Limits...
Loss of the lupus autoantigen Ro52/Trim21 induces tissue inflammation and systemic autoimmunity by disregulating the IL-23-Th17 pathway.

Espinosa A, Dardalhon V, Brauner S, Ambrosi A, Higgs R, Quintana FJ, Sjöstrand M, Eloranta ML, Ní Gabhann J, Winqvist O, Sundelin B, Jefferies CA, Rozell B, Kuchroo VK, Wahren-Herlenius M - J. Exp. Med. (2009)

Bottom Line: Ro52/Trim21 is targeted as an autoantigen in systemic lupus erythematosus and Sjögren's syndrome.Ro52, which was recently identified as an E3 ligase, mediates ubiquitination of several members of the interferon regulatory factor (IRF) family, and the Ro52-deficient mice have an enhanced production of proinflammatory cytokines that are regulated by the IRF transcription factors, including cytokines involved in the Th17 pathway (interleukin [IL] 6, IL-12/IL-23p40, and IL-17).These data reveal that the lupus-associated Ro52 protein is an important negative regulator of proinflammatory cytokine production, and they provide a mechanism by which a defective Ro52 function can lead to tissue inflammation and systemic autoimmunity through the IL-23-Th17 pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Karolinska Institute, Stockholm SE-171 77, Sweden.

ABSTRACT
Ro52/Trim21 is targeted as an autoantigen in systemic lupus erythematosus and Sjögren's syndrome. Polymorphisms in the Ro52 gene have been linked to these autoimmune conditions, but the molecular mechanism by which Ro52 may promote development of systemic autoimmune diseases has not been explored. To address this issue, we generated Ro52- mice (Ro52(-/-)), which appear phenotypically normal if left unmanipulated. However, Ro52(-/-) mice develop severe dermatitis extending from the site of tissue injury induced by ear tags. The affected mice further develop several signs of systemic lupus with hypergammaglobulinemia, autoantibodies to DNA, proteinuria, and kidney pathology. Ro52, which was recently identified as an E3 ligase, mediates ubiquitination of several members of the interferon regulatory factor (IRF) family, and the Ro52-deficient mice have an enhanced production of proinflammatory cytokines that are regulated by the IRF transcription factors, including cytokines involved in the Th17 pathway (interleukin [IL] 6, IL-12/IL-23p40, and IL-17). Loss of IL-23/IL-17 by genetic deletion of IL-23/p19 in the Ro52(-/-) mice conferred protection from skin disease and systemic autoimmunity. These data reveal that the lupus-associated Ro52 protein is an important negative regulator of proinflammatory cytokine production, and they provide a mechanism by which a defective Ro52 function can lead to tissue inflammation and systemic autoimmunity through the IL-23-Th17 pathway.

Show MeSH

Related in: MedlinePlus

Ro52 is expressed in the immune system and induced by IFNs. (A) Targeting construct for generating Ro52−/− mice. Primer pairs “a” and “b” were used for genotyping WT and targeted allele, respectively. (B) Genotyping PCR for WT (340 bp) and Ro52- (700 bp) alleles by PCR in Ro52+/+, Ro52+/−, and Ro52−/− mice. (C) Immunoblotting for Ro52 and GFP expression in splenocyte cell extracts demonstrates loss of Ro52 expression and gain of GFP expression in Ro52+/− and Ro52−/− mice. Data are representative for three independent experiments. (D) Ro52 is specifically expressed in immune tissues as shown by GFP expression (brown) in organ sections (five per organ) from Ro52+/− mice investigated by immunohistochemistry using an anti-GFP antibody. Slides were counterstained with hematoxylin. Data are representative of five independent mice and represent one of two experimental repeats. The original magnification was 10×. Bar, 100 µm. (E) Ro52 is expressed in all major leukocyte populations, as shown by GFP expression in Ro52+/− splenocytes using flow cytometry. The numbers in the quadrants represent the percentage of cells in each quadrant and are representative of 12 mice analyzed in four independent experiments. (F) Ro52 is induced by type I and II IFNs. GFP expression in cells from bone marrow of Ro52+/− mice cultured for 24 h with medium (gray line) or medium + cytokine (black line). Filled histogram represents cells from Ro52+/+ mice. Data are representative of three independent experiments.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC2722164&req=5

