- Email: [email protected]
Reconstruction of hierarchical differentiation processes based on the temporal clonal readout in distinct hematopoietic lineages
Poster Presentations/ Experimental Hematology 53 (2017) S54-S136
3164 - INVOLVEMENT OF THE VASCULAR NICHE IN THE REGULATION OF MURINE CHRONIC MYELOID LEUKEMIA Sonika Godavarthy1, Stefanie Herkt2, Eva Weissenberger1, Djamel Aggoune1, Kuan-Ting Pan3, Thomas Oellerich4, Vivian Oehler5, Richard Van Etten6, Joern Lausen7, and Daniela Krause1
S103
ing us to speculate about the plausibility of the particular assumptions. Our results highlight the necessity to understand the mechanisms of immune control in leukemia therapy to employ this effect for optimal treatment results and to furthermore identify the clinically accessible measures that allow to derive better predictions about the outcome of treatment cessation.
1
Georg-Speyer-Haus, Frankfurt, Germany; 2Institute for Transfusion Medicine DRKBlutspendedienst Baden-Wurttemburg-Hessen, Frankfurt, Germany; 3Bioanalytical mass spectrometry group, Max Planck Institute for Biophysical Chemistry, Frankfurt, Germany; 4Department of Internal Medicine, Haematology/Oncology, University Hospital Frankfurt, Frankfurt, Germany; 5Fred Hutchinson Cancer Research Centre, Seattle, WA, USA, Washington, United States; 6Fred Hutchinson Cancer Research Centre, Seattle, WA, USA, California, United States; 7Institute for Transfusion Medicine, DRK- Blutspendedienst Baden-W€urttemberg-Hessen, Frankfurt, Germany The endosteal bone marrow niche is known to protect leukemic stem cells (LSC) from chemotherapy in acute myeloid leukemia, but the knowledge about the role of the vascular niche in the bone marrow microenvironment in leukemia is limited. E-selectin, which is expressed on bone marrow endothelium, has been described to regulate the dormancy of normal hematopoietic stem cells. In chronic myeloid leukemia (CML) E-selectin and the adhesion molecule CD44, a ligand for E-selectin, which is overexpressed on CML-initiating cells, was shown to be responsible for homing and engraftment of LSC in a murine model of CML. However, exact mechanisms of the interactions between BCR-ABL1, the causative oncogene in CML, CD44 expression and interactions with the vascular niche are unclear. Hypothesizing that E-selectin influences the LSC cell cycle similar to HSC we treated murine recipients of CML-initiating cells in the retroviral transduction/transplantation model with the E -selectin inhibitor GMI-1271 and imatinib, considered standard of care in CML. This increased the survival of mice compared to animals treated with imatinib alone and decreased the engraftment of CML-initiating cells in this model, as well as in a xenotransplantation model of NOD SCID IL2 receptor knockout (NSG) mice injected with human CML cells. Further, treatment with GMI-1271 in vitro increased the cell cycle of LSC, with a concomitant increase in expression of the SCL/TAL1 gene, a transcriptional regulator. Tal1 was shown to negatively regulate CD44 expression, which was potentiated by imatinib. Downstream, imatinib reduced the AKTmediated phosphorylation of Tal1 (T90 and S122), which are known negative regulatory phosphorylation sites of Tal1 activity. In summary, inhibition of E-selectin in murine CML may lead to ‘non-adherence’ of LSC to the vascular niche and improved eradication by imatinib, as well as reduced engraftment of LSC, likely via an increase in cell cycle and an increase of Tal1 expression, which is shown to be regulated by BCR-ABL1. These data connect the role of CD44 for engraftment of LSC in CML with its regulation by Tal1 and BCR-ABL1 and the biology of the vascular niche in CML.
