Identification of the gene transcription and apoptosis mediated by TGF‐β‐Smad2/3‐Smad4 signaling

Abstract

Transforming growth factor-beta (TGF-beta) signaling is known to depend on the formation of Smad2/3-Smad4 transcription regulatory complexes. However, the signaling functions of Smad2/3-Smad4 during TGF-beta-induced responses are obscure as TGF-beta also initiates a number of other signaling pathways. In this study, we systematically assessed the contribution of TGF-beta-Smad2/3-Smad4 signaling to both target gene transcription and apoptosis. Individual Smads were selectively knocked down in Hep3B cells by stable RNA interference (RNAi). We identified TGF-beta-responsive genes using genome-wide oligonucleotide microarrays and confirmed their dependency on Smad2, Smad3, or Smad4 by the combination of RNAi and microarray assay. The major finding from our microarray analysis was that of the 2,039 target genes seen to be regulated via TGF-beta induction, 190 were differentially transcriptionally controlled by Smad2-Smad4 and Smad3-Smad4 signaling and the latter control mechanism appeared to be functionally more important. We also found indirect evidence of competition between Smad2 and Smad3 for their activation when controlling the transcription of target genes. Functional analysis revealed that Smad3 and Smad4 were the predominant mediators of TGF-beta-induced apoptosis in Hep3B cells. We provide evidence that up-regulation of Bcl-2-interacting mediator of cell death (Bim), under the transcriptional control of Smad3-Smad4 signaling, is crucial to TGF-beta-induced apoptosis in Hep3B cells.

Nephronectin Expression Is Up-Regulated by BMP-2

“…[18][19][20] As for the expression of Npnt, Tsukasaki et al reported its suppression by Smad2, 21) while the present findings demonstrated its activation by Smad4. Together, these observations led us to speculate that the Npnt expression is also regulated by a transcription co-activator protein such as CBP/p300, and mediated by the competition between Smad2 and Smad4, though additional experiments are needed to more fully reveal the mechanism details.…”

Smad3 Is a Key Nonredundant Mediator of Transforming Growth Factor β Signaling in Nme Mouse Mammary Epithelial Cells

“…In accordance with our results obtained in the Nme cells used here, the data collected from these other cell types point to Smad3 as an essential mediator of responses to TGFβ. Three reports from studies in human epithelial cell lines, including keratinocytes or hepatocytes, showed that Smad3 also has a prominent role in TGFβ-mediated cell cycle arrest and apoptosis (25)(26)(27).…”

p21-activated Kinase 2 (PAK2) Inhibits TGF-β Signaling in Madin-Darby Canine Kidney (MDCK) Epithelial Cells by Interfering with the Receptor-Smad Interaction

“…4E). Finally, we tested the expression of several TGF-␤ target genes, including Smad7, c-Myc, and Bim (32)(33)(34)(35)(36). As shown in Fig.…”