PROJECT SUMMARY Elevated psychosocial stress – a hallmark of modern, fast-paced societies – has been repeatedly associated with altered immune function and increased coronary heart disease (CHD) risk, but the mechanisms underlying these associations are unclear. DNA methylation, one of the critical and most studied epigenetic processes in humans, has emerged as a key link between environmental exposures and human health. Our long-term goal is to elucidate the epigenetic and other molecular mechanisms through which psychosocial stress contributes to atherosclerotic disease. The overall objective of this application is to define the methylomic differences associated with stress both in whole blood and in distinct immune cell types and to determine how these differences can shape immune function and predict CHD risk. The central hypothesis is that stress drives methylomic patterns that epigenetically upregulate proinflammatory and other immune mediators across distinct blood cell types, thereby contributing to incident CHD. The rationale for this application is that determining stress-associated epigenomic patterns and their functional sequelae in peripheral blood immune cells can yield novel composite predictors and molecular targets that can be leveraged to enhance CHD prevention and treatment. The central hypothesis will be tested by combining large-scale analyses in human cohorts and mechanistic investigations in cell models. Three distinct but complementary specific aims will be pursued: 1) Identify stress-associated methylomic profiles in whole blood that predict incident CHD; 2) Define cell type- specific methylomic patterns that are associated with stress and predict CHD; and 3) Mechanistically dissect how stress epigenetically regulates monocyte function in culture. Aims 1 and 2 will leverage multiple large cohort studies that participate in the NHLBI Trans-Omics for Precision Medicine (TOPMed) program and have suitable psychosocial stress, whole-blood DNA methylation, and/or incident CHD data. Cell type-specific methylomic patterns will be deconvoluted using cutting-edge computational methods recently developed by the research team. Key epigenetic signals will be mechanistically dissected using cell models of monocytes and monocyte- derived macrophages undergoing exposure to physiological levels of stress hormones and targeted DNA methylation editing in culture. This interdisciplinary proposal is innovative as it will integrate large-scale association efforts that apply novel computational methods across multiple TOPMed studies with cutting-edge mechanistic work in immune cell models. The proposed research is significant as it is expected to yield innovative epigenetic predictors and actionable molecular targets that can be leveraged to enhance prevention and treatment of stress-associated CHD.