3165 - A MATHEMATICAL MODELING APPROACH TOWARDS IMMUNOLOGICAL CONTROL OF MINIMAL RESIDUAL DISEASE IN CML PATIENTS Ingmar Glauche1, Artur Fassoni2, Tom Haehnel3, Christoph Baldow3, and Ingo Roeder3
3166 - RECONSTRUCTION OF HIERARCHICAL DIFFERENTIATION PROCESSES BASED ON THE TEMPORAL CLONAL READOUT IN DISTINCT HEMATOPOIETIC LINEAGES Ingmar Glauche and Christoph Baldow
1 IMB, TU Dresden, Dresden, Germany; 2Universidade Federal de Itajuba, Itajuba, Brazil; 3TU Dresden, Dresden, Germany
IMB, TU Dresden, Dresden, Germany
There is increasing evidence in patients with Chronic Myeloid Leukemia (CML) pointing to the role of the immune system in the sustained control of residual leukemic cells after cessation of treatment with tyrosine-kinase inhibitors. It has been speculated that, once the treatment decreases the leukemic burden below some threshold, the immune cells are capable to keep it at a residual level, or even to completely eliminate the leukemic clone. However, at the moment these mechanisms are poorly understood. Specifically, since the immune control appears to be effective only at a low level of leukemic cells, it is important to understand the interaction between two phenomena: (i) the stimulation of immune cells by the presence of leukemic cells and (ii) the elimination of these cells. We develop a mathematical framework describing CML progression and treatment in terms of ordinary differential equations. Within this model we make different assumptions about the mechanisms by which (i) immune cells are stimulated by leukemic cells and (ii) how leukemic cells are targeted by immune cells. The combination of the different assumptions leads to several structurally different models, which are characterized by the existence of different numbers of steady states. We compare our conceptual results with available data sets from CML patients after TKI cessation, thereby allow-
The differentiation process of hematopoietic stem cells towards mature blood cells is generally depicted as a hierarchical decision process. Although the principle structure of such a branching tree is widely accepted, increasing evidence about complementary and alternative differentiation pathways question the concept of a fixed hierarchy. Clonal tracing studies allow quantifying to which extend and in which lineages marked hematopoietic stem cells contribute to the production of mature blood cells. However, the reconstruction of a hierarchical decision tree based on the temporal clonal readout in distinct blood lineages is computationally challenging. We use a computational approach to investigate how and under which conditions the original branching tree can be faithfully reconstructed. To this end we use a mathematically model to generate prototypic differentiation processes through a hierarchical decision tree and produce clonal readouts in different lineages over time that closely resample the data available from corresponding experimental and clinical studies. Applying a numerical optimization procedure, we estimate the extend of necessary data (e.g. with respect to measured compartments and timing of subsequent measures) that is needed for the reconstruction process in order to obtain identifiable results. We complement the model-based investigations with available data from clonal tracing studies and test our suggested approaches for reconstruction of
S104
Poster Presentations/ Experimental Hematology 53 (2017) S54-S136
hierarchical decision trees in real-world settings. Our analysis is a necessary and complementary prerequisite to support experimental approaches for the reconstruction of hierarchical, hematopoietic decision trees. In particular, the modeling approach allows, based on the available data, to estimate whether certain decision processes can be uniquely reconstructed or not.
3168 - TRACING THE EMERGENCE OF HEMATOPOIETIC STEM CELLS FROM THE EXTRA-EMBRYONIC YOLK SAC Yasamine Ghorbanian1, Lydia Lee2, Hanna Mikkola2, and Matthew Inlay1 1
University of California, Irvine, Irvine, United States; 2University of California, Los Angeles, Los Angeles, United States While iPSCs offer the potential for an unlimited source of patient HSCs, the generation of transplantable HSCs from iPSCs has yet to be achieved. This suggests that a better understanding of how HSCs arise naturally in the course of embryonic development is needed. The first primitive waves of hematopoiesis arise from the embryonic yolk sac (YS), but definitive waves arise later and produce the first HSCs. Which tissues produce definitive hematopoiesis, and thereby HSCs, remains unclear. The aorta-gonad-mesonephros (AGM), placenta, YS, and vitelline vessels are all potential sources. Using a Lyve1-Cre lineage tracing reporter, we previously published with Dr. Hanna Mikkola’s lab that Lyve1-Cre marks definitive hematopoietic cells that arise primarily in the YS, with some labeling in the vitelline vessels, and show that about one-third of adult HSCs are derived from Lyve1-expressing precursors. Our central hypothesis is that the extra-embryonic YS is a significant source of adult HSCs. We have found that Lyve1-derived cells contribute to each definitive wave of embryonic hematopoiesis. Lyve1-marked pre-HSCs from the early embryo are transplantable and give rise to adult HSCs. Lyve1-derived adult HSCs contribute to cells in all of the major blood lineages. Interestingly, there seems to be a bias in the development of lymphoid cells, suggesting that Lyve1-derived HSCs have functional properties distinct from HSCs that arise from other tissues. Taken together, this data suggests that Lyve1-derived cells give rise to engraftable, multipotent, and long term self-renewing HSCs. We interpret these results to support the notion that the extra-embryonic YS is an HSC nice in the embryo and contributes to adult hematopoiesis.
