SPDEF Induces Quiescence of Colorectal Cancer Cells by Changing the Transcriptional Targets of β-catenin

BACKGROUND & AIMS: The canonical Wnt signaling pathway activates the transcriptional activity of beta-catenin. This pathway is often activated in colorectal cancer cells, but strategies to block it in tumors have not been effective. The SAM pointed domain containing ETS transcription factor (SPDEF) suppresses formation of colon tumors by unclear mechanisms. We investigated these mechanisms and the effects of SPDEF on beta-catenin activity in mouse models of colorectal cancer (CRC), CRC cell lines, and mouse and human normal and cancer colonoids. METHODS: We performed studies of Lgr5(CreERT2); beta-cateninexon3; Rosa26(LSL-rtta-ires-EGFP); TRE-Spdef mice, which express an oncogenic form of beta-catenin in Lgr5-positive ISCs upon administration of tamoxifen and SPDEF upon administration of tetracycline. CRC lines (HCT116 and SW480) were engineered to express inducible tagged SPDEF or vector (control) and subcutaneously injected into immunodeficient NSG mice. We generated SPDEF-inducible human colonoids, including a line derived from normal rectal mucosa (control) and an adenocarcinoma line derived from a patient with germline MUTYH mutation. Full-length and truncated forms of SPDEF were expressed in CRC cells; cells were assayed for beta-catenin activity and studied in immunoprecipitation and chromatin immunoprecipitation assays. RESULTS: Expression of SPDEF was sufficient to inhibit intestinal tumorigenesis by activated beta-catenin, block tumor cell proliferation, and restrict growth of established tumors. In tumor cells with activated b -catenin, expression of SPDEF induced a quiescent state, which was reversed when SPDEF expression was stopped. In mouse and human normal and tumor-derived enteroids/colonoids, those that expressed SPDEF for 3 days were significantly smaller. SPDEF inhibited the transcriptional activity of beta-catenin via a protein-protein interaction, independent of SPDEF DNA binding capacity. SPDEF disrupted beta-catenin binding to TCF1 and TCF3, displacing beta-catenin from enhancer regions of genes that regulate the cell cycle but not genes that regulate stem cell activities. CONCLUSIONS: In studies of mice and human CRC, we found that SPDEF induces a quiescent state in CRC cells by disrupting binding of beta-catenin to TCF1 and TCF3 and regulation of genes that control the cell cycle. In this model, beta-catenin activity determines the proliferation or quiescence of CRC cells based on the absence or presence of SPDEF.