). Nuclei were visualized by staining with DAPI throughout this study. The fluorescent image was quantified in the area indicated by blue line (right most panel). Quantification results from mock- or SeV-infected cells are shown at the bottom. Fluorescence of DAPI corresponds to area in the nucleus. The mitochondria heavily stained with MitoTracker but lightly stained with anti-FLAG are shown by arrows. B, IPS-1-HeLa cells were mock-treated or infected with SeV for 12 h. Cells were stained with anti-FLAG antibody (FLAG-IPS-1) and anti-ERAB antibody (ERAB).
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ppat-1001012-g002: Redistribution of IPS-1 in SeV-infected cells.A, The IPS-1-HeLa clone #2 was mock-treated or infected with SeV for 12 h and stained with MitoTracker (Mitochondria) and anti-FLAG antibody (FLAG-IPS-1). Nuclei were visualized by staining with DAPI throughout this study. The fluorescent image was quantified in the area indicated by blue line (right most panel). Quantification results from mock- or SeV-infected cells are shown at the bottom. Fluorescence of DAPI corresponds to area in the nucleus. The mitochondria heavily stained with MitoTracker but lightly stained with anti-FLAG are shown by arrows. B, IPS-1-HeLa cells were mock-treated or infected with SeV for 12 h. Cells were stained with anti-FLAG antibody (FLAG-IPS-1) and anti-ERAB antibody (ERAB).

Mentions: Like endogenous IPS-1, FLAG-tagged IPS-1 is expressed on mitochondria in uninfected cells as shown by co-staining with MitoTracker (Fig. 2A, Mock). However, compared to the even cytoplasmic staining in the mock-infected cells, the staining pattern of IPS-1 became noticeably speckled in SeV-infected cells (Fig. 2A, SeV). Quantification of the fluorescence image revealed that mitochondria heavily stained with MitoTracker but lightly stained with anti-FLAG antibody were produced in SeV-infected cells. This redistribution was also observed with another mitochondrial marker, endoplasmic reticulum-associated amyloid β-peptide-binding protein (ERAB) (Fig. 2B), and different viruses (Newcastle disease virus (NDV), Sindbis virus, EMCV, Influenza virus, and Vesicular stomatitis virus (VSV)) (Fig. 3).


Virus-infection or 5'ppp-RNA activates antiviral signal through redistribution of IPS-1 mediated by MFN1.

Onoguchi K, Onomoto K, Takamatsu S, Jogi M, Takemura A, Morimoto S, Julkunen I, Namiki H, Yoneyama M, Fujita T - PLoS Pathog. (2010)

Redistribution of IPS-1 in SeV-infected cells.A, The IPS-1-HeLa clone #2 was mock-treated or infected with SeV for 12 h and stained with MitoTracker (Mitochondria) and anti-FLAG antibody (FLAG-IPS-1). Nuclei were visualized by staining with DAPI throughout this study. The fluorescent image was quantified in the area indicated by blue line (right most panel). Quantification results from mock- or SeV-infected cells are shown at the bottom. Fluorescence of DAPI corresponds to area in the nucleus. The mitochondria heavily stained with MitoTracker but lightly stained with anti-FLAG are shown by arrows. B, IPS-1-HeLa cells were mock-treated or infected with SeV for 12 h. Cells were stained with anti-FLAG antibody (FLAG-IPS-1) and anti-ERAB antibody (ERAB).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2908619&req=5

ppat-1001012-g002: Redistribution of IPS-1 in SeV-infected cells.A, The IPS-1-HeLa clone #2 was mock-treated or infected with SeV for 12 h and stained with MitoTracker (Mitochondria) and anti-FLAG antibody (FLAG-IPS-1). Nuclei were visualized by staining with DAPI throughout this study. The fluorescent image was quantified in the area indicated by blue line (right most panel). Quantification results from mock- or SeV-infected cells are shown at the bottom. Fluorescence of DAPI corresponds to area in the nucleus. The mitochondria heavily stained with MitoTracker but lightly stained with anti-FLAG are shown by arrows. B, IPS-1-HeLa cells were mock-treated or infected with SeV for 12 h. Cells were stained with anti-FLAG antibody (FLAG-IPS-1) and anti-ERAB antibody (ERAB).
Mentions: Like endogenous IPS-1, FLAG-tagged IPS-1 is expressed on mitochondria in uninfected cells as shown by co-staining with MitoTracker (Fig. 2A, Mock). However, compared to the even cytoplasmic staining in the mock-infected cells, the staining pattern of IPS-1 became noticeably speckled in SeV-infected cells (Fig. 2A, SeV). Quantification of the fluorescence image revealed that mitochondria heavily stained with MitoTracker but lightly stained with anti-FLAG antibody were produced in SeV-infected cells. This redistribution was also observed with another mitochondrial marker, endoplasmic reticulum-associated amyloid β-peptide-binding protein (ERAB) (Fig. 2B), and different viruses (Newcastle disease virus (NDV), Sindbis virus, EMCV, Influenza virus, and Vesicular stomatitis virus (VSV)) (Fig. 3).

Bottom Line: 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.Our study suggests that mitochondria participate in the segregation of IPS-1 through their fusion processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics, Institute for Virus Research, Kyoto University, Kyoto, Japan.

ABSTRACT
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.

Show MeSH
Related in: MedlinePlus