3167 - THE ROLE OF THE GATA2 TRANSCRIPTIONAL PROGRAM IN THE DEVELOPMENT OF ACUTE MYELOID LEUKEMIA Emanuele Gioacchino1, Emma de Pater2, Ivo Touw1, Hans de Looper1, Paulette Strien1, Remco Hoogenboezem1, Eric Bindels1, Elaine Dzierzak3, and Polynikis Kaimakis4 Erasmus MC, Rotterdam (NL), The Netherlands; 2Erasmus MC, Rotterdam, Netherlands; 3Edinburgh Medical School, Edinburgh, United Kingdom; 4Centro de Biologıa Molecular Severo Ochoa, Madrid, Spain
3169 - THE ROLE OF FUBP1 IN HEMATOPOIETIC STEM CELLS AND DURING ERYTHROID MATURATION Katharina Gerlach1, Josephine Wesely2, Marlene Steiner1, Michael Rieger3, and Martin Z€ornig1 1
Georg-Speyer-Haus, Frankfurt, Germany; 2Icahn School of Medicine at Mount Sinai, New York, United States; 3University Hospital Frankfurt, Frankfurt, Germany
1
Myeloid dysplastic syndrome (MDS) and acute myeloid leukemia (AML) are aggressive forms of blood cancer that are still fatal in more than 50% of patients. Familial forms of MDS/AML have very poor cure rates. Germline mutations in GATA2, resulting in haploinsufficiency of the transcription factor GATA2 (Gata2 in animals) is the most commonly mutated gene in childhood MDS/AML. Between 36-71% of patients with germline mutations in GATA2 develop AML. Although the disease initiating mutation is known, it is unclear how GATA2 haploinsufficiency predisposes to leukemia and contributes to the progression to MDS/AML. Because GATA2 is pivotal for hematopoietic stem cell (HSC) generation in the embryo, the diseasedriving defects likely already affect the hematopoietic system during fetal development. Therefore, we hypothesize that the defect in the hematopoietic system of patients with a GATA2 haploinsufficiency in newly generated HSCs or in amplifying HSCs contributes to leukemia predisposition. We have performed RNA sequencing on WT and Gata2+/- mouse embryonic and adult phenotypic HSCs and found that Gata2 transcriptional levels normalize during HSC development. Despite this normalization, the transcriptome of WT and Gata2+/- adult bone marrow HSCs is still transcriptionally different. This points to an underlying epigenetic reprogramming during the embryonic stages. We have found that cell cycle and metabolic signatures are different in embryonic HSCs compared to WT. We will further dissect these phenotypes in the zebrafish model in which we have generated a knockout of gata2b and several reporter lines to study these processes in vivo.
We previously found out that the transcriptional regulator and established activator of c-myc gene transcription, FUSE-binding protein 1 (FUBP1), functions as an important oncoprotein in hepatocellular carcinoma (HCC). In addition, two different gene trap mouse models with impaired FUBP1 function and prospectively isolated murine Fubp1 knockdown hematopoietic stem cells (HSCs) were analyzed to unravel the physiological function of FUBP1. The results of serial competitive transplantation experiments, ex vivo single cell tracking and quantitative real-time PCR analyses led us to conclude that FUBP1 fulfills an essential role in LT-HSC self-renewal by promoting survival and proliferation of HSCs via the regulation of gene expression. We then addressed the question whether FUBP1 also influences the differentiation capacity of hematopoietic progenitor cells. Indeed, flow cytometric analyses of FUBP1deficient fetal liver cells and Fubp1 knockout embryonic stem cell clones revealed an impaired terminal erythroid maturation. This finding corresponds with the severe anemia observed in mouse embryos lacking functional FUBP1 and might explain their lethality at day E15.5 of embryonic development. Our findings suggest at least two functions for FUBP1 in the hematopoietic system as an HSC self-renewal factor and as a transcriptional regulator of erythroid maturation. We continue to analyze the regulatory transcriptional network that is controlled by FUBP1 in HSCs and during erythroid maturation. The ongoing establishment of a conditional Fubp1 knockout mouse line will provide a valuable tool for our current studies.
