Lyn kinase plays important roles in erythroid expansion, maturation and erythropoietin receptor signalling by regulating inhibitory signalling pathways that control survival

2014 ◽  
Vol 459 (3) ◽  
pp. 455-466 ◽  
Author(s):  
Neli S. Slavova-Azmanova ◽  
Nicole Kucera ◽  
Alison Louw ◽  
Jiulia Satiaputra ◽  
Adley Handoko ◽  
...  
Keyword(s):  
Cell Survival ◽  
Cell Development ◽  
Erythropoietin Receptor ◽  
Erythroid Cell ◽  
Signalling Pathways ◽  
Erythroid Cells ◽  
Src Family Kinase ◽  
Receptor Signalling ◽  
Lyn Kinase ◽  
Control Survival

In erythroid cells both positive viability signals and feedback inhibitory signalling require the Src family kinase Lyn, influencing cell survival and their ability to differentiate. This illustrates that Lyn is critical for normal erythropoiesis and erythroid cell development.

Sox6 cell-autonomously stimulates erythroid cell survival, proliferation, and terminal maturation and is thereby an important enhancer of definitive erythropoiesis during mouse development

Blood ◽  
2006 ◽  
Vol 108 (4) ◽  
pp. 1198-1207 ◽  
Author(s):  
Bogdan Dumitriu ◽  
Michael R. Patrick ◽  
Jane P. Petschek ◽  
Srujana Cherukuri ◽  
Ursula Klingmuller ◽  
...  
Keyword(s):  
Cell Survival ◽  
Cell Fate ◽  
Fetal Liver ◽  
Cell Development ◽  
Erythroid Cell ◽  
Erythroid Cells ◽  
Mouse Development ◽  
Hematopoietic Stem ◽  
Definitive Erythropoiesis ◽  
Erythrocyte Cytoskeleton

Abstract Erythropoiesis, the essential process of hematopoietic stem cell development into erythrocytes, is controlled by lineage-specific transcription factors that determine cell fate and differentiation and by the hormone erythropoietin that stimulates cell survival and proliferation. Here we identify the Sry-related high-mobility-group (HMG) box transcription factor Sox6 as an important enhancer of definitive erythropoiesis. Sox6 is highly expressed in proerythroblasts and erythroblasts in the fetal liver, neonatal spleen, and bone marrow. Mouse fetuses and pups lacking Sox6 develop erythroid cells slowly and feature misshapen, short-lived erythrocytes. They compensate for anemia by elevating the serum level of erythropoietin and progressively enlarging their erythropoietic tissues. Erythroid-specific inactivation of Sox6 causes the same phenotype, demonstrating cell-autonomous roles for Sox6 in erythroid cells. Sox6 potentiates the ability of erythropoietin signaling to promote proerythroblast survival and has an effect additive to that of erythropoietin in stimulating proerythroblast and erythroblast proliferation. Sox6 also critically facilitates erythroblast and reticulocyte maturation, including hemoglobinization, cell condensation, and enucleation, and ensures erythrocyte cytoskeleton long-term stability. It does not control adult globin and erythrocyte cytoskeleton genes but acts by stabilizing filamentous actin (F-actin) levels. Sox6 thus enhances erythroid cell development at multiple levels and thereby ensures adequate production and quality of red blood cells.

The SCL Transcription Factor Supports Erythroid Cell Survival and Differentiation Downstream of the Erythropoietin Receptor.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 818-818
Author(s):  
Rachid Lahlil ◽  
Richard Martin ◽  
Peter D. Aplan ◽  
C. Glenn Begley ◽  
Jacqueline E. Damen ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Cell Survival ◽  
Cell Fate ◽  
Genetic Interaction ◽  
Fetal Liver ◽  
Erythropoietin Receptor ◽  
Erythroid Cell ◽  
Loss Of Function

