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IL-2 and long-term T cell activation induce physical and functional interaction between STAT5 and ETS transcription
P Rameil1, P Lécine, J Ghysdael
1INSERM U119, Marseille, France.
Abstract:
Activation of Stat5 by many cytokines implies that it cannot alone insure the specificity of the regulation of its target genes. We have evidenced a physical and functional interaction between members of two unrelated transcription factor families, Ets-1, Ets-2 and Stat5, which could contribute to the proliferative response to interleukin 2. Competition with GAS- and EBS-specific oligonucleotides and immunoassays with a set of anti-Stat and anti-Ets families revealed that the IL-2-induced Stat5-Ets complex recognizes several GAS motifs identified as target sites for activated Stat5 dimers. Coimmunoprecipitation experiments evidenced that a Stat5/Ets-1/2 complex is formed in vivo in absence of DNA. GST-pull down experiments demonstrated that the C-terminal domain of Ets-1 is sufficient for this interaction in vitro. Cotransfection experiments in Kit225 T cells resulted in cooperative transcriptional activity between both transcription factors in response to a combination of IL-2, PMA and ionomycin. A Stat5-Ets protein complex was the major inducible DNA-binding complex bound to the human IL-2rE GASd/EBSd motif in long-term proliferating normal human T cells activated by CD2 and CD28. These results suggest that the inducible Stat5-Ets protein interaction plays a role in the regulation of gene expression in response to IL-2 in human T lymphocytes.
Insights
Transcription factors Ets and Stat5 interact physically and functionally to regulate gene expression in response to interleukin-2 (IL-2). This Stat5-Ets complex specifically binds to DNA, contributing to T cell proliferation.
Area of Science:
- Molecular Biology
- Immunology
- Cell Biology
Background:
- Cytokine signaling often lacks specificity, necessitating co-regulatory mechanisms for precise gene expression.
- Stat5 activation by cytokines is crucial but insufficient for specific target gene regulation.
- Interleukin-2 (IL-2) signaling involves T cell proliferation, a process potentially mediated by combinatorial transcription factor activity.
Purpose of the Study:
- To investigate the physical and functional interaction between Ets transcription factors (Ets-1, Ets-2) and Stat5.
- To determine the role of this interaction in the context of IL-2-mediated T cell responses.
- To elucidate the molecular mechanisms underlying Stat5-Ets complex formation and DNA binding.
Main Methods:
- Oligonucleotide competition assays using GAS and EBS motifs.
- Immunoassays with anti-Stat and anti-Ets antibodies.
- Coimmunoprecipitation to detect in vivo complex formation.
- GST-pull down assays for in vitro interaction studies.
- Cotransfection experiments in T cells to assess transcriptional activity.
Main Results:
- A physical and functional interaction between Ets-1/2 and Stat5 was identified.
- The Stat5-Ets complex binds to GAS motifs, previously known Stat5 target sites.
- Complex formation occurs in vivo and is mediated by the C-terminal domain of Ets-1.
- Cooperative transcriptional activity was observed in response to IL-2 and other stimuli.
- A Stat5-Ets complex was the predominant inducible DNA-binding factor on the IL-2 receptor regulatory element in activated human T cells.
Conclusions:
- The inducible interaction between Stat5 and Ets transcription factors plays a significant role in regulating gene expression.
- This interaction contributes to the specific response of human T lymphocytes to IL-2.
- The findings reveal a novel mechanism for achieving specificity in cytokine-driven gene regulation.