3164 - INVOLVEMENT OF THE VASCULAR NICHE IN THE REGULATION OF MURINE CHRONIC MYELOID LEUKEMIA Sonika Godavarthy1, Stefanie Herkt2, Eva Weissenberger1, Djamel Aggoune1, Kuan-Ting Pan3, Thomas Oellerich4, Vivian Oehler5, Richard Van Etten6, Joern Lausen7, and Daniela Krause1
S103
ing us to speculate about the plausibility of the particular assumptions. Our results highlight the necessity to understand the mechanisms of immune control in leukemia therapy to employ this effect for optimal treatment results and to furthermore identify the clinically accessible measures that allow to derive better predictions about the outcome of treatment cessation.
1
Georg-Speyer-Haus, Frankfurt, Germany; 2Institute for Transfusion Medicine DRKBlutspendedienst Baden-Wurttemburg-Hessen, Frankfurt, Germany; 3Bioanalytical mass spectrometry group, Max Planck Institute for Biophysical Chemistry, Frankfurt, Germany; 4Department of Internal Medicine, Haematology/Oncology, University Hospital Frankfurt, Frankfurt, Germany; 5Fred Hutchinson Cancer Research Centre, Seattle, WA, USA, Washington, United States; 6Fred Hutchinson Cancer Research Centre, Seattle, WA, USA, California, United States; 7Institute for Transfusion Medicine, DRK- Blutspendedienst Baden-W€urttemberg-Hessen, Frankfurt, Germany The endosteal bone marrow niche is known to protect leukemic stem cells (LSC) from chemotherapy in acute myeloid leukemia, but the knowledge about the role of the vascular niche in the bone marrow microenvironment in leukemia is limited. E-selectin, which is expressed on bone marrow endothelium, has been described to regulate the dormancy of normal hematopoietic stem cells. In chronic myeloid leukemia (CML) E-selectin and the adhesion molecule CD44, a ligand for E-selectin, which is overexpressed on CML-initiating cells, was shown to be responsible for homing and engraftment of LSC in a murine model of CML. However, exact mechanisms of the interactions between BCR-ABL1, the causative oncogene in CML, CD44 expression and interactions with the vascular niche are unclear. Hypothesizing that E-selectin influences the LSC cell cycle similar to HSC we treated murine recipients of CML-initiating cells in the retroviral transduction/transplantation model with the E -selectin inhibitor GMI-1271 and imatinib, considered standard of care in CML. This increased the survival of mice compared to animals treated with imatinib alone and decreased the engraftment of CML-initiating cells in this model, as well as in a xenotransplantation model of NOD SCID IL2 receptor knockout (NSG) mice injected with human CML cells. Further, treatment with GMI-1271 in vitro increased the cell cycle of LSC, with a concomitant increase in expression of the SCL/TAL1 gene, a transcriptional regulator. Tal1 was shown to negatively regulate CD44 expression, which was potentiated by imatinib. Downstream, imatinib reduced the AKTmediated phosphorylation of Tal1 (T90 and S122), which are known negative regulatory phosphorylation sites of Tal1 activity. In summary, inhibition of E-selectin in murine CML may lead to ‘non-adherence’ of LSC to the vascular niche and improved eradication by imatinib, as well as reduced engraftment of LSC, likely via an increase in cell cycle and an increase of Tal1 expression, which is shown to be regulated by BCR-ABL1. These data connect the role of CD44 for engraftment of LSC in CML with its regulation by Tal1 and BCR-ABL1 and the biology of the vascular niche in CML.
