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Publications (224)
TAZ::CAMTA1 is a fusion protein found in over 90% of Epithelioid Hemangioendothelioma (EHE), a rare vascular sarcoma with an unpredictable disease course. To date, how TAZ::CAMTA1 initiates tumour formation remains unexplained. To study the oncogenic mechanism leading to EHE initiation, we developed a model system whereby TAZ::CAMTA1 expression is...
During embryonic development, all blood progenitors are initially generated from endothelial cells that acquire a hemogenic potential. Blood progenitors emerge through an endothelial-to-hematopoietic transition regulated by the transcription factor RUNX1. To date, we still know very little about the molecular characteristics of hemogenic endotheliu...
During embryonic development, all blood progenitors are initially generated from endothelial cells that acquire a hemogenic potential. Blood progenitors emerge through an endothelial-to-hematopoietic transition regulated by the transcription factor RUNX1. To date, we still know very little about the molecular characteristics of hemogenic endotheliu...
The transcription factor RUNX1 is a master regulator of blood cell specification. During embryogenesis, hematopoietic progenitors are initially generated from hemogenic endothelium through an endothelium-to-hematopoietic transition controlled by RUNX1. Several studies have dissected the expression pattern and role of RUNX1 isoforms at the onset of...
In vitro generation and expansion of hematopoietic stem cells (HSCs) holds great promise for the treatment of any ailment that relies on bone marrow or blood transplantation. To achieve this, it is essential to resolve the molecular and cellular pathways that govern HSC formation in the embryo. HSCs first emerge in the aorta-gonad-mesonephros regio...
Despite long-lasting efforts to generate transplantable Haematopoietic Stem Cells (HSCs) from Embryonic Stem Cells and more recently, to reprogram skin/endothelial cells into HSCs, the results have been limited. The main pitfall is still our only limited understanding of HSC generation during development. The first HSCs trans differentiate from end...
Little is known about the emergence of blood progenitors during human embryogenesis due to ethical reasons and restricted embryo access. The use of human embryonic stem cells (hESCs) as a model system offers unique opportunities to dissect human blood cell formation. Here, we describe a protocol allowing the differentiation of hESCs via embryoid bo...
The transcription factor RUNX1 first came to prominence due to its involvement in the t(8;21) translocation in acute myeloid leukemia (AML). Since this discovery, RUNX1 has been shown to play important roles not only in leukemia but also in the ontogeny of the normal hematopoietic system. Although it is currently still challenging to fully assess t...
Gene expression governs cell fate, and is regulated via a complex interplay of transcription factors and molecules that change chromatin structure. Advances in sequencing-based assays have enabled investigation of these processes genome-wide, leading to large datasets that combine information on the dynamics of gene expression, transcription factor...
RUNX transcription factors orchestrate many different aspects of biology, including basic cellular and developmental processes, stem cell biology and tumorigenesis. In this Primer, we introduce the molecular hallmarks of the three mammalian RUNX genes, RUNX1, RUNX2 and RUNX3, and discuss the regulation of their activities and their mechanisms of ac...
The hematopoietic system is established during embryonic life through a series of developmental steps that culminates with the generation of hematopoietic stem cells. Characterisation of the transcriptional network that regulates blood cell emergence has led to the identification of transcription factors essential for this process. Among the many f...
The differentiation of human embryonic stem cells (hESCs) to hematopoietic lineages initiates with the specification of hemogenic endothelium, a transient specialized endothelial precursor of all blood cells. This in vitro system provides an invaluable model to dissect the emergence of hematopoiesis in humans. However, the study of hematopoiesis sp...
Haematopoietic stem cells (HSCs) are generated from haemogenic endothelial (HE) cells via the formation of intra-aortic haematopoietic clusters (IAHCs) in vertebrate embryos. The molecular events controlling endothelial specification, endothelial-to-haematopoietic transition (EHT) and IAHC formation, as it occurs in vivo inside the aorta, are still...
Erythropoiesis occurs through several waves during embryonic development. Although the source of the primitive wave is well characterized, the origin of erythrocytes later in embryogenesis is less clear due to overlaps between the different erythroid waves. Using the miR144/451-GFP mouse model to track cells expressing the erythroid microRNAs miR14...
