Project Summary/Abstract Colon cancer is the 2nd-leading cause of cancer-related deaths in the United States and is one of the best- characterized solid tumors in terms of its common genetic mutations. However, knowledge gaps exist in the basic understanding of these cancers downstream of their genetic profile. For instance, while ~60% of colon cancers follow the typical WNT-driven adenoma-to-carcinoma pathway, approximately 30% follow the “serrated tumor pathway”- often driven by gain-of-function BRAF mutations. Even though serrated tumors have the worst prognosis of colon cancers, there is a large gap in our understanding of the molecular mechanisms. The proposal’s main objective will focus on the tumor-suppressor gene, SMAD4, which is mutated in approximately 57% of all colon tumors, but for which there is little understanding of its molecular mechanisms in colon cancer. The hypothesis is that SMAD4 plays a critical transcriptional regulatory role in serrated tumor pathway, which promotes cancer development and progression. The rationale for this hypothesis is based on preliminary studies that reveal: 1) SMAD4 loss, when combined with activation of BRAF, can trigger serrated tumor formation as rapidly as 1 month in mice, and 2) that SMAD4 binds to regulatory regions of the genome also bound by ß-catenin – the transcriptional effector of the most commonly mutated signaling pathway in colon cancer, the WNT pathway. Aim 1 will use new mouse models to determine how SMAD4 suppresses BRAF-driven serrated tumorigenesis. The goal is to test the hypothesis that SMAD4 suppresses key signaling pathways that are required for serrated tumor development and progression. Aim 2 will use epigenomic approaches to characterize the role of SMAD4 as a transcriptional regulator and will map the interaction of SMAD4 with ß-catenin at the level of DNA-binding, thus detailing the first intersection of these pathways on the colon cancer genome. The proposed studies will test the hypothesis that SMAD4 directly regulates the WNT- signaling pathway by redirecting ß-catenin to tumor-suppressive gene targets in coordination with RUNX3, thus impacting cancer development. The proposed studies are significant in that they will present a new perspective on an understudied, but more deadly, colon cancer tumor type and also identify the cellular mechanisms of tumor development in a commonly mutated tumor-suppressor gene background. These studies would have broad impacts in the cancer research field, and will reveal new targets to identify and treat patients with serrated tumors. With the co-mentorship of Drs. Michael Verzi and Ronald Hart, this proposal's training plan will prepare me for a path towards independence by 1) bolstering my epigenomics training, and 2) guiding my path from a novice in the cancer research field to that of a budding expert poised to make my own impacts towards diagnosing and treating cancer.