Anti-Inflammatory Cytokine Interleukin-4 Inhibits Inducible Nitric Oxide Synthase Gene Expression in the Mouse Macrophage Cell Line RAW264.7 through the Repression of Octamer-Dependent Transcription
Inducible nitric oxide synthase (iNOS) is a signature molecule involved in the classical activation of M1 macrophages and is induced by the Nos2 gene upon stimulation with Th1-cell derived interferon-gamma (IFNγ) and bacterial lipopolysaccharide (LPS). Although the anti-inflammatory cytokine IL-4 is...
Saved in:
| Main Authors: | , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Wiley
2013-01-01
|
| Series: | Mediators of Inflammation |
| Online Access: | http://dx.doi.org/10.1155/2013/369693 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1850234551242063872 |
|---|---|
| author | Miki Hiroi Yoshiichi Sakaeda Hana Yamaguchi Yoshihiro Ohmori |
| author_facet | Miki Hiroi Yoshiichi Sakaeda Hana Yamaguchi Yoshihiro Ohmori |
| author_sort | Miki Hiroi |
| collection | DOAJ |
| description | Inducible nitric oxide synthase (iNOS) is a signature molecule involved in the classical activation of M1 macrophages and is induced by the Nos2 gene upon stimulation with Th1-cell derived interferon-gamma (IFNγ) and bacterial lipopolysaccharide (LPS). Although the anti-inflammatory cytokine IL-4 is known to inhibit Nos2 gene expression, the molecular mechanism involved in the negative regulation of Nos2 by IL-4 remains to be fully elucidated. In the present study, we investigated the mechanism of IL-4-mediated Nos2 transcriptional repression in the mouse macrophage-like cell line RAW264.7. Signal transducer and activator of transcription 6 (Stat6) knockdown by siRNA abolished the IL-4-mediated inhibition of Nos2 induced by IFNγ/LPS. Transient transfection of a luciferase reporter gene containing the 5′-flanking region of the Nos2 gene demonstrated that an octamer transcription factor (OCT) binding site in the promoter region is required for both positive regulation by IFNγ/LPS and negative regulation by IL-4. Although IL-4 had no inhibitory effect on the DNA-binding activity of constitutively expressed Oct-1, IL-4-induced Nos2-reporter transcriptional repression was partially attenuated by overexpression of the coactivator CREB-binding protein (CBP). These results suggest that a coactivator/cofactor that functionally interacts with Oct-1 is a molecular target for the IL-4-mediated inhibition of Nos2 and that IL-4-activated Stat6 represses Oct-1-dependent transcription by competing with this coactivator/cofactor. |
| format | Article |
| id | doaj-art-0ea657f2edf247818c9c1e4f24d70f4b |
| institution | OA Journals |
| issn | 0962-9351 1466-1861 |
| language | English |
| publishDate | 2013-01-01 |
| publisher | Wiley |
| record_format | Article |
| series | Mediators of Inflammation |
| spelling | doaj-art-0ea657f2edf247818c9c1e4f24d70f4b2025-08-20T02:02:37ZengWileyMediators of Inflammation0962-93511466-18612013-01-01201310.1155/2013/369693369693Anti-Inflammatory Cytokine Interleukin-4 Inhibits Inducible Nitric Oxide Synthase Gene Expression in the Mouse Macrophage Cell Line RAW264.7 through the Repression of Octamer-Dependent TranscriptionMiki Hiroi0Yoshiichi Sakaeda1Hana Yamaguchi2Yoshihiro Ohmori3Division of Microbiology and Immunology, Department of Oral Biology and Tissue Engineering, Meikai University School of Dentistry, 1-1 Keyakidai, Sakado 350-0283, Saitama, JapanDivision of Microbiology and Immunology, Department of Oral Biology and Tissue Engineering, Meikai University School of Dentistry, 1-1 Keyakidai, Sakado 350-0283, Saitama, JapanDivision of Microbiology and Immunology, Department of Oral Biology and Tissue Engineering, Meikai University School of Dentistry, 1-1 Keyakidai, Sakado 350-0283, Saitama, JapanDivision of Microbiology and Immunology, Department of Oral Biology and Tissue Engineering, Meikai University School of Dentistry, 1-1 Keyakidai, Sakado 350-0283, Saitama, JapanInducible nitric oxide synthase (iNOS) is a signature molecule involved in the classical activation of M1 macrophages and is induced by the Nos2 gene upon stimulation with Th1-cell derived interferon-gamma (IFNγ) and bacterial lipopolysaccharide (LPS). Although the anti-inflammatory cytokine IL-4 is known to inhibit Nos2 gene expression, the molecular mechanism involved in the negative regulation of Nos2 by IL-4 remains to be fully elucidated. In the present study, we investigated the mechanism of IL-4-mediated Nos2 transcriptional repression in the mouse macrophage-like cell line RAW264.7. Signal transducer and activator of transcription 6 (Stat6) knockdown by siRNA abolished the IL-4-mediated inhibition of Nos2 induced by IFNγ/LPS. Transient transfection of a luciferase reporter gene containing the 5′-flanking region of the Nos2 gene demonstrated that an octamer transcription factor (OCT) binding site in the promoter region is required for both positive regulation by IFNγ/LPS and negative regulation by IL-4. Although IL-4 had no inhibitory effect on the DNA-binding activity of constitutively expressed Oct-1, IL-4-induced Nos2-reporter transcriptional repression was partially attenuated by overexpression of the coactivator CREB-binding protein (CBP). These results suggest that a coactivator/cofactor that functionally interacts with Oct-1 is a molecular target for the IL-4-mediated inhibition of Nos2 and that IL-4-activated Stat6 represses Oct-1-dependent transcription by competing with this coactivator/cofactor.http://dx.doi.org/10.1155/2013/369693 |
| spellingShingle | Miki Hiroi Yoshiichi Sakaeda Hana Yamaguchi Yoshihiro Ohmori Anti-Inflammatory Cytokine Interleukin-4 Inhibits Inducible Nitric Oxide Synthase Gene Expression in the Mouse Macrophage Cell Line RAW264.7 through the Repression of Octamer-Dependent Transcription Mediators of Inflammation |
| title | Anti-Inflammatory Cytokine Interleukin-4 Inhibits Inducible Nitric Oxide Synthase Gene Expression in the Mouse Macrophage Cell Line RAW264.7 through the Repression of Octamer-Dependent Transcription |
| title_full | Anti-Inflammatory Cytokine Interleukin-4 Inhibits Inducible Nitric Oxide Synthase Gene Expression in the Mouse Macrophage Cell Line RAW264.7 through the Repression of Octamer-Dependent Transcription |
| title_fullStr | Anti-Inflammatory Cytokine Interleukin-4 Inhibits Inducible Nitric Oxide Synthase Gene Expression in the Mouse Macrophage Cell Line RAW264.7 through the Repression of Octamer-Dependent Transcription |
| title_full_unstemmed | Anti-Inflammatory Cytokine Interleukin-4 Inhibits Inducible Nitric Oxide Synthase Gene Expression in the Mouse Macrophage Cell Line RAW264.7 through the Repression of Octamer-Dependent Transcription |
| title_short | Anti-Inflammatory Cytokine Interleukin-4 Inhibits Inducible Nitric Oxide Synthase Gene Expression in the Mouse Macrophage Cell Line RAW264.7 through the Repression of Octamer-Dependent Transcription |
| title_sort | anti inflammatory cytokine interleukin 4 inhibits inducible nitric oxide synthase gene expression in the mouse macrophage cell line raw264 7 through the repression of octamer dependent transcription |
| url | http://dx.doi.org/10.1155/2013/369693 |
| work_keys_str_mv | AT mikihiroi antiinflammatorycytokineinterleukin4inhibitsinduciblenitricoxidesynthasegeneexpressioninthemousemacrophagecelllineraw2647throughtherepressionofoctamerdependenttranscription AT yoshiichisakaeda antiinflammatorycytokineinterleukin4inhibitsinduciblenitricoxidesynthasegeneexpressioninthemousemacrophagecelllineraw2647throughtherepressionofoctamerdependenttranscription AT hanayamaguchi antiinflammatorycytokineinterleukin4inhibitsinduciblenitricoxidesynthasegeneexpressioninthemousemacrophagecelllineraw2647throughtherepressionofoctamerdependenttranscription AT yoshihiroohmori antiinflammatorycytokineinterleukin4inhibitsinduciblenitricoxidesynthasegeneexpressioninthemousemacrophagecelllineraw2647throughtherepressionofoctamerdependenttranscription |