fig1: Ro52 is expressed in the immune system and induced by IFNs. (A) Targeting construct for generating Ro52−/− mice. Primer pairs “a” and “b” were used for genotyping WT and targeted allele, respectively. (B) Genotyping PCR for WT (340 bp) and Ro52- (700 bp) alleles by PCR in Ro52+/+, Ro52+/−, and Ro52−/− mice. (C) Immunoblotting for Ro52 and GFP expression in splenocyte cell extracts demonstrates loss of Ro52 expression and gain of GFP expression in Ro52+/− and Ro52−/− mice. Data are representative for three independent experiments. (D) Ro52 is specifically expressed in immune tissues as shown by GFP expression (brown) in organ sections (five per organ) from Ro52+/− mice investigated by immunohistochemistry using an anti-GFP antibody. Slides were counterstained with hematoxylin. Data are representative of five independent mice and represent one of two experimental repeats. The original magnification was 10×. Bar, 100 µm. (E) Ro52 is expressed in all major leukocyte populations, as shown by GFP expression in Ro52+/− splenocytes using flow cytometry. The numbers in the quadrants represent the percentage of cells in each quadrant and are representative of 12 mice analyzed in four independent experiments. (F) Ro52 is induced by type I and II IFNs. GFP expression in cells from bone marrow of Ro52+/− mice cultured for 24 h with medium (gray line) or medium + cytokine (black line). Filled histogram represents cells from Ro52+/+ mice. Data are representative of three independent experiments.

Mentions: We inserted an IRES-GFP reporter cassette into the Ro52 locus to generate Ro52-deficient mice in which the GFP expression allowed us to track Ro52-expressing cells in vivo (Fig. 1, A–C). Tissue specificity of Ro52 expression has not been investigated systematically, and using GFP as a reporter for Ro52 expression, we observed that Ro52 was predominantly expressed in the lymphoid compartments of spleen, lymph node, and thymus, with little or no expression in nonimmune tissues including liver, pancreas, heart, kidney, and skin (Fig. 1 D and Fig. S1). Some expression was observed in endothelial cells (Fig. S1). Our findings are supported by microarray analysis, showing that Ro52 messenger RNA is predominantly expressed in lymphoid cell populations in both mice and humans (http://biogps.gnf.org). These observations suggest that the primary function of Ro52 may be in the immune system. The vast majority of leukocytes in the bone marrow, thymus, spleen, lymph nodes, and peripheral blood expressed Ro52, as detected by GFP expression, with the highest expression in CD3+ and CD11b+GR-1+ cells (Fig. 1 E and not depicted). Using GFP as the reporter, we noted that, as has previously been reported (Rhodes et al., 2002; Strandberg et al., 2008), the expression of Ro52 was induced by IFN-γ and, to a lesser extent, by IFN-α and IFN-β (Fig. 1 F). Other cytokines (TGF-β or TNF-α) did not affect the expression of Ro52 (Fig. 1 F).


Loss of the lupus autoantigen Ro52/Trim21 induces tissue inflammation and systemic autoimmunity by disregulating the IL-23-Th17 pathway.

Espinosa A, Dardalhon V, Brauner S, Ambrosi A, Higgs R, Quintana FJ, Sjöstrand M, Eloranta ML, Ní Gabhann J, Winqvist O, Sundelin B, Jefferies CA, Rozell B, Kuchroo VK, Wahren-Herlenius M - J. Exp. Med. (2009)

Ro52 is expressed in the immune system and induced by IFNs. (A) Targeting construct for generating Ro52−/− mice. Primer pairs “a” and “b” were used for genotyping WT and targeted allele, respectively. (B) Genotyping PCR for WT (340 bp) and Ro52- (700 bp) alleles by PCR in Ro52+/+, Ro52+/−, and Ro52−/− mice. (C) Immunoblotting for Ro52 and GFP expression in splenocyte cell extracts demonstrates loss of Ro52 expression and gain of GFP expression in Ro52+/− and Ro52−/− mice. Data are representative for three independent experiments. (D) Ro52 is specifically expressed in immune tissues as shown by GFP expression (brown) in organ sections (five per organ) from Ro52+/− mice investigated by immunohistochemistry using an anti-GFP antibody. Slides were counterstained with hematoxylin. Data are representative of five independent mice and represent one of two experimental repeats. The original magnification was 10×. Bar, 100 µm. (E) Ro52 is expressed in all major leukocyte populations, as shown by GFP expression in Ro52+/− splenocytes using flow cytometry. The numbers in the quadrants represent the percentage of cells in each quadrant and are representative of 12 mice analyzed in four independent experiments. (F) Ro52 is induced by type I and II IFNs. GFP expression in cells from bone marrow of Ro52+/− mice cultured for 24 h with medium (gray line) or medium + cytokine (black line). Filled histogram represents cells from Ro52+/+ mice. Data are representative of three independent experiments.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2722164&req=5

