-Author name in bold denotes the presenting author
-Asterisk * with author name denotes a Non-ASH member
Clinically Relevant Abstract denotes an abstract that is clinically relevant.

PhD Trainee denotes that this is a recommended PHD Trainee Session.

Ticketed Session denotes that this is a ticketed session.

2687 Myelo-Erythroid Lineage Segregation Is Regulated By the GATA-2 Interactome

Program: Oral and Poster Abstracts
Session: 501. Hematopoietic Stem and Progenitor Cells and Hematopoiesis: Basic and Translational: Poster II
Hematology Disease Topics & Pathways:
Research, Fundamental Science
Sunday, December 10, 2023, 6:00 PM-8:00 PM

Alexandre Fagnan, PhD1, Roy Drissen, PhD2*, Cristina Di Genua3*, Yiran Meng4 and Claus Nerlov, PhD4*

1MRC - Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, ENG, United Kingdom
2MRC - Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, United Kingdom
3University of Oxford, Oxford, ENG, GBR
4Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, United Kingdom

Hematopoiesis arises from hematopoietic stem cell differentiating into downstream progenitors through a multistep process of sequential lineage potential restriction leading to the final production of >10 hematopoietic cell type.The regulation of hematopoietic lineage development involves synergistic and antagonistic interactions between transcription factors (TF) with lineage-restricted expression patterns, both through protein-protein interaction and direct transcriptional activation.Eosinophils, basophils and mast cells are key myeloid cell types involved in allergic inflammation and have been proposed to arise from a granulocyte-macrophage progenitor (GMP), that also generates neutrophils and macrophages, with GATA-2 up-regulation as a critical determinant. However, recent work identified a distinct basophil/eosinophil/mast cells progenitor (BEMP) that arises from multi-potent erythroid-primed multipotent progenitors (EMPP) where GATA-2 is already expressed (Drissen et al., 2016). Here, we show that the GATA-2 interactome, rather than GATA-2 expression, controls myeloid lineage choice in EMPPs.

To investigate the molecular mechanism instructing lineage decision in EMPP, we performed gene-expression profiling (RNA-seq) comparing its downstream progeny, early restricted erythroid/megakaryocyte progenitor (preMegE) and BEMP. This analysis identified differential expressed genes, including antagonist gene-expression pattern between the LIM-only protein 2 (Lmo2) and protein 4 (Lmo4), both previously shown as GATA-2 interactor. While Lmo2 function in hematopoiesis have been exhaustively studied, the role of Lmo4 remains unknown.

In colony forming unit (CFU)-assay, overexpression of Lmo4 in EMPP mediated by lentiviral delivery, resulted in an increased number of mast-colonies associated with a decreased of erythroid/megakaryocyte colonies, while the total colony number was unchanged. In addition, transplantation of hematopoietic lineage-negative cells transduced with Lmo4/empty(Ctrl) lentiviral particles, showed an increased frequency of basophils and eosinophils associated with a decrease of erythroid cells in Lmo4 overexpressing cells compared to Ctrl, in both bone marrow and spleen of recipient mice, 8 weeks after transplantation. Analysis of transduced Lineage-negative Kit+ (LK) compartment of recipient mice, also revealed an increased frequency of BEMP associated with a decreased frequency of preMegE in Lmo4overexpressing cells compared to Ctrl. Taken together, these data suggested that Lmo4 overexpression instruct lineage decision of EMPP toward basophil/eosinophil/mast lineages.Over-expression followed by co-immunoprecipitation in HEK-293T cell line revealed that LMO4 compete with LMO2 binding to GATA-2, suggesting a mutually exclusive transcriptional complex. More importantly, lack of interaction between LMO4 and GATA-2 mediated by a point mutation in the zinc-finger 1 domain (ZnF1) of GATA-2 (GATA2-G320D), frequently identified in AML/AEL, resulted in an absence of mast lineage potential of EMPP compared to GATA2+/+. Moreover, over-expression of Lmo4 in EMPP from GATA2G320D/G320D mice compared to GATA2+/+, failed to instruct myeloid fate in vitro.

Overall, our data identified a specific molecular mechanism of hematopoietic cell fate decision driven by protein-protein interaction with GATA-2. Perturbation of GATA-2 interactome through over-expression of the cofactor Lmo4 or specific point mutation in GATA2 ZnF1 domain results in lineage fate switch of EMPP toward the erythroid/megakaryocyte or basophil/eosinophil/mast lineages.

Disclosures: No relevant conflicts of interest to declare.

<< Previous Abstract | Next Abstract
*signifies non-member of ASH