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Zinc Finger Transcription Factors as Molecular Targets for Nitric Oxide-mediated Immunosuppression: Inhibition of IL-2 Gene Expression in Murine Lymphocytes

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Abstract

Background

Nitric oxide (NO) has frequently been shown to display immunosuppressive activities. We describe here a molecular mechanism contributing to this effect.

Materials and Methods

Murine T cell lymphoma EL4-6.1 cells were activated with the physiological stimulus interleukin (IL)-1β to express IL-2 mRNA in the presence or absence of subtoxic concentrations of the physiological spontaneous NO donor S-nitrosocysteine (SNOC). Subsequently, semiquantitative RT-PCR and gel shift assays with nuclear extracts were performed to analyze the effects of NO on IL-2 mRNA expression and on the activity of the dominant regulating transcription factors Spl, EGR-1, and NFATc.

Results

NO inhibits IL-1β-induced IL-2 mRNA expression in EL4-6.1 cells. The suppressive activity of NO was concentration dependent and found to be completely reversible. Importantly, NO at the concentrations used induced neither apoptosis nor necrosis. Dominant regulation of IL-2 gene expression is known to reside in the zinc finger transcription factors Sp1 or EGR-1 and in the non-zinc finger protein NFAT. NO abrogates the DNA binding activities of recombinant Sp1 and EGR-1. More importantly, gel shift assays also showed a lack of DNA binding of native Sp1 derived from NO-treated nuclear extracts and that from NO-treated viable lymphocytes. This effect is selective, as the DNA binding activity of recombinant NFATc was not affected by NO.

Conclusion

Inactivation of zinc finger transcription factors by NO appears to be a molecular mechanism in the immunosuppressive activity of NO in mammals, thus contributing to NO-mediated inhibition of IL-2 gene expression after physiological stimuli. The exact understanding of the molecular mechanism leading to NO-mediated, fully reversible suppression of immune reactions may lead to use of this naturally occuring tool as an aid in inflammatory diseases.

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Acknowledgments

We thank Dr. V. Burckhardt (Diabetes Research Center at the Heinrich-Heine University of Düsseldorf) for supplying EL4-6.1 mouse lymphoma cells. Dr. C. Esser (Medical Institute for Environmental Hygiene, Düsseldorf) for supplying mouse IL-2 primer, O. Grapenthin (Institute of Pharmaceutical Chemistry, Heinrich-Heine University of Düsseldorf) for providing PAPA/NO, and M. Turken for the photographic work. This work was supported by grants from the Deutsche Forschungsgemeinschaft (Kr 1443/3–1 and 3–2 to K.D.K).

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Authors and Affiliations

  1. Research Group Immunobiology 14.80, MED-Heinrich-Heine University of Düsseldorf, P.O. Box 10 10 07, D-40001, Düsseldorf, Germany

    Denise Berendji, Victoria Kolb-Bachofen &  Klaus-D. Kröncke

  2. Research Group of Biomolecular Medicine, Bernhard Nocht Institute for Tropical Medicine, Hamburg, Germany

    Peter F. Zipfel & Christine Skerka

  3. The Institute of Physiological Chemistry I, MED-Heinrich-Heine University of Düsseldorf, Düsseldorf, Germany

    Carsten Carlberg

Authors
  1. Denise Berendji
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  2. Victoria Kolb-Bachofen
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  3. Peter F. Zipfel
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  4. Christine Skerka
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  5. Carsten Carlberg
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  6. Klaus-D. Kröncke
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Correspondence to Klaus-D. Kröncke.

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Berendji, D., Kolb-Bachofen, V., Zipfel, P.F. et al. Zinc Finger Transcription Factors as Molecular Targets for Nitric Oxide-mediated Immunosuppression: Inhibition of IL-2 Gene Expression in Murine Lymphocytes. Mol Med 5, 721–730 (1999). https://doi.org/10.1007/BF03402096

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