PROJECT SUMMARY/ABSTRACT During aging, the ability of neural stem cells (NSCs) in the brain to form new neurons is reduced, but the molecular mechanisms underlying the deterioration of NSC function remain unclear. There is currently a critical need to understand the mechanisms by which NSCs are activated to form neurons, and why this process declines with age. The long term goal is to identify the mechanisms responsible for the loss of NSC function with age, and discover interventions that harness the regenerative capacity of these cells to increase cognitive function in aged and diseased individuals. The objective of this proposal is to identify the mechanisms by which the conserved “pro-longevity” transcription factor, FOXO3, preserves NSC quiescence during aging. The central hypothesis is that FOXO3 directly regulates a network of target genes and pathways that are critical for preserving NSCs during aging. This hypothesis will be tested by pursuing the following specific aims: 1) Determine the specific pathways regulated by FOXO3 in NSCs that preserve the quiescent state; 2) Investigate how FOXO3 and ASCL1 govern the balance between stem cell preservation and neurogenesis, a process that is drastically altered with age; and 3) Determine the extrinsic inputs that control FOXO3 activity and function. The first aim will be accomplished by combining a model of primary adult mouse NSC quiescence with loss of function and overexpression approaches to test the hypothesis that FOXO3 directly promotes quiescence by regulating specific genes and pathways. The second aim will be performed using methods to reveal the dynamic and antagonistic interaction between FOXO3 and ASCL1, and test the extent to which levels or activity of these factors are responsible for reduced activation of NSCs with age. The third aim will be accomplished using a combination of mouse genetics and molecular methods to test the hypothesis that BMP signaling directly regulates FOXO3 expression in vivo to promote NSC quiescence during aging. The outcome of this project will be the identification of the mechanisms by which FOXO3 regulates NSC function, how these mechanisms deteriorate with age, and reveal a strategy to counter the loss of NSC function during aging. This work is significant because it will determine why NSC activation is reduced in the aged brain, and uncover strategies to reverse it. This proposed research is innovative because it will use a unique system to elucidate the direct, genome-wide mechanisms that promote adult NSC quiescence, and parlay these findings into the in vivo setting. This work will provide key mechanistic insight into how gene networks are coordinated in young and aging NSCs, and have the potential to reveal new mechanisms underlying cognitive decline during aging.