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Ras controls tumor necrosis factor receptor-associated factor (TRAF)6-dependent induction of nuclear factor-kappa b.
C J Caunt1, E Kiss-Toth, F Carlotti
1Functional Genomics, Division of Molecular and Genetic Medicine, University of Sheffield, S10 2JF, United Kingdom.
Abstract:
In the present study, we show that Ras activity differentially controls interleukin (IL)-1 induced transcription factor activation by selective regulation of responses mediated by receptor complex components. Initial experiments revealed that stimulation with IL-1 caused a rapid, matrix-dependent activation of Ras. The effect was transient, peaking at 5 min and returning to base levels after 30 min. Activation correlated with pronounced changes in cell shape in EGFPH-Ras transfected cells. Transfection with the dominant negative mutant, Ras(Asn-17), inhibited IL-1 induced activation of the IL-8 promoter as well as of NF-kappa B and AP-1 synthetic promoters in transient transfection assays. Furthermore, overexpression of the IL-1 signaling proteins TRAF6 or MyD88 gave characteristic activation of IL-8, which was accentuated in the presence of IL-1. Co-transfection with Ras(Asn-17) gave a dose-dependent inhibition of TRAF6-induced responses in the presence and absence of IL-1, but had no effect on MyD88 mediated activity. Similarly, induction of NF-kappa B was abolished by Ras(Asn-17) only in TRAF6-transfected cells. In contrast, inhibiting Ras activity limited AP-1-mediated responses through both receptor complex proteins. Constitutively active Ras(Val-12) increased the TRAF6 induced activity of the NF-kappa B pathway similar to the effect induced by IL-1, while the Ras(Val-12) induced activity was not inhibited by co-transfection with a dominant negative TRAF6. Our data show that activation of the Ras GTPase is an early, matrix-dependent response in IL-1 signaling which participates in structural regulation of IL-1-induced genes. In addition, they show that the Ras induced effect selectively regulates TRAF6-mediated activation of the NF-kappa B pathway, suggesting that Ras GTPase represents a convergence point in structural and cytokine responses, with distinct effects on a subset of downstream signaling events.
Insights
Ras GTPase activation is an early, matrix-dependent response in interleukin-1 (IL-1) signaling, selectively regulating transcription factor activation and downstream signaling pathways like NF-kappa B and AP-1.
Area of Science:
- Molecular Biology
- Cell Signaling
- Immunology
Background:
- Interleukin-1 (IL-1) is a key cytokine in inflammatory and immune responses.
- Ras GTPase signaling plays a crucial role in various cellular processes, including proliferation and differentiation.
Purpose of the Study:
- To investigate the role of Ras activity in IL-1 induced transcription factor activation.
- To determine how Ras selectively regulates signaling pathways downstream of IL-1 receptor complex components.
Main Methods:
- Utilized transient transfection assays with dominant-negative and constitutively active Ras mutants.
- Measured IL-8 promoter activity and activation of NF-kappa B and AP-1 synthetic promoters.
- Investigated the interplay between Ras, TRAF6, and MyD88 in IL-1 signaling.
Main Results:
- IL-1 stimulation rapidly activated Ras in a matrix-dependent manner.
- Dominant-negative Ras inhibited IL-1 induced activation of IL-8, NF-kappa B, and AP-1 promoters.
- Ras selectively regulated TRAF6-mediated NF-kappa B activation but not MyD88-mediated activity.
- Ras inhibition affected AP-1 responses mediated by both TRAF6 and MyD88.
Conclusions:
- Ras GTPase activation is an early, matrix-dependent event in IL-1 signaling.
- Ras selectively modulates TRAF6-mediated NF-kappa B activation, indicating a convergence point in signaling.
- Ras plays a distinct role in regulating specific downstream signaling events in response to IL-1.