The first hematopoietic stem and progenitor cells are generated during development from hemogenic endothelium (HE) through trans-differentiation. The molecular mechanisms underlying this endothelial-to-hematopoietic transition (EHT) remain poorly understood. Here, we explored the role of the epigenetic regulators HDAC1 and HDAC2 in the emergence of...
Video S4. Hdac1/2 Double Knockout Cells Undergo Improved EHT with SB43, Related to Figure 6
Video S2. Hdac1/2 Wild-Type Cultures Show Low Numbers of GFP-Positive Cells, Related to Figure 3
Video S3. Hdac1/2 Double Knockout Cells Undergo Improved EHT with SB43, Related to Figure 6
Video S5. Hdac1/2 Double Knockout Cells Undergo Improved EHT with SB43, Related to Figure 6
Video S1. Hdac1/2 Wild-Type Cultures Show Low Numbers of GFP-Positive Cells, Related to Figure 3
During ontogeny, hematopoietic stem and progenitor cells arise from hemogenic endothelium through an endothelial-to-hematopoietic transition that is strictly dependent on the transcription factor RUNX1. Although it is well established that RUNX1 is essential for the onset of hematopoiesis, little is known about the role of RUNX1 dosage specifically...
Movie S1a. Genesis of CD41+ Hematopoietic Cells, In Vitro, Promoted by HOXB4 (green: HOXB4, red: CD41)
Movie S2a. Endothelial-to-Hematopoietic Transition of HOXB4+ Hemogenic Endothelium (without Runx induction)
Movie S2b. Endothelial-to-Hematopoietic Transition of HOXB4+ Hemogenic Endothelium (after Runx1 induction)
Movie S1b. Genesis of CD41+ Hematopoietic Cells, In Vitro, Promoted by HOXB4 (phase contrast)
Generation of hematopoietic stem cells (HSCs) from pluripotent stem cells, in vitro, holds great promise for regenerative therapies. Primarily, this has been achieved in mouse cells by overexpression of the homeotic selector protein HOXB4. The exact cellular stage at which HOXB4 promotes hematopoietic development, in vitro, is not yet known. Howeve...
In recent years, highly detailed characterization of adult bone marrow (BM) myeloid progenitors has been achieved and, as a result, the impact of somatic defects on different hematopoietic lineage fate decisions can be precisely determined. Fetal liver (FL) hematopoietic progenitor cells (HPCs) are poorly characterized in comparison, potentially hi...
Characterization of Runx1-null progenitors (Relating to Fig 5).
A-B. Characterization of myeloid progenitor populations in wild type, Runx1-del/+ and Runx1-del/del E14.5 FL. A. Representative FACS plots of MEP, CMP, GMP and MkP populations. B. Quantitation of myeloid hematopoietic progenitors. N = 6.
C-F. Differential CFU-C activity of CD31- MEPs (...
Primers used for mouse genotyping PCRs.
(DOCX)
Details of flow cytometry reagents.
(DOCX)
Lineage output of Lin- Sca1high cKithigh (LSK), GMP and MkP Hematopoietic Stem and Progenitor Cells (Relating to Fig 1).
A. Differential CFU-C activity of E14.5 fetal liver cKit- cells, GMPs and MkPs. N = 3.
B-F. Flow cytometric analysis of wild type (WT) E14.5 fetal liver GMPs and MkPs. B. Representative FACS plot of CD55/CD150 expression in GMPs....
Lineage output of E12.5 and E13.5 Fetal Liver Runx1 P1-GFP::P2-hCD4 CMPs (Relating to Fig 3).
A-B. Differential CFU-C activity of E12.5 (A, N = 3) and E13.5 (B, N = 4) fetal liver wild type, P2-hCD4- and P2-hCD4+ CMPs.
C-F. Lineage output of day 7 OP9 co-cultured fetal liver wild type, P2-hCD4- and P2-hCD4+ CMPs. C-D. Representative FACS plots of C...
Differentially expressed genes in P2+ MEPs with respect to P2- MEPs.
(XLSX)
Characterization of CD31-/+ MEPs and CD31low/high CMPs (Relating to Fig 4).
