PROJECT SUMMARY The mitochondrial network is shaped by fusion and fission dynamics that ultimately influence the mitochondrial capacity to utilize fuels. Our recent dissection of metabolic circuits in diffuse large B-cell lymphoma (DLBCL) has identified heterogeneity of mitochondrial architecture and biochemical networks in DLBCL subtypes with distinct patterns of fuel utilization. OxPhos-DLBCLs show a net increase in mitochondrial fragmentation and rely on mitochondrial fatty acid oxidation (FAO) for survival and proliferation independent of B-cell receptor (BCR) signaling. This is distinct from non-OxPhos/Warburg type DLBCLs that are BCR-dependent, rely on glycolysis and have connected mitochondrial network. Importantly, blocking fragmentation in OxPhos-DLBCLs reduces mitochondrial FA utilization capacity but does not alter consumption of other fuels. The above observations indicate a specific requirement for fragmentation in facilitating mitochondrial handling of FAs, and link mitochondrial morphologic heterogeneity to fuel choice and metabolic specialization in DLBCL subtypes. In response to RFA-CA-17-017 PQ5, the proposed studies examine the mechanisms and consequences of this link and its relevance to tumorigenesis. In Aim 1, we will define the mechanistic determinants of the net increase in mitochondrial fragmentation in OxPhos- vs BCR-DLBCLs, including changes in fusion and fission rates at the level of individual mitochondria and alterations in mitochondria-shaping proteins. We will also address the long-term consequences of altered mitochondrial fragmentation in growth and survival of DLBCL subtypes in vitro and in vivo. In Aim 2, we will learn about the consequence of mitochondrial fragmentation for fuel utilization in general and FAO in particular. A combination of carbon tracing and biochemical studies will be undertaken to determine the mechanisms underlying regulation of mitochondrial FA handling by mitochondrial fragmentation in OxPhos-DLBCLs. In Aim 3, we will determine how mitochondrial architecture and fuel metabolism are modulated by BCR-initiated signals, and probe the relevance of these mitochondrial pathways to the sensitivity of BCR-DLBCLs to clinically-relevant BCR inhibitors. Together, these studies can provide important conceptual advancement and mechanistic insights into how the mitochondrial morphologic specializations in DLBCLs are intertwined with fuel utilization to support tumor growth.