Cellular barcoding of developmental hematopoiesis

PROJECT SUMMARY/ ABSTRACT Hematopoietic stem cells (HSCs) lie at the top of the blood hierarchy and are capable of giving rise to all blood cells of an organism. Consequently, their use has enormous therapeutic potential for the treatment of blood diseases, and generation of HSCs in vitro is a central aim in regenerative biology. Despite this clinical need, we

lack protocols that allow us to efficiently generate HSCs in vitro that are capable of long-term engraftment and multi-lineage output. A major hindrance is our incomplete understanding of how HSCs are made in vivo. For instance, although it is established that blood cells develop from endothelial cells in multiple sites throughout the

embryo, we still do not know which embryonic sites produce long-term HSCs, nor how site of origin impacts on life-long stem cell function or behavior. Furthermore, we are limited in our understanding of the intrinsic and extrinsic cues driving functional heterogeneity in hemogenic endothelial cells. This project proposes to use

powerful next-generation barcoding technology to enrich our understanding of the embryonic origin of HSCs and the hemogenic endothelial cell states that give rise to blood to allow us to harness the process in vitro. Dr. Bowling conducted her graduate work in developmental biology and during her postdoctoral training

has focused on the development of next-generation DNA barcoding tools for performing single cell, inducible cell lineage tracing in vivo. Equipped with this skillset, Dr. Bowling plans to use innovative cellular barcoding techniques to, for the first time, catalog the precise endothelial origins of long-lived blood progenitors in the

mammalian embryo (Aim 1). Furthermore, she will perform in-depth characterization of the heterogeneous endothelial cell states that give rise to distinct blood cells in the embryo (Aim 2). The knowledge generated from these experiments have the potential to answer major, long-standing questions in the field of developmental

hematopoiesis and transform our basic understanding of the steps leading to blood generation, and therefore revolutionize protocols for HSC generation in vitro. Dr. Bowling is supported by a panel of mentors and consultants who are world-class researchers in hematology, developmental biology, and technology development. Her mentors Drs. Fernando Camargo and

Leonard Zon have made exceptional contributions to the field of hematopoiesis and also have outstanding track- records for mentorship. Dr. Bowling will gain further scientific training and career development support from her scientific committee: Drs Jay Shendure, Trista North and Berthold Gottgens. Finally, she will benefit from carrying

out her research program in the scientifically stimulating and resource-rich environment of Boston Children’s Hospital and Harvard Medical School. The aims in this proposal will allow Dr. Bowling to build on her skills to gain expert knowledge in the computational analysis of sequencing datasets and the use of induced pluripotent

stem cells, in preparation for her transition to independence. As a result, she will establish a unique niche for resolving important, clinically-relevant questions in hematopoietic development as an independent researcher.