A WNT/p21 Circuit Directed by the C-Clamp, a Sequence-Specific DNA Binding Domain in TCFs

Abstract

The lymphoid enhancer factor 1/T cell factor (LEF/TCF) family of transcription factors are downstream effectors of the WNT signaling pathway, which drives colon tumorigenesis. LEF/TCFs have a DNA sequence-specific high-mobility group (HMG) box that binds Wnt response elements (WREs). The "E tail" isoforms of TCFs are alternatively spliced to include a second DNA binding domain called the C-clamp. We show that induction of a dominant negative C-clamp version of TCF1 (dnTCF1E) induces p21 expression and a stall in the growth of DLD1 colon cancer cells. Induction of a C-clamp mutant did not efficiently induce p21, nor did it stall cell growth. Microarray analysis revealed that induction of p21 by wild-type dnTCF1E (dnTCF1E WT ) correlated with a decrease in expression of multiple p21 suppressors that act at multiple levels from transcription (SP5, YAP1, and RUNX1), RNA stability (MSI2), and protein stability (CUL4A). We show that the C-clamp is a sequence-specific DNA binding domain that can make contacts with 5=-RCCG-3= elements upstream or downstream of WREs. The C-clamp-RCCG interaction was critical for TCF1E-mediated transcriptional control of p21-connected target gene promoters. Our results indicate that a rapid-response WNT/p21 circuit is driven by C-clamp target gene selection.

Sp5 negatively regulates the proliferation of HCT116 cells by upregulating the transcription of p27

“…The present study showed that the expression of Sp5 is not upregulated in HCT116 cells, in which Wnt signaling is constitutively activated. Although other studies have reported that Sp5 represses the expression of p21 and promotes the growth of MCF-7 cells (11,14,17), no significant changes could be detected in the p21 expression in the HCT116 cells transfected with Sp5 in the present study. This discrepancy may be due to the differences in cell types and the different expression levels of the Sp family of proteins.…”

“…It has been reported that overexpression of Sp5 suppresses p21 expression (11,14,17). However, in the present study, RT-qPCR analysis revealed that overexpression of mSp5 did not affect the expression levels of p21 in HCT116 cells (Fig.…”

“…Sp5 contains three conserved Cys2His2 zinc finger domains at its C-terminus, which bind the GC-box (GGGCGG) or closely associated sequences (15,16). Sp5 directly represses the transcription of the cell cycle inhibitor p21 by binding to the GC-boxes located in its promoter region, and exogenous expression promotes the growth of breast cancer MCF-7 cells, which express no detectable Sp5 protein (11,14,17). By contrast, overexpression of Sp5 inhibits cell growth and transformation mediated by oncogenic KRAS in NIH3T3 cells (18).…”

Differential Effects of Hepatocyte Nuclear Factor 4α Isoforms on Tumor Growth and T-Cell Factor 4/AP-1 Interactions in Human Colorectal Cancer Cells

“…The full-length human HNF4␣2 (NM_000457) and HNF4␣8 (NM_175914) cDNAs in pcDNA3.1 were gifts from Christophe Rachez at Pasteur Institute, Paris, France (14,46). The human ApoB.-85.-47.E4.Luc luciferase reporter construct and Flag-dnTCF1E1 (Flag.dnTCF1) and Flag-TCF4E2 (Flag.TCF4) expression vectors have been described previously (29,47).…”

A Genomic Analysis Pipeline and Its Application to Pediatric Cancers

“…The targets and gene programs that are misregulated in these cells are specified by the DNA binding specificities of the LEF/TCFs. The factor studied here, TCF1E, contains two DNA binding domains, a WRE binding domain and a C-clamp binding domain [77].…”