Abstract 4774: JC virus T-antigen-dependent activation of Wnt target genes and cell cycle progression in colon cancer.

Abstract

Abstract Background: The Wnt signaling pathway is aberrantly activated in many different types of cancer including colon cancer. Transcription of target genes in the Wnt pathway promotes strong proliferation and differentiation signals in the cell. This transcription is mediated by β-catenin, the central signaling protein in the Wnt pathway. β-catenin targets include potent transcription factors and cell cycle regulatory proteins such as c-myc and cyclin D1. We have previously shown that a strong regulator of β-catenin-binding is the polyomavirus JC virus protein large T-antigen (TAg). According to sero-epidemiological studies, JCV establishes a latent infection in 70-90% of the general population by adulthood and TAg has been associated with numerous types of cancer in the CNS and GI tract. We and others have previously shown that TAg is expressed in colon cancer samples and has a significant association with β-catenin in the nuclei of colon cancer cells. Our hypothesis is that the viral protein T-antigen (TAg) interacts with β-catenin in colonic epithelial cells to promote transcription of β-catenin target genes particularly c-myc and cyclin D1, which affect cell cycle progression and lead to unchecked cellular growth. Materials and Methods: To test this hypothesis we transfected colon cancer cell lines with a plasmid containing the TAg gene and measured β-catenin target gene transcription. We used the TOPflash reporter construct to determine general β-catenin / TCF4-mediated promoter activity and reporter constructs for the β-catenin target genes c-myc and cyclin D1. We performed RT-qPCR and Western blot analysis to determine mRNA and protein levels of c-myc and cyclin D1 in TAg- and empty vector-transfected cells. We also utilized β-catenin siRNA or the dominant negative TCF4 plasmid to inhibit Wnt signaling in these groups and measured the same endpoints. Results: TCF-dependent promoter activity with a TOPflash reporter plasmid was increased in cells transfected with the TAg gene. This increased activity resulted in elevated levels of β-catenin target genes c-myc and cyclin D1. Moreover, inhibition of β-catenin, either by β-catenin siRNA or co-transfection with DN-TCF4 ablated this response. Finally, we found that transfection with TAg altered cell cycle distribution in these colon cancer cells with an increased proportion of cells in the G2/M phase. Conclusions: We found that the TAg protein activates the Wnt signaling pathway in our cellular model of colon cancer. This activation was determined to be β-catenin-dependent and mediated by TCF4 binding to promoter regions. Moreover, because Wnt pathway activation in these cells leads to cell cycle progression we suggest an oncogenic role for TAg in colon cancer. Considering the large proportion of the population seropositive for JC virus, it is imperative to elucidate its role in the oncogenesis of colon cancer. Citation Format: Michael J. Ripple, Amanda Struckhoff, Robin McGoey, Luis Del Valle. JC virus T-antigen-dependent activation of Wnt target genes and cell cycle progression in colon cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4774. doi:10.1158/1538-7445.AM2013-4774