Project Funding Details

The master transcription factor (TF) TCF7L2 physically interacts with the co-activator beta-catenin to orchestrate epithelial cell proliferation in the normal intestine and instigate key oncogenic programs in colorectal cancer (CRC). Despite being one of the eight most frequently mutated genes in microsatellite stable CRC patients (approximately 12% frequency), the functional implications of TCF7L2 mutations remain enigmatic. Our preliminary findings suggest that these mutations lead to a loss-of-function hypomorphic phenotype, which reduces the dependency of CRC tumours on TCF7L2 signalling for growth and survival. We propose that deleterious mutations in TCF7L2 reshape beta-catenin-driven transcription and cellular responses by engaging alternative partnerships with distinct TFs and co-regulators. These alternative hubs are expected to modulate gene expression by redefining the beta-catenin cistrome and influencing chromatin accessibility. Our goal is to elucidate how TCF7L2 mutations impact the interplay between cis- and trans-acting elements and chromatin organisation in CRC. Through a multidimensional interactome analysis, we seek to map and functionally investigate: i) the impact of mutations in TCF7L2 on its ability to interact with transcriptional regulators, chromatin modifiers and DNA responsive elements ii) the composition of the multiprotein assemblies that nucleate around beta-catenin in the presence of wild-type or mutant TCF7L2; iii) the protein-DNA associations governed by the TFs, co-activators and co-repressors operating in such assemblies; and iv) the chromatin accessibility landscapes that typify TCF7L2 wild-type and mutant CRC, and the relationships between chromatin states and TCF7L2/beta-catenin-regulated DNA regions dictating defined transcriptional outputs. This comprehensive census of proteomic, epigenetic and transcriptional connectivities will lay the groundwork for characterising critical components serving as potential therapeutic targets with applicative significance. TCF7L2 wild-type and mutant patient-derived CRC tumouroids will be utilised for all investigations. Affinity purification and mass spectrometry-based proteomics will be deployed to identify functionally relevant proteins showing distinct binding ability to beta-catenin and/or TCF7L2 based on TCF7L2 mutational status. The chromatin-binding patterns of beta-catenin complexes in CRC tumouroids with wild-type and mutant TCF7L2 and the accessibility of TCF7L2/beta-catenin-regulated chromatin regions will be explored through ChIP-seq and ATAC-seq analyses, respectively. Drugs that directly or indirectly target the TCF7L2-beta-catenin complex, epigenetic inhibitors, and compounds against newly identified interactors endowed with potential pharmacological actionability will be tested in TCF7L2 wild-type and mutant tumouroids, followed by in vivo validation in matched patient-derived xenografts. The project integrates genetic, biological, and computational approaches to yield a functionally validated catalogue of trans- and cis-regulatory elements and chromatin-remodelling cofactors that differentially specify beta-catenin's tumorigenic programs in TCF7L2 wild-type and mutant CRC. Ultimately, the definition of a multidimensional protein-protein and protein-DNA interactome will provide a foundational understanding of beta-catenin-dependent transcriptional control, offering potential targets for rational therapeutic strategies. The proposed research will unveil novel insights into the role of the beta-catenin-TCF7L2 complex in CRC, shedding light on its regulatory activities and opening potential avenues for pharmacological intervention. The identification of specific cofactor interactions affecting TF gene regulation holds promise for delivering original and impactful biological discoveries, with translational implications for advancing CRC treatment.