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IL-1α is a DNA damage sensor linking genotoxic stress signaling to sterile inflammation and innate immunity.

Idan C, Peleg R, Elena V, Martin T, Cicerone T, Mareike W, Lydia B, Marina F, Gerhard M, Elisa FM, Dinarello CA, Ron AN, Robert S - Sci Rep (2015)

Bottom Line: Environmental signals can be translated into chromatin changes, which alter gene expression.Thus, in addition to its role as a danger signal, which occurs when the cytokine is passively released by cell necrosis, IL-1α could directly sense DNA damage and act as signal for genotoxic stress without loss of cell integrity.Here we demonstrate localization of the cytokine to DNA-damage sites and its subsequent secretion.

View Article: PubMed Central - PubMed

Affiliation: Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS UMR 7104, INSERM U 964, Université de Strasbourg, 67404 Illkirch, France.

ABSTRACT
Environmental signals can be translated into chromatin changes, which alter gene expression. Here we report a novel concept that cells can signal chromatin damage from the nucleus back to the surrounding tissue through the cytokine interleukin-1alpha (IL-1α). Thus, in addition to its role as a danger signal, which occurs when the cytokine is passively released by cell necrosis, IL-1α could directly sense DNA damage and act as signal for genotoxic stress without loss of cell integrity. Here we demonstrate localization of the cytokine to DNA-damage sites and its subsequent secretion. Interestingly, its nucleo-cytosolic shuttling after DNA damage sensing is regulated by histone deacetylases (HDAC) and IL-1α acetylation. To demonstrate the physiological significance of this newly discovered mechanism, we used IL-1α knockout mice and show that IL-1α signaling after UV skin irradiation and DNA damage is important for triggering a sterile inflammatory cascade in vivo that contributes to efficient tissue repair and wound healing.

No MeSH data available.


