Project Summary Gain-of-function Notch mutations are among the most frequent mutations in small B cell lymphomas, including chronic lymphocytic leukemia (CLL) and mantle cell lymphoma (MCL). Harboring Notch mutation is also a predictor of poor outcomes in small B cell lymphomas, which despite recent advances remain incurable with chemotherapy. Together, these observations provide a compelling rationale for focusing on Notch inhibitors as potential therapeutic options in small B cell lymphomas. Unfortunately, progress in treating patients with Notch inhibitors has been stymied partly due to a limited understanding of underlying mechanisms of sensitivity and resistance to Notch inhibitors. Here, we represent a concerted effort to overcome these limitations and fill major gaps in current knowledge by testing our central hypothesis that Notch-driven activation and positioning of enhancers synergize Notch and several crucial signaling pathways to co-activate pro-growth and survival genes in small B cell lymphomas. This premise is based on our preliminary data showing an important consequence of Notch inhibition is decommissioning and repositioning of putative enhancer elements associated with key genes in primary small B cell lymphoma samples, including several components of B cell receptor (BCR) and cytokine signaling pathways. We will test this hypothesis by using cutting-edge functional genomics, chromatin conformation capture, genome engineering, and single-cell resolution genomics and optical assays. We will systematically determine: i) the Notch-dependent enhancer activation and nuclear positioning, and their genomic requirements that together mediate crosstalk between Notch, BCR and cytokine signaling pathways; ii) the rewiring of Notch-driven epigenetic program that enables drug-resistant cells to bypass effects of Notch inhibitors and maintain the expression of critical Notch targets. Together, these complementary studies will greatly expand understanding of epigenetic mechanisms underpinning sensitivity and resistance to Notch inhibitors and provide a clearer rationale for targeting Notch in small B cell lymphoma.