ABSTRACT
BACKGROUND: T-cell factor (TCF) proteins represent key transcription factors that activate Wnt/ß-catenin signalling. We have reported that a pair of TCF-4 isoforms (TCF-4C and TCF-4D) exhibit differential TCF transcriptional activity in hepatocellular carcinoma (HCC) cells, although their structure differs by only the presence (TCF-4D) or absence (TCF-4C) of exon 4. AIM: To demonstrate a regulatory role of exon 4 in HCC development. METHODS: TCF-4C and TCF-4D expression profiles were examined in 27 pairs of human HCC and adjacent liver tissues. The functional role of the TCF-4 isoforms was evaluated in OUMS-29 (an immortalized hepatocyte-derived) and HAK-1A (a well-differentiated HCC) cell lines using stable clones overexpressing the TCF-4 isoforms. RESULTS: TCF-4C was significantly upregulated in HCC tissues compared with corresponding peritumour and normal liver tissues; in contrast, there was no difference in TCF-4D expression. TCF-4C clones derived from both cell lines exhibited increased TCF activity, Wnt-responsive target genes, cell proliferation, cell cycle progression and resistance to chemotherapeutic drugs compared with TCF-4D clones. Capability of cell migration and colony formation was significantly higher in TCF-4C than TCF-4D clones. In a nude mice xenograft model, the HAK-1A-derived TCF-4C clone rapidly developed tumours compared with the TCF-4D clone. TCF-4C clone-derived tumours exhibited upregulation of Wnt-responsive target genes compared with the slow developing and small TCF-4D-derived tumours. CONCLUSION: These results demonstrate that the TCF-4C isoform lacking exon 4 is associated with a malignant phenotype compared with the exon 4-harbouring TCF-4D isoform, indicating that exon 4 of TCF-4 plays a prominent role in HCC development.
Subject(s)
Carcinoma, Hepatocellular/genetics , Liver Neoplasms/genetics , Transcription Factor 7-Like 2 Protein/genetics , Transcription Factor 7-Like 2 Protein/metabolism , Animals , Blotting, Western , Cell Cycle/genetics , Cell Line, Tumor , Cell Movement/genetics , Cell Proliferation , Colony-Forming Units Assay , Exons/genetics , Fluorescent Antibody Technique , Gene Expression Profiling , Humans , Immunoprecipitation , Mice , Mice, Nude , Protein Isoforms/genetics , Protein Isoforms/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Statistics, NonparametricABSTRACT
Hepatocellular carcinoma (HCC) is the third leading cause of cancer mortality worldwide. However, there is little known about targeted therapeutics for the treatment of this devastating tumor. Among the growth factor signaling cascades deregulated in HCC, evidences suggest that the WNT/Frizzled-mediated signaling pathway plays a key role in the hepatic carcinogenesis. Aberrant activation of the signaling in HCC is mostly due to deregulated expression of the Wnt/ß-catenin signaling components. This leads to the activation of the ß-catenin/TCF dependent target genes, which controls cell proliferation, cell cycle, apoptosis or motility. It has been shown that disruption of the Wnt/ß-catenin signaling cascade displayed anti-cancer properties in HCC. Currently, no therapeutic molecules targeting the WNT pathway are available or under clinical evaluation for the treatment of HCC. This review will discuss the identified potential molecular targets related to the canonical WNT signaling pathway and their potential therapeutic usefulness.
Subject(s)
Carcinoma, Hepatocellular/drug therapy , Carcinoma, Hepatocellular/metabolism , Liver Neoplasms/drug therapy , Liver Neoplasms/metabolism , Molecular Targeted Therapy , Wnt Proteins/metabolism , beta Catenin/metabolism , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/metabolism , Antibodies, Monoclonal , Carcinoma, Hepatocellular/genetics , Dishevelled Proteins , Frizzled Receptors/antagonists & inhibitors , Frizzled Receptors/metabolism , Gene Knockdown Techniques , Humans , Liver Neoplasms/genetics , Phosphoproteins/genetics , Phosphoproteins/metabolism , Signal Transduction , Wnt Proteins/antagonists & inhibitors , Wnt Proteins/immunology , beta Catenin/geneticsABSTRACT
The Wnt/ß-catenin signaling pathway is frequently activated in hepatocellular carcinoma (HCC). Downstream signaling events involving the Wnt/ß-catenin cascade occur through T-cell factor (TCF) proteins. The human TCF-4 gene is composed of 17 exons with multiple alternative splicing sites. However, the role of different TCF-4 isoforms in the pathogenesis of HCC is unknown. The purpose of this study was to identify and characterize TCF-4 isoforms in HCC. We identified 14 novel TCF-4 isoforms from four HCC cell lines. Functional analysis following transfection and expression in HCC cells revealed distinct effects on the phenotype. The TCF-4J isoform expression produced striking features of malignant transformation characterized by high cell proliferation rate, migration and colony formation even though its transcriptional activity was low. In contrast, the TCF-4K isoform displayed low TCF transcriptional activity; cell proliferation rate and colony formation were reduced as well. Interestingly, TCF-4J and TCF-4K differed by only five amino acids (the SxxSS motif). Thus, these studies suggest that conserved splicing motifs may have a major influence on the transcriptional activity and functional properties of TCF-4 isoforms and alter the characteristics of the malignant phenotype.
