Enter the text here that is the new abstract information for your application. This section must be no longer than 30 lines of text. During fetal life or in adulthood, hematopoietic stem cells (HSCs) are positioned adjacent to venous sinusoids, specialized veins that provide support for stem cell homeostasis. Previously, we found that sinusoids interact directly with HSCs in a process we termed “cuddling,” and this supports HSC division. We investigated the transcriptional program of venous sinusoids and published a method to overexpress transcription factors (TFs) to reprogram ectopic regions of the zebrafish embryo to become venous sinusoids. These new vessels support local blood stem cell self-renewal. We now plan to examine vascular reprogramming in adults to provide an alternative location for hematopoietic stem cells to form blood during stress or infiltrative disorders of the marrow. Through comparing single cell RNA-seq analyses of adult liver sinusoids and marrow sinusoids from zebrafish, mice and humans, we found five new TFs specifically expressed in the marrow sinusoids. We plan to overexpress these TFs in the hope of transforming the adult liver, which typically does not support hematopoiesis, into a hematopoietic site. Preliminary data overexpressing TFEC and MAFB using a liver-sinusoid-specific enhancer demonstrates activation of much of the marrow sinusoid fate in vivo. Overexpression of these factors in human iPSC-derived venous sinusoids leads to enhanced proliferation of hematopoietic stem and progenitor cells (HSPCs) in co-culture. We plan to overexpress the human orthologs of other TFs found in zebrafish to examine the support of HSPCs in vitro. The reprogrammed venous sinusoids could be used to expand HSCs in vitro. In an effort to understand how niche endothelial cells (ECs) cuddle HSCs, we undertook a chemical genetic screen to find small molecules that regulate the process. We found that antagonists of the G-coupled receptor, cysteinyl leukotriene receptor 1 (CysLTR1), stimulate stem cell division specifically nearby cuddling ECs in vivo. The receptor is specifically expressed by the sinusoids in zebrafish and humans. Using lipidomics, we found that the CysLTR1 ligand, LTD4, is present in the zebrafish larval niche where HSCs divide. LTD4 suppresses the HSC division nearby cuddling ECs. Using genetics, chemical biology, and biochemistry, we plan to define the mechanism by which LTD4 regulates endothelial cells to support HSC self-renewal. Our work will provide a new understanding of the mechanism of endothelial cuddling and its role in HSC division, potentially leading to new chemical or cell therapy to induce HSC division and maintenance for the treatment of blood disorders.