Modulation of endogenous Smad expression in normal skin fibroblasts by transforming growth factor-β

Yasuji Mori, Shu Jen Chen, John Varga

Research output: Contribution to journalArticlepeer-review

93 Scopus citations

Abstract

Transforming growth factor-β (TGF-β) stimulation of collagen synthesis plays a fundamental role in physiological tissue repair as well as pathological fibrosis. Members of the SMAD family of intracellular proteins are phosphorylated by TGF-β receptors and convey signals to specific TGF-β- inducible genes. Ligand binding initiates signaling through the SMAD pathway, but it is unknown how signaling is terminated. The expression and regulation of Smads have been characterized mostly in transformed cells using transient expression systems. In this study, we investigated the physiological regulation of endogenous Smads by TGF-β in nontransformed normal skin fibroblasts in vitro. Treatment with TGF-β resulted in time and dose- dependent translocation of SMAD3 and SMAD4 from the cytoplasm to the nucleus. The levels of SMAD3 and Smad3 mRNA were profoundly down-regulated by TGF-β1 or TGF-β3 in a time-dependent manner, whereas expression of antagonistic Smad7 was rapidly and transiently induced. The stability of Smad3 mRNA transcripts was unaffected by TGF-β. Cycloheximide prevented the inhibition of Smad3, but not the induction of Smad7, mRNA expression by TGF- β1, identifying Smad7 as an immediate-early gene target of TGF-β in fibroblasts. In Smad4-deficient breast cancer cells, TGF-β failed to modulate Smad expression, suggesting that SMADs mediate their own regulation in response to ligand. These results demonstrate that TGF-β not only triggers functional activation of the SMAD signaling cascade in primary skin fibroblasts, but also simultaneously exerts potent effects on endogenous SMAD expression and intracellular trafficking. Taken together with recent reports implicating ubiquitination in SMAD turnover, these findings indicate the existence of multiple levels of control for modulating SMAD-mediated TGF-β signaling in fibroblasts.

Original languageEnglish (US)
Pages (from-to)374-383
Number of pages10
JournalExperimental Cell Research
Volume258
Issue number2
DOIs
StatePublished - Aug 1 2000

Keywords

  • Skin fibroblasts
  • Smad signaling
  • Smad3
  • Smad7
  • TGF-β

ASJC Scopus subject areas

  • Cell Biology

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