A. Principal component analysis of the single cell RNA sequencing expression data from CD41- CD150+
P2-hCD4- and P2-hCD4+ MEPs and CMPs (P2- MEP, P2+ MEP, P2- CMP, P2+ CMP). Top: cells are color-coded according to the sorted population. Middle and Bottom plots: cells are co...
Characterization of Runx1-P1-MRIPV Fetal Liver progenitors (Relating to Fig 6).
A-B. Characterization of myeloid progenitor populations in wild type, Runx1-P1-MRIPV/+ and Runx1-P1-MRIPV/MRIPV E14.5 fetal liver. A. Representative FACS plots of MEP, CMP, GMP and MkP populations. B. Quantitation of myeloid hematopoietic progenitors. N = 7.
C-F. Differ...
Lineage output of E12.5 and E13.5 Fetal Liver Runx1-P1-GFP::P2-hCD4 MEPs (Relating to Fig 2).
A-B. Differential CFU-C activity of E12.5 (A, N = 3) and E13.5 (B, N = 4) fetal liver wild type, P2-hCD4- and P2-hCD4+ MEPs. C-F. Lineage output of day 7 OP9 co-cultured fetal liver wild type, P2-hCD4- and P2-hCD4+ MEPs.
C-D. Representative FACS plots of T...
Characterization of E16.5 Fetal Liver CMPs and MEPs (Relating to Fig 4).
A-B. Expression of Runx1-P1-GFP and P2-hCD4 in P1-GFP::P2-hCD4/+ E16.5 fetal liver CD41- CD150+ MEPs and CMPs. A. Representative FACS plots. B. Quantitation of the proportions of P1-
P2-, P1+
P2-, P1+
P2+ and P1-
P2+ MEPs and CMPs as a percentage of total live red blood cell l...
Impaired Mk/Ery Maturation in Runx1-null Fetal Liver (Relating to Fig 5).
A. RUNX1/ACTINB Western blot of total protein extract from wild type, Runx1-del/+ and Runx1-del/del E14.5 fetal liver. Representative of 3 independent experiments.
B-E. Characterization of erythroid lineage subsets S0-S5 in wild type, Runx1-del/+ and Runx1-del/del E14.5 fetal...
Mk/Ery maturation in Runx1-P1-MRIPV Fetal Liver (Relating to Fig 6).
A. RUNX1/ACTINB Western blot of total protein extract from wild type, Runx1-P1-MRIPV/+ and Runx1-P1-MRIPV/MRIPV E14.5 fetal liver. Representative of 3 independent experiments.
B-E. Characterization of erythroid lineage subsets S0-S5 in wild type, Runx1-P1-MRIPV/+ and Runx1-MRIPV/M...
Primers and probes used for qPCR.
(DOCX)
Differentially expressed genes in P2+ CMPs with respect to P2- CMPs.
(XLSX)
Antibody combinations used for flow cytometric analysis and sorting (see S2 Table for clone and supplier details).
(DOCX)
Primitive erythroblasts are the first blood cells generated during embryonic hematopoiesis. Tracking their emergence both in vivo and in vitro has remained challenging due to the lack of specific cell surface markers. To selectively investigate primitive erythropoiesis, we have engineered a new transgenic embryonic stem (ES) cell line, where eGFP e...
Sox7−/− embryos have defects in the dorsal aorta at E8.5. Whole mount PECAM1 staining of E8.5 Sox7+/− and Sox7−/− embryos. Arrows indicate posterior section of dorsal aorta.
RUNX1 is crucial for the regulation of megakaryocyte specification, maturation, and thrombopoiesis. Runx1 possesses 2 promoters: the distal P1 and proximal P2 promoters. The major protein isoforms generated by P1 and P2 are RUNX1C and RUNX1B, respectively, which differ solely in their N-terminal amino acid sequences. RUNX1C is the most abundantly e...
The transcriptional program that regulates the differentiation of endothelial precursor cells into a highly organized vascular network is still poorly understood. Here we explore the role of SOX7 during this process, performing a detailed analysis of the vascular defects resulting from either a complete deficiency in Sox7 expression or from the con...