Related in: MedlinePlus

IL-1α acetylation within the nuclear localization sequence impacts on IL1α subcellular localisation.(a) IL-1α precursor is recognized by a pan acetyl antibody. Endogenous IL-1α was immunoprecipitated (IP) from nuclear extracts of Raw 264.7 cells, either induced or non-induced with 100 ng/ml LPS. Total IP proteins were separated over 15% SDS PAGE, transferred to nitrocellulose membranes and blotted with anti-mouse IL-1α (top panel) or anti-Kac (bottom panel). Acetylated IL-1α is marked by arrows and IP antibody light and heavy chain signals are indicated. (b) Annotated MS/MS spectrum of the tryptic peptide VTVSATSSN(Deam)GK(Acetyl)ILK (MH2 + 724.40 Da) showing acetylation of IL-1α (Uniprot ID P01582) at K82 and N80 deamidation. (c) PrecIL-1α K82 mutants affect IL-1α sub-cellular localization. Confocal microscopic analysis of GFP tagged WT IL-1α and mutations of precIL-1α K82 to glutamine (precIL-1α K82Q, mimicking acetylation) and to arginine (precIL-1α K82R non-acetylateable). White scale bars, 20 μm (d) IL-1α K82 mutations reduce cytokine secretion after DNA damage. Mouse B16 cells were transfected with the indicated GFP IL-1α plasmids. The cells were then subjected to 100 μM H2O2. 16h after stress induction levels of secreted GFP IL-1α in cell growth medium was measured using a GFP ELISA. GFP IL-1α levels in cell lysates were used to normalize for transfection efficiencies and non-transfected cells were used as negative controls. Data are expressed as mean ± SD of three independent experiments. (e) Histone deacetylase inhibition by TSA increases IL-1α nuclear localization. Images of cells expressing GFP IL-1α either non-treated (control) or treated with TSA (100 ng/ml) were collected every hour for 22 h and representative images for three time points (0, 11 and 22 hours) are shown (For averaged fluorescence intensities of nuclear/cytoplasmic ratios see Supplementary Figure 1b). (f) HDAC-1 and IL-1α can co-localize at DNA damage lesions. Cells expressing GFP IL-1α were laser-microirradiated for the induction of DNA damage. Localization of HDAC-1 and IL-1α–GFP were visualized by confocal microscopic analysis.
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f3: IL-1α acetylation within the nuclear localization sequence impacts on IL1α subcellular localisation.(a) IL-1α precursor is recognized by a pan acetyl antibody. Endogenous IL-1α was immunoprecipitated (IP) from nuclear extracts of Raw 264.7 cells, either induced or non-induced with 100 ng/ml LPS. Total IP proteins were separated over 15% SDS PAGE, transferred to nitrocellulose membranes and blotted with anti-mouse IL-1α (top panel) or anti-Kac (bottom panel). Acetylated IL-1α is marked by arrows and IP antibody light and heavy chain signals are indicated. (b) Annotated MS/MS spectrum of the tryptic peptide VTVSATSSN(Deam)GK(Acetyl)ILK (MH2 + 724.40 Da) showing acetylation of IL-1α (Uniprot ID P01582) at K82 and N80 deamidation. (c) PrecIL-1α K82 mutants affect IL-1α sub-cellular localization. Confocal microscopic analysis of GFP tagged WT IL-1α and mutations of precIL-1α K82 to glutamine (precIL-1α K82Q, mimicking acetylation) and to arginine (precIL-1α K82R non-acetylateable). White scale bars, 20 μm (d) IL-1α K82 mutations reduce cytokine secretion after DNA damage. Mouse B16 cells were transfected with the indicated GFP IL-1α plasmids. The cells were then subjected to 100 μM H2O2. 16h after stress induction levels of secreted GFP IL-1α in cell growth medium was measured using a GFP ELISA. GFP IL-1α levels in cell lysates were used to normalize for transfection efficiencies and non-transfected cells were used as negative controls. Data are expressed as mean ± SD of three independent experiments. (e) Histone deacetylase inhibition by TSA increases IL-1α nuclear localization. Images of cells expressing GFP IL-1α either non-treated (control) or treated with TSA (100 ng/ml) were collected every hour for 22 h and representative images for three time points (0, 11 and 22 hours) are shown (For averaged fluorescence intensities of nuclear/cytoplasmic ratios see Supplementary Figure 1b). (f) HDAC-1 and IL-1α can co-localize at DNA damage lesions. Cells expressing GFP IL-1α were laser-microirradiated for the induction of DNA damage. Localization of HDAC-1 and IL-1α–GFP were visualized by confocal microscopic analysis.

Mentions: Since the function and dynamics of many DNA damage response factors and complexes involved in DNA repair are regulated by posttranslational modifications (PTM), we examined the possibility that the cytoplasmic localization of IL-1α after DNA damage is also regulated by PTMs. To investigate the role of PTMs in IL-1α function we searched for novel IL-1α modifications. By using immunoprecipitation (IP), we observed that mouse IL-1α protein is strongly recognized by an anti-Acetyl lysine (KAct) antibody (Fig. 3a), suggesting a novel IL-1α acetylation. To corroborate this finding and to map the acetylation sites we performed mass spectrometry analysis and detected acetylation of lysine K82 within the IL-1α NLS (Fig. 3b). To better understand the role of IL-1α acetylation, we expressed GFP tagged IL-1α mutated on K82 to an arginine (K82R) or glutamine (K82Q), which are non-acetylatable or an acetylation mimic, respectively. Interestingly, expressing the K82Q mutation we detected increased IL-1α nuclear signal, whereas the K82R mutant seems to increase the cytoplasmic IL-1α signal without affecting overall cellular expression levels (Fig. 3c and Supplementary Figure 3a). Furthermore, the K82R mutation resulted in increased IL-1α release from UV irradiated cells, whereas release of the cytokine was lower when IL-1α was mutated to K82Q (Fig. 3d). Together these results strongly suggest that IL-1α acetylation can regulate its cellular localization.


