CDX2 inhibits the proliferation and tumor formation of colon cancer cells by suppressing Wnt/β-catenin signaling via transactivation of GSK-3β and Axin2 expression

Abstract

Caudal-related homeobox transcription factor 2 (CDX2), an intestine-specific nuclear transcription factor, has been strongly implicated in the tumourigenesis of various human cancers. However, the functional role of CDX2 in the development and progression of colorectal cancer (CRC) is not well known. In this study, CDX2 knockdown in colon cancer cells promoted cell proliferation in vitro, accelerated tumor formation in vivo, and induced a cell cycle transition from G0/G1 to S phase, whereas CDX2 overexpression inhibited cell proliferation. TOP/FOP-Flash reporter assay showed that CDX2 knockdown or CDX2 overexpression significantly increased or decreased Wnt signaling activity. Western blot assay showed that downstream targets of Wnt signaling, including β-catenin, cyclin D1 and c-myc, were up-regulated or down-regulated in CDX2-knockdown or CDX2-overexpressing colon cancer cells. In addition, suppression of Wnt signaling by XAV-939 led to a marked suppression of the cell proliferation enhanced by CDX2 knockdown, whereas activation of this signaling by CHIR-99021 significantly enhanced the cell proliferation inhibited by CDX2 overexpression. Dual-luciferase reporter and quantitative chromatin immunoprecipitation (qChIP) assays further confirmed that CDX2 transcriptionally activates glycogen synthase kinase-3β (GSK-3β) and axis inhibition protein 2 (Axin2) expression by directly binding to the promoter of GSK-3β and the upstream enhancer of Axin2. In conclusion, these results indicated that CDX2 inhibits the proliferation and tumor formation of colon cancer cells by suppressing Wnt/β-catenin signaling.

DGCR8/ZFAT-AS1 Promotes CDX2 Transcription in a PRC2 Complex-Dependent Manner to Facilitate the Malignant Biological Behavior of Glioma Cells

“…Studies have found that CDX2 inhibits colon cancer cell proliferation and tumor formation. 25 During the development of mice, the spontaneous loss of CDX2 heterozygosity causes the transformation of squamous epithelial cells, leading to colon tumors. 44,45 In the current study, we discovered that CDX2 expression was increased in glioma tissues, U87 cells, and U251 cells, and CDX2 knockdown significantly inhibited glioma cell proliferation, migration, and invasion, and it promoted apoptosis.…”

“…Studies have found that overexpression of CDX2 inhibits the proliferation and tumor formation of colon cancer cells. 25 CDX2 expression decreased progressively with the advancing stage of gastric cancers and showed a negative correlation with the depth of tumor invasion. 26 RRM2B encodes one of two forms of the ribonucleotide reductase R2 subunit and is necessary for normal DNA repair and mtDNA synthesis in non-proliferating cells.…”

“…Previous studies have reported that CDX2 transcriptionally activates the expression of glycogen synthase kinases GSK-3b and Axin2 by directly binding to the promoter of GSK-3b and an enhancer upstream of Axin2. 25 Comparison of the adult mouse and human intestinal cells revealed that CDX2 binds to and activates different target genes during development. 65 To elucidate the mechanism through which CDX2 regulates RRM2B expression, we used the bioinformatics software JASPAR and identified that the RRM2B promoter region possessed a potential binding site for CDX2.…”

“…A Pierce magnetic RNA-protein pull-down kit (Thermo Fisher Scientific, USA) was used to detect the interaction between ZFAT-AS1 and EZH2 protein according to the manufacturer's instructions. ZFAT transcripts, ZFAT-AS1, and negative RNA control (poly(A) 25 RNA) were labeled using desthiobiotinylated cytidine bisphosphate and T4 RNA ligase according to the Pierce RNA 3 0 -End Desthiobiotinylation Kit procedure. The biotin-labeled negative RNA control (poly(A) 25 RNA), ZFAT transcripts, and ZFAT-AS1 were captured using streptavidin magnetic beads in RNA capture buffer for 30 min at room temperature.…”

“…ZFAT transcripts, ZFAT-AS1, and negative RNA control (poly(A) 25 RNA) were labeled using desthiobiotinylated cytidine bisphosphate and T4 RNA ligase according to the Pierce RNA 3 0 -End Desthiobiotinylation Kit procedure. The biotin-labeled negative RNA control (poly(A) 25 RNA), ZFAT transcripts, and ZFAT-AS1 were captured using streptavidin magnetic beads in RNA capture buffer for 30 min at room temperature. Beads were washed twice in 20 mM Tris (pH 7.5), once in protein-RNA binding buffer, and U87 extract was added.…”