The core objective of this Program Project is to define the relationships and interconnections between hematopoiesis (the differentiation of hematopoietic stem cells, or HSCs) and the progression of cardiovascular diseases (CVDs). To close this gap in knowledge, the Program Project will bring together field-leading researchers to investigate currently unknown factors that mediate hematopoiesis following CVD processes including myocardial infarction, atherosclerosis, and stroke, while also the dissecting contribution of these factors to CVD progression. Therefore, there exists a great opportunity to leverage the capabilities imparted by platform technologies like genome editing to engineer HSCs ex vivo and in mouse models of CVD to help address these outstanding questions. Advances in genome-editing technologies have enabled researchers to precisely alter DNA sequences and gene expression, allowing the rapid generation of biological models to test functional significance of sequence variants. Genome editing enables the permanent heritable alteration of genetic sequence by inducing targeted double-strand breaks (DSBs) in DNA at specified sequences. A complementary approach called epigenome editing leverages the DNA targeting capabilities of the nuclease platforms to transiently alter gene expression by recruiting heterologous effector domains capable of modulating transcriptional states to regulatory regions of the genome, such as promoters or enhancers, without the need for DSBs. Thus, the long-term goal of our Genome Engineering Core (Core 2) is to concurrently develop and optimize genome- and epigenome editing technologies that will enable the rapid, safe, and efficient editing of HSCs to elucidate the connection between hematopoiesis and CVD. Major objectives of Core 2 include supporting the Projects of this proposal by optimizing experimental approaches, and providing genome and epigenome editing reagents and expertise for key experiments of Projects 1 through 4. Core 2 will also develop novel and innovative genome editing, epigenome editing, and lineage tracing platforms to characterize genes, processes, and differentiation of HSCs. Specifically, Core 2 will support Projects 1-4 by enabling efficient and specific genome editing strategies in mouse HSCs, by developing novel epigenome editing reagents for use in HSCs, and by implementing and developing editing-based lineage tracing methods. Successful optimization and development of editing in mouse HSCs pursued by this Core will expedite the ability of Projects 1 through 4 to examine the relationships between hematopoiesis and CVD and more broadly, these advances will be widely applicable to the study and therapeutic intervention of cardiovascular and hematopoietic disorders, and the ex vivo genome and epigenome editing of HSCs.
Genome-editing nucleases are broadly enabling technologies that have revolutionized biomedical research and hold tremendous promise for treating genetic diseases. One important area of research interest is the ex vivo manipulation of hematopoietic stem cells (HSCs) to better understand the link between hematopoiesis and cardiovascular disease processes such as inflammation, myocardial infarction, atherosclerosis, and stroke. The studies proposed herein will characterize and optimize critical genome editing parameters in HSCs, and, most importantly, will support and enable the genome and epigenome editing needs of Projects 1-4 by rapidly implementing and developing effective strategies to manipulate genes related to hematopoiesis and cardiovascular disease.
