Concealed expansion of immature precursors underpins acute burst of adult HSC activity in foetal liver

One day prior to mass emergence of haematopoietic stem cells (HSCs) in the foetal liver at E12.5, the embryo contains only a few definitive HSCs. It is thought that the burst of HSC activity in the foetal liver is underpinned by rapid maturation of immature embryonic precursors of definitive HSCs, termed pre-HSCs. However, because pre-HSCs are not detectable by direct transplantations into adult irradiated recipients, the size and growth of this population, which represents the embryonic rudiment of the adult haematopoietic system, remains uncertain. Using a novel quantitative assay, we demonstrate that from E9.5 the pre-HSC pool undergoes dramatic growth in the aorta-gonad-mesonephros region and by E11.5 reaches the size that matches the number of definitive HSCs in the E12.5 foetal liver. Thus, this study provides for the first time a quantitative basis for our understanding of how the large population of definitive HSCs emerges in the foetal liver

Recent advances in the structural molecular biology of Ets transcription factors: interactions, interfaces and inhibition

The Ets family of eukaryotic transcription factors is based around the conserved Ets DNA-binding domain. Although their DNA-binding selectivity is biochemically and structurally well characterized, structures of homodimeric and ternary complexes point to Ets domains functioning as versatile protein-interaction modules. In the present paper, we review the progress made over the last decade to elucidate the structural mechanisms involved in modulation of DNA binding and protein partner selection during dimerization. We see that Ets domains, although conserved around a core architecture, have evolved to utilize a variety of interaction surfaces and binding mechanisms, reflecting Ets domains as dynamic interfaces for both DNA and protein interaction. Furthermore, we discuss recent advances in drug development for inhibition of Ets factors, and the roles structural biology can play in their future

Hierarchical organization and early hematopoietic specification of the developing HSC lineage in the AGM region

A CD45-negative population of pre-HSCs develops into definitive HSCs in the AGM region of the embryo

A molecular roadmap of the AGM region reveals BMP ER as a novel regulator of HSC maturation

In the developing embryo, hematopoietic stem cells (HSCs) emerge from the aorta-gonad-mesonephros (AGM) region, but the molecular regulation of this process is poorly understood. Recently, the progression from E9.5 to E10.5 and polarity along the dorso-ventral axis have been identified as clear demarcations of the supportive HSC niche. To identify novel secreted regulators of HSC maturation, we performed RNA sequencing over these spatiotemporal transitions in the AGM region and supportive OP9 cell line. Screening several proteins through an ex vivo reaggregate culture system, we identify BMP ER as a novel positive regulator of HSC development. We demonstrate that BMP ER is associated with BMP signaling inhibition, but is transcriptionally induced by BMP4, suggesting that BMP ER contributes to the precise control of BMP activity within the AGM region, enabling the maturation of HSCs within a BMP-negative environment. These findings and the availability of our transcriptional data through an accessible interface should provide insight into the maintenance and potential derivation of HSCs in culture.Peer reviewe

Runx1 is required for progression of CD41+ embryonic precursors into HSCs but not prior to this

Haematopoiesis in adult animals is maintained by haematopoietic stem cells (HSCs), which self-renew and can give rise to all blood cell lineages. The AGM region is an important intra-embryonic site of HSC development and a wealth of evidence indicates that HSCs emerge from the endothelium of the embryonic dorsal aorta and extra-embryonic large arteries. This, however, is a stepwise process that occurs through sequential upregulation of CD41 and CD45 followed by emergence of fully functional definitive HSCs. Although largely dispensable at later stages, the Runx1 transcription factor is crucially important during developmental maturation of HSCs; however, exact points of crucial involvement of Runx1 in this multi-step developmental maturation process remain unclear. Here, we have investigated requirements for Runx1 using a conditional reversible knockout strategy. We report that Runx1 deficiency does not preclude formation of VE-cad+CD45-CD41+ cells, which are phenotypically equivalent to precursors of definitive HSCs (pre-HSC Type I) but blocks transition to the subsequent CD45+ stage (pre-HSC Type II). These data emphasise that developmental progression of HSCs during a very short period of time is regulated by precise stage-specific molecular mechanisms

A gene trap transposon eliminates haematopoietic expression of zebrafish Gfi1aa, but does not interfere with haematopoiesis

