Abstract: The activities of innate and adaptive immune cells need to be precisely coordinated for an effective immune response to a wide-range of pathogens. There are many similarities between immune responses in different species, but each species also has adapted to combat unique pathogenic insults. Mechanistically, structural variation in the genome between species plays a prominent role in the acquisition (or loss) of regulatory events involved in defining the functional activity of immune cells. Structural variation refers to genome rearrangements such as translocations, insertions, amplifications, and inversions. Structural variation in the genome between species has the potential to substantially alter the placement of genes and regulatory elements, including splitting apart clusters of genes with similar functions, relocating genes to different chromosomes, and changing the orientation of loci. The functional consequences for changes in the genome between species are based on how each change affects the regulatory events responsible for controlling gene expression. Therefore, it is important to define how structural variation between species affects regulatory principles to improve our ability to relate findings from mechanistic and preclinical studies in model organisms to the regulation of the human immune response. In this application, we will define how structural variation between the mouse and human genomes influences the regulatory events involved in controlling genes that are differentially expressed in mouse and human immune cells. We will determine how structural variation between the mouse and human genomes affects the 1) topology of the genome, 2) regulatory events that position topological domain boundaries, 3) enhancer landscape available to genes, and 4) long-range enhancer-promoter interactions for genes with differences in expression between mouse and human immune cells. We will test the functional consequences for divergent regulatory events between mouse and human immune cells, with a focus on defining the regulatory events contributing to differences in the LPS-inducible expression of Nos2 (encodes inducible nitric oxide synthase; iNOS) in mouse and human macrophages, and we will use this to build a mouse model reflecting human expression. The mouse is one of the most widely used preclinical models of the human immune response, and the studies in our proposal will aid in understanding how structural variation between the genomes of model organisms and the human genome contribute to species acquiring different cell-type and stimulation-dependent gene expression patterns and functions. It will also make it possible to predict how mechanisms defined in mice translate to human as well as build mouse models that better reflect human immune responses for pre-clinical studies.