3165 - A MATHEMATICAL MODELING APPROACH TOWARDS IMMUNOLOGICAL CONTROL OF MINIMAL RESIDUAL DISEASE IN CML PATIENTS Ingmar Glauche1, Artur Fassoni2, Tom Haehnel3, Christoph Baldow3, and Ingo Roeder3
3166 - RECONSTRUCTION OF HIERARCHICAL DIFFERENTIATION PROCESSES BASED ON THE TEMPORAL CLONAL READOUT IN DISTINCT HEMATOPOIETIC LINEAGES Ingmar Glauche and Christoph Baldow
1 IMB, TU Dresden, Dresden, Germany; 2Universidade Federal de Itajuba, Itajuba, Brazil; 3TU Dresden, Dresden, Germany
IMB, TU Dresden, Dresden, Germany
There is increasing evidence in patients with Chronic Myeloid Leukemia (CML) pointing to the role of the immune system in the sustained control of residual leukemic cells after cessation of treatment with tyrosine-kinase inhibitors. It has been speculated that, once the treatment decreases the leukemic burden below some threshold, the immune cells are capable to keep it at a residual level, or even to completely eliminate the leukemic clone. However, at the moment these mechanisms are poorly understood. Specifically, since the immune control appears to be effective only at a low level of leukemic cells, it is important to understand the interaction between two phenomena: (i) the stimulation of immune cells by the presence of leukemic cells and (ii) the elimination of these cells. We develop a mathematical framework describing CML progression and treatment in terms of ordinary differential equations. Within this model we make different assumptions about the mechanisms by which (i) immune cells are stimulated by leukemic cells and (ii) how leukemic cells are targeted by immune cells. The combination of the different assumptions leads to several structurally different models, which are characterized by the existence of different numbers of steady states. We compare our conceptual results with available data sets from CML patients after TKI cessation, thereby allow-
The differentiation process of hematopoietic stem cells towards mature blood cells is generally depicted as a hierarchical decision process. Although the principle structure of such a branching tree is widely accepted, increasing evidence about complementary and alternative differentiation pathways question the concept of a fixed hierarchy. Clonal tracing studies allow quantifying to which extend and in which lineages marked hematopoietic stem cells contribute to the production of mature blood cells. However, the reconstruction of a hierarchical decision tree based on the temporal clonal readout in distinct blood lineages is computationally challenging. We use a computational approach to investigate how and under which conditions the original branching tree can be faithfully reconstructed. To this end we use a mathematically model to generate prototypic differentiation processes through a hierarchical decision tree and produce clonal readouts in different lineages over time that closely resample the data available from corresponding experimental and clinical studies. Applying a numerical optimization procedure, we estimate the extend of necessary data (e.g. with respect to measured compartments and timing of subsequent measures) that is needed for the reconstruction process in order to obtain identifiable results. We complement the model-based investigations with available data from clonal tracing studies and test our suggested approaches for reconstruction of
S104
Poster Presentations/ Experimental Hematology 53 (2017) S54-S136
hierarchical decision trees in real-world settings. Our analysis is a necessary and complementary prerequisite to support experimental approaches for the reconstruction of hierarchical, hematopoietic decision trees. In particular, the modeling approach allows, based on the available data, to estimate whether certain decision processes can be uniquely reconstructed or not.
3168 - TRACING THE EMERGENCE OF HEMATOPOIETIC STEM CELLS FROM THE EXTRA-EMBRYONIC YOLK SAC Yasamine Ghorbanian1, Lydia Lee2, Hanna Mikkola2, and Matthew Inlay1 1
University of California, Irvine, Irvine, United States; 2University of California, Los Angeles, Los Angeles, United States While iPSCs offer the potential for an unlimited source of patient HSCs, the generation of transplantable HSCs from iPSCs has yet to be achieved. This suggests that a better understanding of how HSCs arise naturally in the course of embryonic development is needed. The first primitive waves of hematopoiesis arise from the embryonic yolk sac (YS), but definitive waves arise later and produce the first HSCs. Which tissues produce definitive hematopoiesis, and thereby HSCs, remains unclear. The aorta-gonad-mesonephros (AGM), placenta, YS, and vitelline vessels are all potential sources. Using a Lyve1-Cre lineage tracing reporter, we previously published with Dr. Hanna Mikkola’s lab that Lyve1-Cre marks definitive hematopoietic cells that arise primarily in the YS, with some labeling in the vitelline vessels, and show that about one-third of adult HSCs are derived from Lyve1-expressing precursors. Our central hypothesis is that the extra-embryonic YS is a significant source of adult HSCs. We have found that Lyve1-derived cells contribute to each definitive wave of embryonic hematopoiesis. Lyve1-marked pre-HSCs from the early embryo are transplantable and give rise to adult HSCs. Lyve1-derived adult HSCs contribute to cells in all of the major blood lineages. Interestingly, there seems to be a bias in the development of lymphoid cells, suggesting that Lyve1-derived HSCs have functional properties distinct from HSCs that arise from other tissues. Taken together, this data suggests that Lyve1-derived cells give rise to engraftable, multipotent, and long term self-renewing HSCs. We interpret these results to support the notion that the extra-embryonic YS is an HSC nice in the embryo and contributes to adult hematopoiesis.