Abstract Erythroid cell development critically depends on the SCL/Tal1 transcription factor and on erythropoietin signalling. In the present study, we have taken several approaches to show that the two genes operate within the same pathway to consolidate the erythroid lineage. Signaling through the erythropoietin receptor (EpoR) upregulates SCL protein levels in a clonal cell line (TF-1) in vitro, and in murine fetal liver cells in vivo, when Epor−/− cells were compared to those of wild type littermates at E12.5. In addition, we provide functional evidence for a linear pathway from EpoR to SCL that regulates erythropoiesis. Interfering with SCL induction or SCL function prevents the anti-apoptotic effect of Epo in TF-1 cells and conversely, ectopic SCL expression is sufficient to substitute for Epo to transiently maintain cell survival. In vivo, SCL gain of function complements the cellular defects in Epor−/− embryos to support cell survival and maturation during primitive and definitive erythropoiesis, as assessed by cellular and histological analyses of Epor−/− SCLtg embryos. Moreover, several erythroid specific genes that are decreased in Epor−/− embryos are rescued by the SCL transgene including glycophorinA, bH1 and bmaj globin, providing molecular confirmation of the functional and genetic interaction between Epor and SCL. Conversely, erythropoiesis becomes deficient in compound Epor+/−SCL+/− heterozygote mice, indicating that the genetic interaction between EpoR and SCL is synthetic. Finally, using EpoR mutants that harbour well defined signalling deficiencies, combined with gain and loss of function approaches for specific kinases, we identify MAPK as the major signal transduction pathway downstream of EpoR that upregulates SCL function, necessary for erythroid cell survival and differentiation. Taken together, our observations are consistent with the view that cytokines can influence cell fate by altering the dosage of lineage transcriptional regulators.

Erythropoietin receptor-dependent erythroid colony-forming unit development: capacities of Y343 and phosphotyrosine-null receptor forms

Blood ◽  
2002 ◽  
Vol 99 (3) ◽  
pp. 898-904
Author(s):  
Chris P. Miller ◽  
Destin W. Heilman ◽  
Don M. Wojchowski
Keyword(s):  
Growth Factor Receptor ◽  
Cell Development ◽  
Erythropoietin Receptor ◽  
Janus Kinase ◽  
Erythroid Cell ◽  
Compensatory Mechanisms ◽  
Erythroid Progenitor ◽  
Epo Receptor ◽  
Binding Domains ◽  
Progenitor Cell Proliferation

Red cell development depends on the binding of erythropoietin (EPO) to receptors expressed by erythroid colony-forming units (CFUe) and the subsequent activation of receptor-bound Janus kinase (Jak2). Jak2 then mediates the phosphorylation of receptor tyrosine sites and the recruitment of 25 or more Src homology 2 domain-encoding proteins and associated factors. Previous studies have shown that an EPO receptor form containing Jak2-binding domains plus a single phosphotyrosine343 (PY343)–STAT5-binding site provides all signals needed for erythroid cell development. However, roles for PY343 and STAT5 remain controversial, and findings regarding PY-null receptor activities and erythropoiesis in STAT5-deficient mice are disparate. To study activities of a PY-null EPO receptor in primary cells while avoiding compensatory mechanisms, a form retaining domains for Jak2 binding and activation, but lacking all cytoplasmic tyrosine sites, was expressed in transgenic mice from aGATA1 gene-derived vector as a human epidermal growth factor receptor- murine EPO receptor chimera (EE-T-Y343F). The bio-signaling capacities of this receptor form were investigated in CFUe from thiamphenicol-treated mice. Interestingly, this PY-null EPO receptor form supported CFUe development (in the absence of detectable STAT5 activation) at efficiencies within 3-fold of those levels mediated by either an EE-T-Y343 form or the endogenous EPO receptor. However, EE-T-Y343F–dependent Ter119+ erythroblast maturation was attenuated. In tests of cosignaling with c-Kit, EE-T-Y343F nonetheless retained full capacity to synergize with c-Kit in promoting erythroid progenitor cell proliferation. Thus, EPO receptor PY-dependent events can assist late erythropoiesis but may be nonessential for EPO receptor–c-Kit synergy.

