Date of Award
Doctor of Philosophy (PhD)
Biochemistry & Molecular Biophysics
Frank J. Rauscher III
Epithelial-Mesenchymal transition (EMT) of primary tumor cells is a critical trans-differentiation event that contributes to dissemination and metastasis of solid tumors. The process of EMT is controlled by specific DNA-binding transcription factors (TFs) that reprogram the tumor transcriptome. In particular, the canonical EMT-TFs Twist and Snail can induce an EMT program when overexpressed in cancer cells, and both are found upregulated in metastatic cancers. Twist and Snail bind DNA directly, by recognition to variants of the E-Box sequence CANNTG. However, it is unclear how this binding is regulated. We have used a biochemical approach to dissect DNA binding and protein-protein interactions that occur amongst these proteins in order to determine the molecular mechanisms of this regulation. We find that Twist preferentially recognizes a dyad repeat of E-boxes that are not directly bound by Snail. Our data suggest that Twist use its WR domain to recruit Snail into a binding complex through the Snail zinc-finger motifs. We analyzed Twist-Snail complexes in the breast carcinoma cell line SUM1315 and found evidence that it contains an additional protein partner, Sox9. Notably, we report that a native Twist complex can be displaced from its dyad binding site by consensus DNA binding sites for Snail and Sox9 even though these proteins do not contact the Twist dyad site. Taken together, our findings suggest that Snail and Sox9 interact with Twist to regulate its DNA binding ability via protein-protein interactions, thereby allosterically regulating Twist DNA binding and thereby transcriptional regulation of a broad range of EMT associated genes by regulation of this binding. We designate this ternary complex EMTosome and have elucidated the molecular mechanisms by which the DNA binding of it can be regulated. These results may inform efforts to therapeutically target the EMT program in order to target cancer metastasis.
Mccracken, Daniel, "Twist, Snail, And Sox9 Form An Allosterically Regulated Complex, The Emtosome, On A Bipartite E-Box Site." (2020). Publicly Accessible Penn Dissertations. 3971.