PROJECT 4: ABSTRACT Diffuse large B-cell lymphoma (DLBCL) is the most common form lymphoma and is conventionally treated with a combination of chemotherapeutics with the anti-CD20 antibody, Rituximab. Although more than half of patients can be cured with this approach, the remainder have a dire prognosis with a short survival. DLBCL can be classified into two subtypes defined by gene expression profiling: the activated B-cell (ABC) and germinal center B-cell (GCB) subtypes. The ABC subtype of DLBCL has worse clinical outcome compared to GCB-DLBCL, and is characterized by chronically-active signaling driven by its B-cell receptor (BCR) and other signaling mutations leading to constitutive activation of positive-selection pathways (e.g., NFκB). Despite the variability in patient outcome and attempts to target ABC-DLBCL with inhibitors of BCR signaling, there are currently no routinely utilized molecular biomarkers that can be employed to direct a specific therapy. That is, precision medicine does not currently exist for DLBCL. CREBBP is a lysine acetyltransferases that acts as transcriptional co-activator by acetylating histone H3 lysine 27 (H3K27Ac) and other residues. CREBBP is dynamically loaded onto chromatin in centrocytes at regions enriched for NFκB and IRF transcription factor motifs, and genes enriched for BCR and NFκB signaling. In GCB DLBCL, CREBBP is recurrently mutated and we have characterized the effects of these mutations showing that they cause a loss of antagonism to the BCL6 oncogene and a resulting epigenetic and transcriptional suppression of BCL6 target genes including those in the BCR and NFκB signaling pathways. Thus, CREBBP normally functions to activate the expression of BCL6-repressed BCR and NFκB signaling pathway genes following positive selection in the light zone of the germinal center. In line with this, CREBBP serves as co-activators for multiple transcription factors that are active in centrocytes and ABC DLBCL, including IRF4 and NFκB transcription factors REL and RELA. We therefore hypothesize that, despite acting as tumor suppressors in GCB-DLBCL, the function of CREBBP in driving the expression of BCR and NFκB signaling pathway genes may be required in ABC-DLBCL. We will investigate this within our program project application using a number of model systems, including CRISPR-engineered cell lines with inducible expression of surface BCR, genetically engineered mouse models (Core 1), patient-derived xenograft models (Core 1), and cutting- edge epigenomic approaches such as CUT&RUN (Core 2). These same model systems and approaches will be utilized by other projects, allowing us to compare directly to studies of loss of function mutations in CREBBP (Project 1), the role of histone phosphorylation in signaling to chromatin as a precursor to CREBBP loading (Project 2), and the role of CREBBP in aged/autoimmune B-cells (Project 3). Together, this will define the mechanistic role of CREBBP in responding to and regulat...