PROJECT SUMMARY/ABSTRACT The E2F transcriptional program, which controls cell cycle commitment and progression in proliferating cells, is upregulated following DNA damage in neurons. Neuronal DNA damage and cell cycle dysregulation are features of neurodegeneration that have also been associated with psychiatric diseases. Although the function of E2F during proliferation has been extensively studied, the role of E2F in nonproliferating cells, such as in neurons, has received less attention. Emerging evidence indicates that E2F plays a role in DNA damage repair, independent of its role in cell cycle entry. This role may be especially important in neurons, which are uniquely vulnerable to DNA damage. Although E2F induction has been linked to resolution of DNA damage in bulk-cell preparations of neurons, such bulk analyses fail to account for potentially confounding heterogeneity within samples. Single cell analysis is necessary to examine the relationship between E2F activity and DNA damage and to link E2F dynamics to functional outcomes in the same cells. The Meyer Lab specializes in single-cell analysis of high-throughput microscopy data, and the lab has recently developed a fluorescent biosensor of E2F activity for this purpose. Quantitative microscopy using live- and fixed-cell readouts of E2F activity and DNA damage will allow signaling history to be mapped to cell fate outcomes at single-cell resolution in thousands of cells. My hypothesis is that in postmitotic neurons, E2F is reversibly activated to drive DNA repair without DNA replication following sublethal DNA damage; however, if DNA damage activates E2F beyond the threshold for S-phase entry, E2F induces DNA replication and apoptosis. By characterizing how the E2F program is regulated in neurons and how it contributes to DNA damage repair, this study will support the identification of potential treatment targets for preserving genetic integrity in brain disorders characterized by genotoxic stress. This research project represents an important component of my training for a career as an independent investigator, employing high-throughput single-cell methods to study the biology of complex diseases. My long- term goal is to become a physician-scientist, practicing as a psychiatrist while also running a basic science lab in an academic hospital. The plan outlined in this proposal along with the mentorship of my sponsor, thesis committee, and the leadership of the Tri-I MD- PhD program, will help me achieve these career aspirations.