Dissecting BMP signaling input into the gene regulatory networks driving specification of the blood stem cell lineage

Abstract

Hematopoietic stem cells (HSCs) that sustain lifelong blood production are created during embryogenesis. They emerge from a specialized endothelial population, termed hemogenic endothelium (HE), located in the ventral wall of the dorsal aorta (DA). In Xenopus, we have been studying the gene regulatory networks (GRNs) required for the formation of HSCs, and critically found that the hemogenic potential is defined at an earlier time point when precursors to the DA express hematopoietic as well as endothelial genes, in the definitive hemangioblasts (DHs). The GRN for DH programming has been constructed and, here, we show that bone morphogenetic protein (BMP) signaling is essential for the initiation of this GRN. BMP2, -4, and -7 are the principal ligands expressed in the lineage forming the HE. To investigate the requirement and timing of all BMP signaling in HSC ontogeny, we have used a transgenic line, which inducibly expresses an inhibitor of BMP signaling, Noggin, as well as a chemical inhibitor of BMP receptors, DMH1, and described the inputs from BMP signaling into the DH GRN and the HE, as well as into primitive hematopoiesis. BMP signaling is required in at least three points in DH programming: first to initiate the DH GRN through gata2 expression, then for kdr expression to enable the DH to respond to vascular endothelial growth factor A (VEGFA) ligand from the somites, and finally for gata2 expression in the DA, but is dispensable for HE specification after hemangioblasts have been formed.

The roles and controls of GATA factors in blood and cardiac development

“…Success of the BMP inhibition in each case was monitored by the presence of phosphorylated SMAD‐1, 5 or 8, the downstream mediators of BMP signalling. We found that BMP input is required at three points during the development of the HSCs: firstly, for the initial expression of GATA‐2 in the DLP/DHs, later for the expression of KDR/FLK‐1 required for the response to VEGF‐A signalling, and thirdly for the expression of GATA‐2 in the DA …”

“…However, BMP is required for the whole of lateral plate mesoderm formation and general patterning of the early embryo. Therefore, to test the specific role around the time when GATA‐2 expression was turned on in the DLP/DHs, we had to make use of conditional inhibition of BMP signalling . We therefore obtained a transgenic Xenopus line expressing the BMP inhibitor, Noggin, from a heat shock‐inducible promoter, and separately a chemical inhibitor of BMP signalling that could be added to the water housing the experimental embryos.…”

“…Therefore, to test the specific role around the time when GATA-2 expression was turned on in the DLP/DHs, we had to make use of conditional inhibition of BMP signalling. 16 We therefore obtained a transgenic Xenopus line expressing the BMP inhibitor, Noggin, from a heat shock-inducible promoter, and separately a chemical inhibitor of BMP signalling that could be added to the water housing the experimental embryos. Success of the BMP inhibition in each case was monitored by the presence of phosphorylated SMAD-1, 5 or 8, the downstream mediators of BMP signalling.…”

“…We found that BMP input is required at three points during the development of the HSCs: firstly, for the initial expression of GATA-2 in the DLP/DHs, later for the expression of KDR/FLK-1 required for the response to VEGF-A signalling, and thirdly for the expression of GATA-2 in the DA. 16…”

Gene regulatory networks and network models in development and evolution

“…One thing that is clear from the papers that follow is that the impact of thinking on gene networks in bioscience is truly broad and deep. The topics span a wide range, from the differentiation of germ cells (1), immunology (2,3), and the molecular basis of celltype differentiation (4)(5)(6)(7)(8)(9)(10)(11), to paleontology (12,13) and behavior (14). The central role of network thinking is revealed in the way it allows for both the design of testable experimental frameworks to probe complex biological processes and the conceptual framework for understanding biological mechanisms.…”