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Rapid activation of proteins that interact with the interferon gamma activation site in response to multiple
1Ligand Pharmaceuticals, San Diego, CA 92121.
Abstract:
Many cytokines and growth factors trigger rapid changes in gene expression upon binding to their receptors. In many cases, the mechanism by which these changes are affected is unknown. In this report, we show that interleukin-2 (IL-2), IL-3, IL-4, IL-6, leukemia inhibitory factor (LIF), erythropoietin (Epo), and granulocyte-macrophage colony-stimulating factor (GM-CSF) treatment of cells causes rapid activation of DNA-binding activities that recognize a DNA sequence element previously implicated in regulation of gene expression by interferon gamma (IFN gamma). The IL-4-, IL-6-, and GM-CSF-induced complexes can be distinguished from the recently characterized IFN gamma-activated protein p91 on the basis of mobility in polyacrylamide gels, sequence preferences, and lack of reactivity with an anti-p91 antiserum. The IL-4- and GM-CSF-induced complexes react with antiphosphotyrosine antibodies, demonstrating the presence of phosphotyrosine-containing proteins in these DNA-binding complexes. Transcriptional activation of a reporter gene linked to a synthetic IFN gamma-responsive promoter is observed in response to IFN gamma, IL-6, and LIF. These data suggest a pathway by which cytokines induce rapid changes in gene expression.
Insights
Cytokines like interleukin-6 (IL-6) and leukemia inhibitory factor (LIF) rapidly activate DNA-binding proteins, influencing gene expression. This study identifies novel cytokine-activated pathways distinct from interferon gamma (IFN-γ) signaling.
Area of Science:
- Molecular Biology
- Cell Signaling
- Immunology
Background:
- Cytokines and growth factors rapidly alter gene expression upon receptor binding, but the underlying mechanisms are often unclear.
- Interferon gamma (IFN-γ) activates specific DNA-binding proteins involved in gene regulation.
Purpose of the Study:
- To investigate the mechanisms by which various cytokines induce rapid changes in gene expression.
- To identify and characterize novel cytokine-activated DNA-binding activities and their role in gene regulation.
Main Methods:
- Treatment of cells with specific cytokines including interleukin-2 (IL-2), IL-3, IL-4, IL-6, leukemia inhibitory factor (LIF), erythropoietin (Epo), and granulocyte-macrophage colony-stimulating factor (GM-CSF).
- Electrophoretic mobility shift assays (EMSAs) to detect DNA-binding activities.
- Characterization of induced complexes using gel mobility, sequence preferences, and antibody reactivity (anti-p91, antiphosphotyrosine).
- Reporter gene assays to assess transcriptional activation of IFN-γ-responsive promoters.
Main Results:
- Multiple cytokines (IL-2, IL-3, IL-4, IL-6, LIF, Epo, GM-CSF) rapidly activate DNA-binding activities recognizing an IFN-γ-responsive element.
- IL-4, IL-6, and GM-CSF induced complexes differ from the known IFN-γ-activated protein p91.
- IL-4 and GM-CSF induced complexes contain phosphotyrosine-containing proteins.
- IFN-γ, IL-6, and LIF induced transcriptional activation of a reporter gene linked to a synthetic IFN-γ-responsive promoter.
Conclusions:
- Cytokines utilize distinct signaling pathways to rapidly modulate gene expression.
- Novel phosphotyrosine-containing DNA-binding proteins are activated by certain cytokines, contributing to gene regulation.
- These findings reveal a conserved mechanism for rapid gene expression changes mediated by various cytokines.