fig1: Ro52 is expressed in the immune system and induced by IFNs. (A) Targeting construct for generating Ro52−/− mice. Primer pairs “a” and “b” were used for genotyping WT and targeted allele, respectively. (B) Genotyping PCR for WT (340 bp) and Ro52- (700 bp) alleles by PCR in Ro52+/+, Ro52+/−, and Ro52−/− mice. (C) Immunoblotting for Ro52 and GFP expression in splenocyte cell extracts demonstrates loss of Ro52 expression and gain of GFP expression in Ro52+/− and Ro52−/− mice. Data are representative for three independent experiments. (D) Ro52 is specifically expressed in immune tissues as shown by GFP expression (brown) in organ sections (five per organ) from Ro52+/− mice investigated by immunohistochemistry using an anti-GFP antibody. Slides were counterstained with hematoxylin. Data are representative of five independent mice and represent one of two experimental repeats. The original magnification was 10×. Bar, 100 µm. (E) Ro52 is expressed in all major leukocyte populations, as shown by GFP expression in Ro52+/− splenocytes using flow cytometry. The numbers in the quadrants represent the percentage of cells in each quadrant and are representative of 12 mice analyzed in four independent experiments. (F) Ro52 is induced by type I and II IFNs. GFP expression in cells from bone marrow of Ro52+/− mice cultured for 24 h with medium (gray line) or medium + cytokine (black line). Filled histogram represents cells from Ro52+/+ mice. Data are representative of three independent experiments.
Mentions: We inserted an IRES-GFP reporter cassette into the Ro52 locus to generate Ro52-deficient mice in which the GFP expression allowed us to track Ro52-expressing cells in vivo (Fig. 1, A–C). Tissue specificity of Ro52 expression has not been investigated systematically, and using GFP as a reporter for Ro52 expression, we observed that Ro52 was predominantly expressed in the lymphoid compartments of spleen, lymph node, and thymus, with little or no expression in nonimmune tissues including liver, pancreas, heart, kidney, and skin (Fig. 1 D and Fig. S1). Some expression was observed in endothelial cells (Fig. S1). Our findings are supported by microarray analysis, showing that Ro52 messenger RNA is predominantly expressed in lymphoid cell populations in both mice and humans (http://biogps.gnf.org). These observations suggest that the primary function of Ro52 may be in the immune system. The vast majority of leukocytes in the bone marrow, thymus, spleen, lymph nodes, and peripheral blood expressed Ro52, as detected by GFP expression, with the highest expression in CD3+ and CD11b+GR-1+ cells (Fig. 1 E and not depicted). Using GFP as the reporter, we noted that, as has previously been reported (Rhodes et al., 2002; Strandberg et al., 2008), the expression of Ro52 was induced by IFN-γ and, to a lesser extent, by IFN-α and IFN-β (Fig. 1 F). Other cytokines (TGF-β or TNF-α) did not affect the expression of Ro52 (Fig. 1 F).

Bottom Line: Ro52/Trim21 is targeted as an autoantigen in systemic lupus erythematosus and Sjögren's syndrome.Ro52, which was recently identified as an E3 ligase, mediates ubiquitination of several members of the interferon regulatory factor (IRF) family, and the Ro52-deficient mice have an enhanced production of proinflammatory cytokines that are regulated by the IRF transcription factors, including cytokines involved in the Th17 pathway (interleukin [IL] 6, IL-12/IL-23p40, and IL-17).These data reveal that the lupus-associated Ro52 protein is an important negative regulator of proinflammatory cytokine production, and they provide a mechanism by which a defective Ro52 function can lead to tissue inflammation and systemic autoimmunity through the IL-23-Th17 pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Karolinska Institute, Stockholm SE-171 77, Sweden.

ABSTRACT
Ro52/Trim21 is targeted as an autoantigen in systemic lupus erythematosus and Sjögren's syndrome. Polymorphisms in the Ro52 gene have been linked to these autoimmune conditions, but the molecular mechanism by which Ro52 may promote development of systemic autoimmune diseases has not been explored. To address this issue, we generated Ro52- mice (Ro52(-/-)), which appear phenotypically normal if left unmanipulated. However, Ro52(-/-) mice develop severe dermatitis extending from the site of tissue injury induced by ear tags. The affected mice further develop several signs of systemic lupus with hypergammaglobulinemia, autoantibodies to DNA, proteinuria, and kidney pathology. Ro52, which was recently identified as an E3 ligase, mediates ubiquitination of several members of the interferon regulatory factor (IRF) family, and the Ro52-deficient mice have an enhanced production of proinflammatory cytokines that are regulated by the IRF transcription factors, including cytokines involved in the Th17 pathway (interleukin [IL] 6, IL-12/IL-23p40, and IL-17). Loss of IL-23/IL-17 by genetic deletion of IL-23/p19 in the Ro52(-/-) mice conferred protection from skin disease and systemic autoimmunity. These data reveal that the lupus-associated Ro52 protein is an important negative regulator of proinflammatory cytokine production, and they provide a mechanism by which a defective Ro52 function can lead to tissue inflammation and systemic autoimmunity through the IL-23-Th17 pathway.

Show MeSH
Related in: MedlinePlus