Csnk2β Knockout during Hematopoiesis Results in Lethality at Mid/Late Gestation Mostly Due to Impaired Fetal Erythropoiesis

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4329-4329
Author(s):  
Laura Quotti Tubi ◽  
Sara Canovas Nunes ◽  
Alessandro Casellato ◽  
Elisa Mandato ◽  
Fortunato Zaffino ◽  
...  
Keyword(s):  
Transcription Factors ◽  
High Throughput ◽  
Fetal Liver ◽  
Cell Development ◽  
Conditional Knockout ◽  
Late Gestation ◽  
Cell Maturation ◽  
Erythroid Cell ◽  
Erythroid Cells ◽  
Fetal Erythropoiesis

Abstract Background. CK2, a serine-threonine kinase composed of two catalytic (α) and two regulatory (β) subunits, has been clearly involved in several hematologic malignancies. This kinase regulates the PTEN/PI3K/AKT, Wnt/βcatenin, Hedgehog, JAK-STAT, cMyc and NF-κB signalling cascades, all of which are known to be of critical importance for hematopoietic stem cell (HSC) biology and normal hematopoiesis. However, the role played by CK2 during blood cell development has remained as yet unexplored. Aims and methods. CK2 function in hematopoiesis was investigated generating conditional knockout mice for CK2β by crossing Csnk2β-Flox/Flox mice with Vav1-CRE transgenic mice. Inactivation of Csnk2β started from 9.5 dpc during embryonic development. Histo-cytological methods, FACS analysis, colony-forming assays (CFA), signal transduction studies by western blotting and RT-PCR were employed to characterize the cellular and molecular phenotype. High throughput RNAseq analysis was also performed on purified Ter119-positive erythroid cells from Csnk2β knockout and Csnk2β control mice to identify differentially expressed CK2-dependent transcriptional targets. Results. Csnk2β knockout in hematopoiesis resulted lethal at mid-late gestation. Rarely some pups were found dead at birth. Macroscopic and phenotypic analysis during gestation revealed the appearance of pale and hydropic fetuses after 12.5 dpc. The majority of pups showed teleangiectasic vessels and haemorrhages. Fetal livers appeared smaller and paler. Cytological analysis and CFA studies unveiled a great depletion of hematopoietic elements belonging to both the erythroid, megakaryocytic and granulocytic-monocytic precursors. A more thorough analysis of the erythroid phenotype revealed that Csnk2β loss caused impairment/loss of red cell maturation at two developmental stages: the earlier stages of Megakaryocyte-Erythroid Precursors (MEP) and pro-erythroblasts and the later stages of terminal maturation (orthocromatic erythroblasts towards reticulocytes). Expression analysis of proteins/genes belonging to known hematopoietic and erythroid-regulating pathways showed perturbations in cell cycle regulatory molecules, cellular apoptosis, a marked reduction of total and phosphorylated Akt in Ser473 and Ser129, a decrease of GATA1 protein levels and a decrease of Hedgehog/Wnt target genes such as Gli-1 and Cyclin D1. Erythropoietin-dependent AKT activation and GATA1 phosphorylation was impaired by Csnk2β loss. Moreover, starting at 14.5 dpc, blood cells displayed a massive p53-dependent response, marked by high levels of p21 and a progressive clear apopototic phenotype. At 17.5 dpc residual hematopoietic cells in the fetal liver were represented by dying erythroid cells, immature myelo-monocytic precursors (expressing high CD11b and low Gr1 levels on the surface) and B-cells displaying an aberrant phenotype with low intensity of expression of B220 and CD19 on the surface. High throughput RNAseq analysis of Ter119-expressing fetal liver cells (erythroid lineage) obtained from 14.5 dpc pups revealed the upregulation of 145 transcripts and the downregulation of 68 transcripts. Among the most increased transcripts were the transcription factors Jun/AP1 and stress-related intermediaries and embryonal globin ε and ζ chains. Among the most decreased transcripts were sugar transporters, glycoproteins CD36 and CD59a, Duffy Blood Group Atypical Chemokine Receptor and component members. Conclusions. We found that Csnk2β plays a critical role in mouse blood development by regulating definitive hematopoiesis of all the hematopoietic cell lineages; however, Csnk2β was needed for the early and late erythropoiesis whilst its loss could be compatible with a certain extent of immature/altered myelo-monocytic and B cell development. Among the pathways found targeted by Csnk2β loss were the PI3K/Akt and the p53-p21 cascades. Our data also suggest that Csnk2β might have a role in the proper activation of the erythroid master regulator GATA1. Moreover, RNAseq analysis revealed that this kinase might have a broader impact during erythroid cell maturation by regulating the activity of critical stress related transcription factors, of molecules regulating energy-managing cellular processes and of mechanisms controlling the switch from embryonal to fetal erythropoiesis. Disclosures No relevant conflicts of interest to declare.

