ABSTRACT
The basic helix loop helix (bHLH) transcription factor DEC2 is associated with the regulation of apoptosis, circadian rhythm and the response to hypoxia. However, the significance of DEC2 in pancreatic cancer remains unknown. Here, we showed for the first time that DEC2 inhibits the progression of human pancreatic cancer. Human pancreatic cancer BxPC-3 cells were treated with or without transforming growth factor-ß (TGF-ß), siRNA against DEC2, or a combination of TGF-ß and DEC2 siRNA or DEC2 overexpression. The cells were analyzed by RT-PCR, real-time PCR, western blotting, immunofluorescent staining and ChIP assay. We also performed immunohistochemical analyses of DEC2 expression in surgically-resected pancreatic cancers. The expression of DEC2 was increased in TGF-ß-treated BxPC-3 cells. In the presence of TGF-ß, DEC2 overexpression decreased the migration and invasion of BxPC-3 cells. Knockdown of DEC2 by siRNA in the presence of TGF-ß significantly increased the expression and nuclear concentration of slug. We also showed that DEC2 siRNA decreased the binding of DEC2 to the E-box of the slug promoter. Immunohistochemically, little DEC2 was detected in pancreatic cancer tissues, whereas significant amounts were detected in the adjacent non-cancerous pancreatic tissues. These results indicate that DEC2 has inhibitory effects against human pancreatic cancer that involve TGF-ß and slug.
Subject(s)
Basic Helix-Loop-Helix Transcription Factors/metabolism , Pancreatic Neoplasms/metabolism , Transforming Growth Factor beta/pharmacology , Basic Helix-Loop-Helix Transcription Factors/genetics , Cell Line, Tumor , Cell Movement/drug effects , Cell Movement/genetics , Disease Progression , Gene Expression Regulation, Neoplastic/drug effects , Gene Silencing , Humans , Pancreatic Neoplasms/genetics , RNA Interference , Snail Family Transcription Factors , Transcription Factors/genetics , Transcription Factors/metabolismABSTRACT
Smads are intracellular signaling mediators. Complexes of Smad2 and Smad3 with Smad4 transmit transforming growth factor-beta (TGF-ß) receptor-induced signaling. Snail plays important roles in mesoderm formation, gastrulation, neural crest development, and epithelial mesenchymal transition. However, it remains unknown whether Smad3 and Snail expression is circadian rhythm-dependent. Here, we showed for the first time that Smad3 and Snail show circadian expression in human gingival fibroblasts (HGF-1) and human mesenchymal stem cells (MSC) after serum shock. They also showed circadian expression in the mouse liver. We confirmed that BMAL1/2, DEC1/2, VEGF, and PER1/2/3 also show circadian expression in both HGF-1 and MSC. The mRNA peaks and phases in circadian expression of these genes differed between HGF-1 and MSC. In a luciferase assay, Smad3 promoter activity was upregulated by CLOCK/BMAL1. These findings suggest that Smad3 and Snail have circadian rhythm in vitro and vivo, and that circadian expression of Smad3 depends on CLOCK/BMAL1.
