Gene Regulatory Networks Governing the Generation and Regeneration of Blood

Abstract

Blood is an example of a highly regenerative tissue and its regeneration depends on the presence of stem cells residing in the bone marrow in humans. A better understanding of how these stem cells are programmed would benefit their use in clinical practice and shed light on the mechanisms by which the unique properties of stem cells are established. Our approach is to delineate the gene regulatory networks (GRNs) that specify these cells during their development in the embryo, and we use the amphibian experimental model because a wealth of evidence shows that the mechanisms used are conserved in mammals including humans. Blood stem cells are made during the intraembryonic wave of hematopoiesis during embryonic development where they emerge from endothelial precursors in the floor of the dorsal aorta (DA). These cells are derived from lateral plate mesoderm and so we have focused on the subset of cells in the lateral plate mesoderm fated to become blood and endothelium known as definitive hemangioblasts. We have found that their programming results from the activities of vascular endothelial growth factor A (VEGFA) and bone morphogenetic protein (BMP) signaling and the inhibition by miRNA of transforming growth factor beta signaling. VEGFA is first generated in the somites adjacent to the lateral plate mesoderm, and one of the responses of the lateral plate mesoderm is to activate endogenous VEGFA expression. BMP has multiple inputs into the programming of these cells via the activation of the transcription factor (TF), Gata2, and of the VEGFA receptor. These actions culminate in the expression of the leukemia-associated TF, Scl/Tal1, which is essential for blood fate specification. The activity of VEGFA in driving endothelial development resides in the small isoform, but the medium and large isoforms are required to initiate the blood stem cell program in the floor of the DA. The expression of the small isoform is dependent on the blood TF with leukemia connections, Tel1/Etv6, whereas the larger isoforms depend on another transcription-associated factor with leukemia connections, Eto2, raising the possibility that the regulation of VEGFA expression may be the mode of action of these leukemic factors. The action of Tel1/Etv6 in directly activating VEGFA expression in the somites was unexpected because this TF had only been reported to repress transcription. Using chromatin immunoprecipitation technology, we were able to show that Tel1/Etv6 does indeed work by repressing the expression of a VEGFA repressor, FoxC3, but it also acts directly to activate VEGFA expression, working together with Klf4. Finally, we have also looked at the mesodermal population that gives rise to the earlier waves of hematopoiesis,

Iterative Single-Cell Analyses Define the Transcriptome of the First Functional Hematopoietic Stem Cells

“…This transition involves changes from an endothelial transcriptional program to a program promoting HC identity, morphology, and function (Swiers et al, 2013). Although the precise program directing the generation of HSCs during EHT is as yet unknown, important molecular and physiological aspects of HSC generation are conserved between zebrafish, chick, Xenopus, mouse, and human (Ciau-Uitz and Patient, 2019;Dzierzak and Bigas, 2018;Ivanovs et al, 2017).…”

Unexpected redundancy of Gpr56 and Gpr97 during hematopoietic cell development and differentiation

“…In vertebrate embryos, HSCs arise in the aorta-gonad mesonephros (AGM) region from hemogenic endothelial cells (HECs). HECs undergo a highly-conserved 'endothelial-tohematopoietic transition' (EHT) process (Ciau-Uitz & Patient, 2019, Dzierzak & Bigas, 2018, Dzierzak & Speck, 2008, Jaffredo, Nottingham et al, 2005. Transcriptomics and loss-of-function studies have identified candidate regulators/signaling pathways in transitioning cells (Gao, Chen et al, 2020, Li, Gao et al, 2017, Lichtinger, Ingram et al, 2012, Moignard, Woodhouse et al, 2015, Porcheri, Golan et al, 2020, Swiers, Baumann et al, 2013, Zhou, Li et al, 2016.…”

Gata2-regulated Gfi1b expression controls endothelial programming during endothelial-to-hematopoietic transition

“…In the AGM region, definitive HSPCs are generated through a trans-differentiation process from a specialized endothelial cell (EC) compartment with hematopoietic potential (hemogenic endothelial cells or HECs). This process of endothelial-to-hematopoietic transition (EHT) is conserved between mammalian and non-mammalian vertebrates (Ciau-Uitz and Patient, 2019; Ivanovs et al, 2017; Bertrand et al, 2010; Boisset et al, 2010; Kissa and Herbomel, 2010; Zovein et al, 2008; de Bruijn et al, 2002; North et al, 2002; Medvinsky and Dzierzak, 1996; Muller et al, 1994; Dieterlen-Lievre, 1975) . In mice, EHT events occur between embryonic days (E)10.5-E12.5.…”