A transposon-mediated gene trap screen identified the zebrafish line qmc551 that expresses a GFP reporter in primitive erythrocytes and also in haemogenic endothelial cells, which give rise to haematopoietic stem and progenitor cells (HSPCs) that seed sites of larval and adult haematopoiesis. The transposon that mediates this GFP expression is located in intron 1 of the gfi1aa gene, one of three zebrafish paralogs that encode transcriptional repressors homologous to mammalian Gfi1 and Gfi1b proteins. In qmc551 transgenics, GFP expression is under the control of the endogenous gfi1aa promoter, recapitulates early gfi1aa expression and allows live observation of gfi1aa promoter activity. While the transposon integration interferes with the expression of gfi1aa mRNA in haematopoietic cells, homozygous qmc551 fish are viable and fertile, and display normal primitive and definitive haematopoiesis. Retained expression of Gfi1b in primitive erythrocytes and upregulation of Gfi1ab at the onset of definitive haematopoiesis in homozygous qmc551 carriers, are sufficient to allow normal haematopoiesis. This finding contradicts previously published morpholino data that suggested an essential role for zebrafish Gfi1aa in primitive erythropoiesi

Defects in ErbB-Dependent Establishment of Adult Melanocyte Stem Cells Reveal Independent Origins for Embryonic and Regeneration Melanocytes

Adult stem cells are responsible for maintaining and repairing tissues during the life of an organism. Tissue repair in humans, however, is limited compared to the regenerative capabilities of other vertebrates, such as the zebrafish (Danio rerio). An understanding of stem cell mechanisms, such as how they are established, their self-renewal properties, and their recruitment to produce new cells is therefore important for the application of regenerative medicine. We use larval melanocyte regeneration following treatment with the melanocytotoxic drug MoTP to investigate these mechanisms in Melanocyte Stem Cell (MSC) regulation. In this paper, we show that the receptor tyrosine kinase, erbb3b, is required for establishing the adult MSC responsible for regenerating the larval melanocyte population. Both the erbb3b mutant and wild-type fish treated with the ErbB inhibitor, AG1478, develop normal embryonic melanocytes but fail to regenerate melanocytes after MoTP-induced melanocyte ablation. By administering AG1478 at different time points, we show that ErbB signaling is only required for regeneration prior to MoTP treatment and before 48 hours of development, consistent with a role in establishing MSCs. We then show that overexpression of kitla, the Kit ligand, in transgenic larvae leads to recruitment of MSCs, resulting in overproliferation of melanocytes. Furthermore, kitla overexpression can rescue AG1478-blocked regeneration, suggesting that ErbB signaling is required to promote the progression and specification of the MSC from a pre–MSC state. This study provides evidence that ErbB signaling is required for the establishment of adult MSCs during embryonic development. That this requirement is not shared with the embryonic melanocytes suggests that embryonic melanocytes develop directly, without proceeding through the ErbB-dependent MSC. Moreover, the shared requirement of larval melanocyte regeneration and metamorphic melanocytes that develops at the larval-to-adult transition suggests that these post-embryonic melanocytes develop from the same adult MSC population. Lastly, that kitla overexpression can recruit the MSC to develop excess melanocytes raises the possibility that Kit signaling may be involved in MSC recruitment during regeneration

The murine placenta contains hematopoietic stem cells within the vascular labyrinth region

SummaryIn the midgestation murine embryo, several major vascular tissues contain hematopoietic stem cell (HSC) activity. These include the aorta-gonad-mesonephros region (AGM), yolk sac, and fetal liver. Recently, the placenta was demonstrated to harbor hematopoietic progenitors, but it was not examined for HSC activity. We demonstrate here that the placenta also harbors adult-repopulating HSCs. Placental HSCs begin to be detected at embryonic day (E) 11, and HSC numbers increase dramatically between E11 and E12, exceeding the numbers in the circulating embryonic blood. Furthermore, all placental HSC activity is restricted to the GFP+ fraction of cells in Ly-6A (Sca-1) GFP transgenic embryos. Cells coexpressing GFP and endothelial markers CD34 and CD31 are found in the embryonic vasculature of the placental labyrinth. Moreover, placental cell expression of other HSC markers and transcription factors suggests that HSC emergence may occur in the placenta, as has been proposed for other embryonic hematopoietic sites

Ginger Stimulates Hematopoiesis via Bmp Pathway in Zebrafish

) has been widely used in traditional medicine; however, to date there is no scientific research documenting the potential of ginger to stimulate hematopoiesis. expression in the caudal hematopoietic tissue area. We further confirmed that Bmp/Smad pathway mediates this hematopoiesis promoting effect of ginger by using the Bmp-activated Bmp type I receptor kinase inhibitors dorsomorphin, LND193189 and DMH1.Our study provides a strong foundation to further evaluate the molecular mechanism of ginger and its bioactive components during hematopoiesis and to investigate their effects in adults. Our results will provide the basis for future research into the effect of ginger during mammalian hematopoiesis to develop novel erythropoiesis promoting agents