Snail is a zinc finger transcription factor encoded by the SNAI1 gene and triggers a cellular process termed epithelial-mesenchymal transition (EMT) upon its increased expression and/or functional act Show more
Snail is a zinc finger transcription factor encoded by the SNAI1 gene and triggers a cellular process termed epithelial-mesenchymal transition (EMT) upon its increased expression and/or functional activation. Snail expression and activity are regulated by various extracellular stimuli, including cytokines and environmental factors. Transforming growth factor-β (TGF-β) is a Snail inducer that functions via Smad3-mediated transcriptional activation. In the present study, we identified a distal enhancer that modulates TGF-β-induced SNAI1 expression. ChIP-seq and Hi-C analyses showed that the enhancer is located 46 kb downstream of the SNAI1 gene; in TGF-β-stimulated cells, it associates with Smad3 and interacts with the SNAI1 proximal promoter. Inhibiting the activity of the enhancer using CRISPRi attenuated TGF-β-induced SNAI1 expression, stress fiber formation, and cell motility enhancement, suggesting that the enhancer mediates TGF-β-induced EMT. The enhancer contains a Smad-binding CAGA motif and an activator protein-1 (AP-1) binding motif that function in transcriptional activation. Ras-responsive element binding protein 1 (RREB1), a transcription factor required for TGF-β-induced Snail expression, regulated the basal activity of the enhancer but not its inducibility by TGF-β. In contrast to the enhancer, the association of Smad3 with the proximal promoter was not evident. These findings suggest that the proximal promoter and the distal enhancer respond to distinct signaling cues, integrate them, and cooperatively function to drive SNAI1 expression. Show less
Activation of the Wnt signaling pathway is frequently observed in hepatocellular carcinoma (HCC), though mutation of three of its components, CTNNB1, AXIN1, and AXIN2, is observed substantially less o Show more
Activation of the Wnt signaling pathway is frequently observed in hepatocellular carcinoma (HCC), though mutation of three of its components, CTNNB1, AXIN1, and AXIN2, is observed substantially less often. We examined the relationship between Wnt signaling and epigenetic alteration of secreted frizzled-related protein (SFRP) genes in HCC. We frequently detected the active form of beta-catenin and accumulation of nuclear beta-catenin in liver cancer cell lines. We detected methylation of SFRP family genes in liver cancer cell lines (SFRP1, 9/12, 75%; SFRP2, 7/12, 58%; SFRP4, 3/12, 25%; SFRP5, 7/12, 58%) and primary HCCs (SFRP1, 9/19, 47%; SFRP2, 12/19, 63%; SFRP5, 8/19, 42%), though methylation of SFRP4 was not found in primary HCCs. SFRP methylation also was detected in hepatitis B or C virus-associated chronic hepatitis (SFRP1, 6/37, 16%; SFRP2, 14/37, 38%; SFRP5, 5/37, 14%) and liver cirrhosis (SFRP1, 10/28, 36%; SFRP2, 9/28, 32%; SFRP5, 3/28, 11%), suggesting that methylation of these genes is an early event in liver carcinogenesis. Ectopic expression of SFRPs downregulated T-cell factor/lymphocyte enhancer factor (TCF/LEF) transcriptional activity in liver cancer cells, while overexpression of a beta-catenin mutant and depletion of SFRP1 using siRNA synergistically upregulated TCF/LEF transcriptional activity. Our results confirm the frequent methylation and silencing of Wnt antagonist genes in HCC, and suggest that their loss of function contributes to activation of Wnt signaling during hepatocarcinogenesis. Show less
Comparative proteomic analysis was used to search for characteristic alterations in the sera of hepatocellular carcinoma (HCC) patients who had undergone curative radiofrequency ablation treatment. Se Show more
Comparative proteomic analysis was used to search for characteristic alterations in the sera of hepatocellular carcinoma (HCC) patients who had undergone curative radiofrequency ablation treatment. Serum samples collected from eight patients before and after treatment were subjected to 2-DE. Eighty-eight protein spots differentially expressed with the treatment were selected by clustering analysis, and the proteins were identified by MS based on MALDI-TOF/TOF analysis and public database searches. The statistical analysis suggested that four proteins decreased after treatment (pro-apolipoprotein, alpha2-HS glycoprotein, apolipoprotein A-IV precursor, and PRO1708/PRO2044, which is the carboxy terminal fragment of albumin) and that seven proteins were increased after treatment, including leucine-rich alpha2-glycoprotein and alpha1-antitrypsin. These data facilitate the identification of differentially expressed proteins that are involved in HCC carcinogenesis and provide candidate biomarkers for the development of diagnostic and therapeutic tools. Show less