The transcription factor GATA2 has pivotal roles in hematopoiesis. Germline GATA2 mutations result in GATA2 haploinsufficiency characterized by immunodeficiency, bone marrow failure and predispositions to myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). Clinical symptoms in GATA2 patients are diverse and mechanisms driving GATA2 related phenotypes are largely unknown. To explore the impact of GATA2 haploinsufficiency on hematopoiesis, we generated a zebrafish model carrying a heterozygous mutation in gata2b, an orthologue of GATA2. Morphological analysis revealed progression of myeloid and erythroid dysplasia in gata2b+/- kidney marrow (KM). Single cell RNA sequencing on KM cells showed that the erythroid dysplasia in gata2b+/- zebrafish was preceded by a differentiation block in erythroid progenitors, hallmarked by downregulation of cytoskeletal transcripts, aberrant proliferative signatures and ribosome biogenesis. Additionally, transcriptional and functional analysis of Gata2b haploinsufficient hematopoietic stem cells (HSCs) indicated that proliferative stress within the HSC compartment possibly contributes to the development of myeloid and erythroid dysplasia in gata2b+/- zebrafish.

The first hematopoietic stem cells (HSCs) are formed through endothelial-to-hematopoietic transition (EHT) events during embryonic development. The transcription factor GATA2 is a crucial regulator of EHT and HSC function throughout life. Because GATA2 haploinsufficiency patients have inborn mutations, prenatal defects are likely to have an influence on disease development. In mice, Gata2 haploinsufficiency (Gata2+/-) reduces the number and the functionality of embryonic hematopoietic stem and progenitor cells (HSPCs) generated through EHT. However, the embryonic HSPC pool is heterogeneous and the mechanisms underlying this defect in Gata2+/- embryos are unclear. Here, we investigated whether Gata2 haploinsufficiency selectively affects a cellular subset undergoing EHT. We show that Gata2+/- HSPCs initiate but cannot fully activate hematopoietic programming during EHT. In addition, due to reduced activity of the endothelial repressor Gfi1b, Gata2+/- HSPCs cannot repress the endothelial identity to complete maturation. Finally, we show that hematopoietic-specific induction of gfi1b can restore HSC production in gata2b-null (gata2b-/-) zebrafish embryos. This study illustrates pivotal roles of Gata2 on the regulation of transcriptional network governing HSPC identity throughout EHT.

The transcription factor GATA2 has pivotal roles in hematopoiesis. Germline GATA2 mutations in patients result in GATA2 haploinsufficiency syndrome characterized by immunodeficiency, bone marrow failure, and predispositions to myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). It is unclear how GATA2 haploinsufficiency drives abnormal hematopoiesis, and to date, these phenotypes have not been captured in a cell or animal model to study the mechanism behind GATA2 haploinsufficiency induced bone marrow failure and leukemia predisposition.