Inhibition of Tcf-4 Induces Apoptosis and Enhances Chemosensitivity of Colon Cancer Cells

Jiang Xie,Cong Zhao,Feng Xiong,Hong Wang,Ting-Yu Li,Jie Chen,Xiao-Lei Wang,De-Bing Xiang,Wael El-Rifai

doi:10.1371/journal.pone.0045617

Abstract

Aberrant activation of β-catenin/Tcf-4 signaling has been implicated in human carcinogenesis, including colorectal cancer. In this study, we compared the effects of Tcf-4 knockdown with β-catenin knockdown on cell proliferation, apoptosis, and chemosensitivity in SW480 and HCT116 colon cancer cells using adenoviral vector-mediated short hairpin RNA (shRNA). Our results show that, compared to β-catenin knockdown, Tcf-4 knockdown more effectively inhibited colony formation, induced apoptosis, and increased 5-FU and oxaliplatin-mediated cytotoxicity in colon cancer cells. We further investigated the mechanisms involved in the different efficacies observed with β-catenin and Tcf-4 knockdown in colon cancer cells. FOXO4 is a member of the subfamily of mammalian FOXO forkhead transcription factors and plays a major role in controlling cellular proliferation, apoptosis, and DNA repair. Our data showed that the protein level of FOXO4 did not change after treatment with both β-catenin and Tcf-4 shRNA. However, β-catenin shRNA was found to increase the accumulation of phosphorylated FOXO4 S193 and decrease the expression of FOXO target genes p27Kip1 and MnSOD, whereas Tcf-4 shRNA showed the opposite effect. Therefore, compared to β-catenin knockdown, Tcf-4 knockdown shows better efficacy for inhibiting proliferation and inducing apoptosis of colorectal cancer cells, which may be related to increased FOXO4 transcriptional activity. These results suggest that Tcf-4 is an attractive potential therapeutic target for colorectal cancer therapy.

Highlights

The canonical Wnt signaling pathway plays a central role in numerous cellular processes, from embryonic development to adult tissue homeostasis [1]
In the absence of Wnt signaling, cytoplasmic b-catenin is normally maintained at low levels through continuous ubiquitinproteasome-mediated degradation of b-catenin, which is regulated by a destruction complex composed of adenomatous polyposis coli (APC), glycogen synthase kinase-3b (GSK-3b), Axin/Conductin, Casein Kinase 1a (CK1a), and other proteins that mediate these biochemical reactions
We show that compared to b-catenin knockdown, Tcf-4 knockdown significantly inhibits cell proliferation, induces cell apoptosis, and enhances the chemosensitivity of colon cancer cells through the upregulation of FOXO4 transcriptional activity

Summary

Introduction

The canonical Wnt signaling pathway plays a central role in numerous cellular processes, from embryonic development to adult tissue homeostasis [1]. The activity of this signaling pathway is determined by the amount of b-catenin present in the cytoplasm. Binding of Wnt proteins to the cell surface receptor complex Fz/LRP facilitates the phosphorylation of the cytoplasmic tail of LDL receptor-related protein (LRP) cytoplasmic tail by GSK-3b [2] This triggers the interaction of the Fz/ LRP complex with Dishevelled (Dsh) and Axin, which leads to the inactivation of the destruction complex, resulting in the accumulation of non-phosphorylated b-catenin in the cytoplasm. The accumulated b-catenin translocates into the nucleus and binds to Tcf/Lef transcription factors to regulate downstream target genes, such as c-Myc and Cyclin D1 [3,4,5]

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