Markus Brown, Gabrielle Dotson, Tianzuo Zhan, Juliane Kamlah, Hannah Warshawsky, Indika Rajapakse, Michael Boutros, Thomas Ried

• Submitter: Markus Brown (Student or postbac)
• email: markus.brown@nih.gov

The transcription factor, TCF7L2, is the major effector of the WNT signaling pathway in the colon. The expression of TCF7L2 target genes, such as MYC, is required to maintain the stem cell compartment of the large intestine. We investigated the dynamical influence of TCF7L2 silencing on gene expression patterns and nuclear structure in the SW480 colorectal cancer cell line by performing RNA sequencing and Hi-C at multiple time points. Generally, we found that TCF7L2 silencing results in a disproportionate upregulation of gene expression, suggesting an overall repressive role for this transcription factor. We observed changes in topological domain (TAD) structure, which occurred prior to a shift in gene expression. However, many genes with a significant change in expression did not exhibit a change in nuclear structure suggesting that the chromatin conformation was, in most cases, already in a sufficiently accessible conformation. Silencing TCF7L2 lead to the over-expression of LEF1, another WNT signaling transcription factor. LEF1 over-expression over-compensated for the loss of TCF4 in terms of WNT signaling activity as determined by a WNT reporter assay. Despite over-compensation, LEF1 did not rescue TCF4 target gene expression, as determined by MYC and G2/M gene set enrichment analysis (GSEA). We then identified genes which were regulated primarily by TCF4 or LEF1, further supporting our hypothesis that LEF1 drives a distinct transcriptional program. TCF4 and LEF1 also demonstrated cooperativity at specific gene loci, such as MYC. In conclusion, we find that TCF7L2 silencing has a profound effect on the gene expression program of colorectal cancer cells, due in large part to LEF1, which results in the diversified use of WNT signaling activity.