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c-myc is a downstream target of the Smad pathway
Ken Yagi1, Masao Furuhashi, Hiromasa Aoki
1Department of Biochemistry, The Cancer Institute of the Japanese Foundation for Cancer Research and Research for the Future Program, Japan Society for the Promotion of Science, 1-37-1 Kami-ikebukuro, Toshima-ku, Tokyo 170-8455, Japan.
Abstract:
c-Myc is one of the most potent regulators of cell cycle progression in higher eukaryotes. Down-regulation of c-Myc is a critical event for growth inhibition induced by transforming growth factor-beta (TGF-beta) and is frequently impaired in cancer cells. We determined a Smad-responsive element in the c-myc promoter. This element is a complex of the TGF-beta1 inhibitory element (TIE) originally identified in the transin/stromelysin promoter and an E2F site responsible for transcriptional activation of the c-myc promoter. Smad3 and E2F-4 directly bound to the element (TIE/E2F), and substitution of two nucleotides in TIE/E2F impaired binding of both Smad3 and E2F-4 as well as serum-induced activation and TGF-beta-induced suppression of the c-myc promoter activity. Smads bound TIE/E2F within 1 h after stimulation with TGF-beta, before the suppression of c-myc transcription, whereas binding of p130 to TIE/E2F became augmented later than 12 h. TGF-beta signaling did not compete with E2F-4 for binding to TIE/E2F, but reduced p300 co-immunoprecipitating with E2F-4. Therefore, TGF-beta signaling may suppress c-myc promoter activity by dissociating p300 from E2F-4.
Insights
Transforming growth factor-beta (TGF-β) suppresses c-Myc, a key cell cycle regulator, by altering its promoter activity. This involves Smad proteins and E2F-4, impacting cancer cell growth.
Area of Science:
- Molecular Biology
- Cell Biology
- Cancer Research
Background:
- c-Myc is a crucial regulator of cell cycle progression.
- TGF-β-induced growth inhibition requires c-Myc down-regulation, often defective in cancers.
Purpose of the Study:
- To elucidate the mechanism by which TGF-β signaling suppresses c-Myc transcription.
- To identify the specific DNA elements and protein interactions involved.
Main Methods:
- Analysis of the c-myc promoter to identify a novel Smad-responsive element (TIE/E2F).
- Electrophoretic mobility shift assays (EMSAs) and chromatin immunoprecipitation (ChIP) to assess protein binding.
- Reporter gene assays to measure promoter activity.
Main Results:
- A complex TIE/E2F element in the c-myc promoter binds both Smad3 and E2F-4.
- Mutations in TIE/E2F disrupt Smad3 and E2F-4 binding and affect promoter activity.
- TGF-β signaling leads to Smad binding and subsequent dissociation of p300 from E2F-4, preceding c-Myc suppression.
Conclusions:
- TGF-β signaling suppresses c-Myc transcription through a Smad-dependent mechanism at the TIE/E2F element.
- The suppression involves the dissociation of the co-activator p300 from E2F-4, impacting cell cycle control.