3167 - THE ROLE OF THE GATA2 TRANSCRIPTIONAL PROGRAM IN THE DEVELOPMENT OF ACUTE MYELOID LEUKEMIA Emanuele Gioacchino1, Emma de Pater2, Ivo Touw1, Hans de Looper1, Paulette Strien1, Remco Hoogenboezem1, Eric Bindels1, Elaine Dzierzak3, and Polynikis Kaimakis4 Erasmus MC, Rotterdam (NL), The Netherlands; 2Erasmus MC, Rotterdam, Netherlands; 3Edinburgh Medical School, Edinburgh, United Kingdom; 4Centro de Biologıa Molecular Severo Ochoa, Madrid, Spain
3169 - THE ROLE OF FUBP1 IN HEMATOPOIETIC STEM CELLS AND DURING ERYTHROID MATURATION Katharina Gerlach1, Josephine Wesely2, Marlene Steiner1, Michael Rieger3, and Martin Z€ornig1 1
Georg-Speyer-Haus, Frankfurt, Germany; 2Icahn School of Medicine at Mount Sinai, New York, United States; 3University Hospital Frankfurt, Frankfurt, Germany
1
Myeloid dysplastic syndrome (MDS) and acute myeloid leukemia (AML) are aggressive forms of blood cancer that are still fatal in more than 50% of patients. Familial forms of MDS/AML have very poor cure rates. Germline mutations in GATA2, resulting in haploinsufficiency of the transcription factor GATA2 (Gata2 in animals) is the most commonly mutated gene in childhood MDS/AML. Between 36-71% of patients with germline mutations in GATA2 develop AML. Although the disease initiating mutation is known, it is unclear how GATA2 haploinsufficiency predisposes to leukemia and contributes to the progression to MDS/AML. Because GATA2 is pivotal for hematopoietic stem cell (HSC) generation in the embryo, the diseasedriving defects likely already affect the hematopoietic system during fetal development. Therefore, we hypothesize that the defect in the hematopoietic system of patients with a GATA2 haploinsufficiency in newly generated HSCs or in amplifying HSCs contributes to leukemia predisposition. We have performed RNA sequencing on WT and Gata2+/- mouse embryonic and adult phenotypic HSCs and found that Gata2 transcriptional levels normalize during HSC development. Despite this normalization, the transcriptome of WT and Gata2+/- adult bone marrow HSCs is still transcriptionally different. This points to an underlying epigenetic reprogramming during the embryonic stages. We have found that cell cycle and metabolic signatures are different in embryonic HSCs compared to WT. We will further dissect these phenotypes in the zebrafish model in which we have generated a knockout of gata2b and several reporter lines to study these processes in vivo.
We previously found out that the transcriptional regulator and established activator of c-myc gene transcription, FUSE-binding protein 1 (FUBP1), functions as an important oncoprotein in hepatocellular carcinoma (HCC). In addition, two different gene trap mouse models with impaired FUBP1 function and prospectively isolated murine Fubp1 knockdown hematopoietic stem cells (HSCs) were analyzed to unravel the physiological function of FUBP1. The results of serial competitive transplantation experiments, ex vivo single cell tracking and quantitative real-time PCR analyses led us to conclude that FUBP1 fulfills an essential role in LT-HSC self-renewal by promoting survival and proliferation of HSCs via the regulation of gene expression. We then addressed the question whether FUBP1 also influences the differentiation capacity of hematopoietic progenitor cells. Indeed, flow cytometric analyses of FUBP1deficient fetal liver cells and Fubp1 knockout embryonic stem cell clones revealed an impaired terminal erythroid maturation. This finding corresponds with the severe anemia observed in mouse embryos lacking functional FUBP1 and might explain their lethality at day E15.5 of embryonic development. Our findings suggest at least two functions for FUBP1 in the hematopoietic system as an HSC self-renewal factor and as a transcriptional regulator of erythroid maturation. We continue to analyze the regulatory transcriptional network that is controlled by FUBP1 in HSCs and during erythroid maturation. The ongoing establishment of a conditional Fubp1 knockout mouse line will provide a valuable tool for our current studies.