DNA Sequence Specificity of Alternatively Spliced TCF1E

Abstract

The purpose of this study is to define the activities of a unique DNA binding domain called the C‐clamp, present in alternatively spliced TCF1E. All isoforms of TCF1 contain a sequence specific high mobility group (HMG) box that binds Wnt response elements to control target gene expression. To determine if the C‐clamp also has DNA sequence specificity, Cyclic Amplification and Selection of Targets (CASTing) was performed by isolation of TCF1E‐DNA complexes and progressively enriching high affinity binding sequences with PCR. CASTing revealed that the C‐clamp of TCF1E contains sequence specificity for 5′‐RCCG‐3′ elements independent of the HMG box. Inclusion of RCCG elements in TCF luciferase reporters increased the transcriptional output of TCF1E, validating the CASTing data. The recognition of RCCG elements by TCF1E may be important for its specific activation of target genes such as SP5 and LEF1, which contain RCCG elements adjacent to Wnt response elements in their promoters. The DNA binding activities of the C‐clamp are also important to the regulation of p21 expression. Expression of TCF1E in colon cancer cells caused a p21 dependent stall in cell growth, whereas expression of a non‐DNA binding C‐clamp mutant TCF1E did not upregulate p21 and did not stall cell growth. Thus, the DNA binding activities of the C‐clamp play an important role in determining the transcriptional output of TCF1E.