IL-1α is a DNA damage sensor linking genotoxic stress signaling to sterile inflammation and innate immunity.

Idan C, Peleg R, Elena V, Martin T, Cicerone T, Mareike W, Lydia B, Marina F, Gerhard M, Elisa FM, Dinarello CA, Ron AN, Robert S - Sci Rep (2015)

IL-1α acetylation within the nuclear localization sequence impacts on IL1α subcellular localisation.(a) IL-1α precursor is recognized by a pan acetyl antibody. Endogenous IL-1α was immunoprecipitated (IP) from nuclear extracts of Raw 264.7 cells, either induced or non-induced with 100 ng/ml LPS. Total IP proteins were separated over 15% SDS PAGE, transferred to nitrocellulose membranes and blotted with anti-mouse IL-1α (top panel) or anti-Kac (bottom panel). Acetylated IL-1α is marked by arrows and IP antibody light and heavy chain signals are indicated. (b) Annotated MS/MS spectrum of the tryptic peptide VTVSATSSN(Deam)GK(Acetyl)ILK (MH2 + 724.40 Da) showing acetylation of IL-1α (Uniprot ID P01582) at K82 and N80 deamidation. (c) PrecIL-1α K82 mutants affect IL-1α sub-cellular localization. Confocal microscopic analysis of GFP tagged WT IL-1α and mutations of precIL-1α K82 to glutamine (precIL-1α K82Q, mimicking acetylation) and to arginine (precIL-1α K82R non-acetylateable). White scale bars, 20 μm (d) IL-1α K82 mutations reduce cytokine secretion after DNA damage. Mouse B16 cells were transfected with the indicated GFP IL-1α plasmids. The cells were then subjected to 100 μM H2O2. 16h after stress induction levels of secreted GFP IL-1α in cell growth medium was measured using a GFP ELISA. GFP IL-1α levels in cell lysates were used to normalize for transfection efficiencies and non-transfected cells were used as negative controls. Data are expressed as mean ± SD of three independent experiments. (e) Histone deacetylase inhibition by TSA increases IL-1α nuclear localization. Images of cells expressing GFP IL-1α either non-treated (control) or treated with TSA (100 ng/ml) were collected every hour for 22 h and representative images for three time points (0, 11 and 22 hours) are shown (For averaged fluorescence intensities of nuclear/cytoplasmic ratios see Supplementary Figure 1b). (f) HDAC-1 and IL-1α can co-localize at DNA damage lesions. Cells expressing GFP IL-1α were laser-microirradiated for the induction of DNA damage. Localization of HDAC-1 and IL-1α–GFP were visualized by confocal microscopic analysis.
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f3: IL-1α acetylation within the nuclear localization sequence impacts on IL1α subcellular localisation.(a) IL-1α precursor is recognized by a pan acetyl antibody. Endogenous IL-1α was immunoprecipitated (IP) from nuclear extracts of Raw 264.7 cells, either induced or non-induced with 100 ng/ml LPS. Total IP proteins were separated over 15% SDS PAGE, transferred to nitrocellulose membranes and blotted with anti-mouse IL-1α (top panel) or anti-Kac (bottom panel). Acetylated IL-1α is marked by arrows and IP antibody light and heavy chain signals are indicated. (b) Annotated MS/MS spectrum of the tryptic peptide VTVSATSSN(Deam)GK(Acetyl)ILK (MH2 + 724.40 Da) showing acetylation of IL-1α (Uniprot ID P01582) at K82 and N80 deamidation. (c) PrecIL-1α K82 mutants affect IL-1α sub-cellular localization. Confocal microscopic analysis of GFP tagged WT IL-1α and mutations of precIL-1α K82 to glutamine (precIL-1α K82Q, mimicking acetylation) and to arginine (precIL-1α K82R non-acetylateable). White scale bars, 20 μm (d) IL-1α K82 mutations reduce cytokine secretion after DNA damage. Mouse B16 cells were transfected with the indicated GFP IL-1α plasmids. The cells were then subjected to 100 μM H2O2. 16h after stress induction levels of secreted GFP IL-1α in cell growth medium was measured using a GFP ELISA. GFP IL-1α levels in cell lysates were used to normalize for transfection efficiencies and non-transfected cells were used as negative controls. Data are expressed as mean ± SD of three independent experiments. (e) Histone deacetylase inhibition by TSA increases IL-1α nuclear localization. Images of cells expressing GFP IL-1α either non-treated (control) or treated with TSA (100 ng/ml) were collected every hour for 22 h and representative images for three time points (0, 11 and 22 hours) are shown (For averaged fluorescence intensities of nuclear/cytoplasmic ratios see Supplementary Figure 1b). (f) HDAC-1 and IL-1α can co-localize at DNA damage lesions. Cells expressing GFP IL-1α were laser-microirradiated for the induction of DNA damage. Localization of HDAC-1 and IL-1α–GFP were visualized by confocal microscopic analysis.
Mentions: Since the function and dynamics of many DNA damage response factors and complexes involved in DNA repair are regulated by posttranslational modifications (PTM), we examined the possibility that the cytoplasmic localization of IL-1α after DNA damage is also regulated by PTMs. To investigate the role of PTMs in IL-1α function we searched for novel IL-1α modifications. By using immunoprecipitation (IP), we observed that mouse IL-1α protein is strongly recognized by an anti-Acetyl lysine (KAct) antibody (Fig. 3a), suggesting a novel IL-1α acetylation. To corroborate this finding and to map the acetylation sites we performed mass spectrometry analysis and detected acetylation of lysine K82 within the IL-1α NLS (Fig. 3b). To better understand the role of IL-1α acetylation, we expressed GFP tagged IL-1α mutated on K82 to an arginine (K82R) or glutamine (K82Q), which are non-acetylatable or an acetylation mimic, respectively. Interestingly, expressing the K82Q mutation we detected increased IL-1α nuclear signal, whereas the K82R mutant seems to increase the cytoplasmic IL-1α signal without affecting overall cellular expression levels (Fig. 3c and Supplementary Figure 3a). Furthermore, the K82R mutation resulted in increased IL-1α release from UV irradiated cells, whereas release of the cytokine was lower when IL-1α was mutated to K82Q (Fig. 3d). Together these results strongly suggest that IL-1α acetylation can regulate its cellular localization.

