The Murine Gastrin Promoter Is Synergistically Activated by Transforming Growth Factor-β/Smad and Wnt Signaling Pathways

Abstract

The transforming growth factor-␤ (TGF-␤) and Wnt/ wingless pathways play critical roles in the specification of cell fate during development and also contribute to cancer formation and progression. Whereas Wnt signaling is clearly pro-oncogenic, TGF-␤ signaling is celland context-dependent, manifesting both inhibitory and proliferative effects. The growth factor, gastrin, has previously been shown to be a downstream target of the Wnt pathway and a promoter of gastrointestinal cancer. In this study, we show that the mouse gastrin promoter is regulated synergistically by TGF-␤/Smads and ␤-catenin/T-cell factor (TCF). Co-transfection of Smad3/Smad4 and ␤-catenin expression constructs synergistically activated mouse gastrin promoter activity 30 -60-fold in AGS cells with minimal effect seen with either construct alone. This activation was further potentiated by TGF-␤1 treatment. Mutating either the TCF binding site or the Smad-binding element (SBE) diminished the activation of gastrin expression by Smad3/Smad4 and ␤-catenin and led to a loss of gastrin promoter responsiveness to TGF-␤1 treatment. Wnt and TGF-␤ regulated endogenous gastrin mRNA levels in AGS cells in a similar fashion, as revealed by small interference RNA studies or overexpression of Smads and TCF4/␤-catenin. Electrophoretic mobility shift assays and DNA affinity precipitation assays showed that the putative SBE and T-cell factor (TCF) sites were able to bind a complex containing Smads and ␤-catenin/TCF4. In addition, the synergy between Smads and ␤-catenin/TCF4 was dependent on CREB-binding protein (CBP)/P300, as demonstrated by overexpression of CBP or E1A. Moreover, by using a heterogeneous promoter reporter system, we showed that this complex containing Smads/TCF4/␤-catenin complex was able to up-regulate transcription at isolated SBE or TCF sites. Thus, the Wnt signaling pathway is able to activate some target genes through its actions as a co-activator at non-TCF sites and has the potential to profoundly alter transcriptional responses to TGF-␤ signaling.

Annexin II binds progastrin and gastrin-like peptides, and mediates growth factor effects of autocrine and exogenous gastrins on colon cancer and intestinal epithelial cells

“…Upregulation of ANX II was also discovered in gastric epithelial cells infected with Helicobacter pylori (Das et al, 2005). At the same time, H. pylori infection of gastric mucosa, and growth factors such as EGF and TGF-b1 have been reported to increase the expression of gastrin gene, directly or indirectly (Lei et al, 2004;Rengifo-Cam and Singh, 2004). These findings suggest the intriguing possibility that the direct (EGF, TGF-b1, oncogenic K-ras) and indirect (H. pylori) growth effects of these agents may be magnified by an upregulation of both the gastrin gene (PG) and its binding 'receptor' protein, ANX II.…”

“…Recent studies further highlight an important role of colon cancerassociated oncogenic pathways (mutant Ras and Wnt/ b-catenin) in upregulating the gastrin gene expression in CRCs (Lei et al, 2004;Chakladar et al, 2005).…”

Smad4 cooperates with lymphoid enhancer-binding factor 1/T cell-specific factor to increase c- myc expression in the absence of TGF-β signaling

“…The first evidence that Smad and LEF/TCF functionally interact came from studies of the Xtwn promoter in Xenopus, which contains both SBE and TBE elements (13,14). In mouse, the Smad-LEF/TCF complex is implicated in the gene expression of Emx2, MSX2, and gastrin and also in the process of palatal development (16,17,39,40). The general model proposed on the basis of these studies is that in the presence of either Wnt or BMP/TGF signaling, the target genes are activated moderately, but when both signals are activated, the target genes are synergistically activated by the formation of R-Smad-Smad4-LEF/ TCF-␤-catenin complex.…”

Transcription Factor NF-Y Is a Functional Regulator of the Transcription of Core Clock Gene Bmal1

“…Preparation of Nuclear Extract and DNA Affinity Precipitation Assays (DAPA)-Nuclear extracts from 24 h post-serumshocked NIH 3T3 cells were prepared as described previously (19). DAPA have been described previously (19).…”