Atherosclerosis is an inflammatory disease of the arterial walls and is the major cause of heart attack and stroke. Atherosclerosis is localized to curves or branches in the vasculature where disturbed blood flow alters endothelial cell (EC) gene expression and induces EC dysfunction. Epigenetics controls aberrant gene expression in many diseases, but the mechanism of flow-induced epigenetic gene regulation in ECs via DNA methylation has not been well studied until very recently. The goal of this project was to determine how the DNA methylome responds to flow, causes altered gene expression, and regulates atherosclerosis development. Here, we found that d-flow increases DNA Methyltransferase 1 (DNMT1) expression in ECs, and we hypothesized that this causes a shift in the EC methylome and transcriptome towards a pro-inflammatory, pro-atherosclerotic gene expression program, and further that this leads to atherosclerosis development. To test this hypothesis, we employed both in vitro and in vivo experimental approaches combined with genome-wide studies of the transcriptome and DNA methylome according to the following three specific aims: 1) to elucidate the role of DNA Methyltransferase 1 in EC function, 2) to uncover the DNA methylation-dependent EC gene expression response to flow, and 3) to discover and examine master regulators of EC function that are controlled by DNA methylation. The work presented here has resulted in new knowledge about the epigenetic EC shear response, details the previously unstudied EC methylome, and implicates specific loci within the genome for additional studies on their role in EC biology and atherosclerosis. This work provides a foundation for novel and more targeted therapeutic strategies for CVD.