Bottom Line: Environmental signals can be translated into chromatin changes, which alter gene expression.Thus, in addition to its role as a danger signal, which occurs when the cytokine is passively released by cell necrosis, IL-1α could directly sense DNA damage and act as signal for genotoxic stress without loss of cell integrity.Here we demonstrate localization of the cytokine to DNA-damage sites and its subsequent secretion.

View Article: PubMed Central - PubMed

Affiliation: Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS UMR 7104, INSERM U 964, Université de Strasbourg, 67404 Illkirch, France.

ABSTRACT
Environmental signals can be translated into chromatin changes, which alter gene expression. Here we report a novel concept that cells can signal chromatin damage from the nucleus back to the surrounding tissue through the cytokine interleukin-1alpha (IL-1α). Thus, in addition to its role as a danger signal, which occurs when the cytokine is passively released by cell necrosis, IL-1α could directly sense DNA damage and act as signal for genotoxic stress without loss of cell integrity. Here we demonstrate localization of the cytokine to DNA-damage sites and its subsequent secretion. Interestingly, its nucleo-cytosolic shuttling after DNA damage sensing is regulated by histone deacetylases (HDAC) and IL-1α acetylation. To demonstrate the physiological significance of this newly discovered mechanism, we used IL-1α knockout mice and show that IL-1α signaling after UV skin irradiation and DNA damage is important for triggering a sterile inflammatory cascade in vivo that contributes to efficient tissue repair and wound healing.

No MeSH data available.


Related in: MedlinePlus