Aberrant WNT signaling drives colorectal cancer (CRC). Here, we identify TIAM1 as a critical antagonist of CRC progression through inhibiting TAZ and YAP, effectors of WNT signaling. We demonstrate that TIAM1 shuttles between the cytoplasm and nucleus antagonizing TAZ/YAP by distinct mechanisms in the two compartments. In the cytoplasm, TIAM1 local...
RUNX transcription factors belong to a highly conserved class of transcriptional regulators which play various roles in the development of the majority of metazoans. In this review we focus on the founding member of the family, RUNX1, and its role in the transcriptional control of blood cell development in mammals. We summarize data showing that RU...
The types of progenitors generated during the successive stages of embryonic blood development are now fairly well characterized. The terminology used to describe these waves, however, can still be confusing. What is truly primitive? What is uniquely definitive? These questions become even more challenging to answer when blood progenitors are deriv...
Endothelial to hematopoietic transition (EHT) is a dynamic process involving the shutting down of endothelial gene expression and switching on of hematopoietic gene transcription. Whilst the factors regulating EHT in hemogenic endothelium (HE) of the dorsal aorta have been relatively well studied, the molecular regulation of yolk sac HE remains poo...
The transmission of extracellular signals into the nucleus involves inducible transcription factors, but how different signaling pathways act in a cell type-specific fashion is poorly understood. Here, we studied the regulatory role of the AP-1 transcription factor family in blood development using embryonic stem cell differentiation coupled with g...
Key Points
ENG regulatory elements target hemogenic mesoderm and hemogenic endothelium. Hemogenic progenitors can be enriched using these elements as molecular probes to discover novel regulators of hematopoiesis.
The first hematopoietic cells are generated very early in ontogeny to support the growth of the embryo and to provide the foundation to the adult hematopoietic system. There is a considerable therapeutic interest in understanding how these first blood cells are generated in order to try to reproduce this process in vitro. This would allow to genera...
: Hematopoietic cell-based therapies are currently available treatment options for many hematological and nonhematological disorders. However, the scarcity of allogeneic donor-derived cells is a major hurdle in treating these disorders. Embryonic stem cell-based directed differentiation and direct reprogramming of somatic cells provide excellent to...
B cell precursor acute lymphoblastic leukemia (BCP-ALL) is the most frequent type of cancer in children. Despite progresses in curative treatment, intensive chemotherapy regimens still cause life threatening complications. A better understanding of the molecular mechanisms underlying the emergence and maintenance of BCP-ALL is fundamental for the d...
Scientific Reports 6: Article number: 2591710.1038/srep25917; published online: May202016; updated: July042016
The original version of this Article contained a typographical error in the spelling of the author Maria Iliut which was incorrectly given as Maria Iluit. This has now been corrected in the PDF and HTML versions of the Article.
Well into the second decade of the 21(st) century, the field of regenerative medicine is bursting with hopes and promises to heal young and old. The bespoken generation of cells is thought to offer unprecedented cures for a vast range of diseases. Hematological disorders have already benefited tremendously from stem cell therapy in the form of bone...
During embryogenesis, the three SOXF transcription factors, SOX7, SOX17 and SOX18, regulate the specification of the cardiovascular system and are also involved in the development of haematopoiesis. The ectopic expression of SOX17 in both embryonic and adult blood cells enhances self-renewal. Likewise, the enforced expression of SOX7 during embryon...
Cardiovascular development during embryogenesis involves complex changes in gene regulatory networks regulated by a variety of transcription factors. In this review we discuss the various reported roles of the SOXF factors: SOX7, SOX17 and SOX18 in cardiac, vascular and lymphatic development. SOXF factors have pleiotropic roles during these process...
Movie S7. Time-Lapse Imaging of E8.5 FLK1+GFP+CD41− Sorted Cells on OP9 Monolayer Cultures Transduced with Control and Runx1 Lentivirus, Related to Figure 6
Movie S5. 3D Reconstruction of z Stacks of E7.5 Etv2::gfp Runx1b::rfp Embryos Stained for CD31, in Magenta; ETV2::GFP, in Green; and RUNX1b::RFP, in Red, Related to Figure 4