ETV6-RUNX1 (TEL-AML1) Deregulates the Erythropoietin Receptor (EPOR) and Promotes Cell Survival.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3641-3641
Author(s):  
Veronica Torrano Moya ◽  
Penny Cardus ◽  
Julia Procter ◽  
Mel Greaves ◽  
Anthony Ford
Keyword(s):  
Cell Surface ◽  
Cord Blood ◽  
Cell Survival ◽  
Blood Cells ◽  
Genetic Alterations ◽  
Erythropoietin Receptor ◽  
Luciferase Reporter ◽  
Erythroid Cell ◽  
Epo Receptor ◽  
Cord Blood Cells

Abstract Abstract 3641 ETV6-RUNX1 (TEL-AML1) fusion is usually an initiating and pre-natal event in childhood acute lymphoblastic leukaemia (ALL). Transformation results in the generation of a persistent pre-leukemic clone, which post-natally converts to ALL following the acquisition of necessary secondary genetic alterations. The erythropoietin receptor gene (EPOR) is consistently highly expressed ectopically in TEL-AML1+ ALL but the presence of a functional receptor on the cell surface and its role, if any, in leukemogenesis driven by TEL-AML1 remains to be confirmed. Similarly, there is much debate as to the role played by the cytokine erythropoietin (EPO) and signaling through EPOR in relation to non-erythroid cell survival. Here, we show by using biotinylated erythropoietin (EPO) and flow analysis that the pre-B ALL TEL-AML1+ cell line (REH) appeared to have higher levels of EPO receptor than other non-TEL-AML1 cell lines. Furthermore, when we “blind screened” CD19+ peripheral blood or bone marrow cells from 10 patients with pre-B ALL, we identified 5 patients that showed high expression of ligand-binding EPO receptor on the cell surface, 4 of which were subsequently shown to be TEL-AML1+. We show that the inducible expression of TEL-AML1 in lymphoid BaF3 cells or its constitutive expression in either a murine transgenic model or in normal human cord blood cells is also sufficient to increase expression of EPOR. In order to further assess the direct regulation of EPOR by TEL-AML1 we next performed EMSA and ChIP experiments to confirm occupancy of AML1 consensus binding sites within the EPOR promoter and luciferase reporter assays to confirm up-regulation of EPOR promoter activity in the presence of TEL-AML1. Given the proposed pro-survival properties of EPO on non-erythroid cells, we next asked if the observed increase in expression of the EPOR in cells expressing TEL-AML1 could correlate with an increased cell survival in the presence of EPO. Cell survival experiments including growth curves, Annexin V staining and analysis of anti-apoptotic gene markers revealed that IL3-dependent cells expressing TEL-AML1 showed a prolonged survival in the presence of EPO alone and the consequent activation of anti-apoptotic pathways. The subsequent removal of TEL-AML1 expression in these cells resulted in cell death even in the presence of EPO, further suggesting that this effect is a consequence solely of fusion gene expression. Moreover, the observed survival in the presence of TEL-AML1 and EPO was enhanced by the co-addition of stem cell factor (SCF). Interestingly, SCF associated intracellular tyrosines have been shown to play an essential role in signaling through EPOR-SCFR interactions. In the presence of EPO, signaling via EPOR was confirmed by phosophorylation of JAK2, STAT5 and AKT and the subsequent up-regulation of BCL-XL. Similarly, EPOR functionality was demonstrated in TEL-AML1 positive patient cells by analysis of JAK2 pathway signaling after EPO induction. We propose that such activation lead to the increase in cell survival also observed in patient cells in the presence of EPO - again suggesting a role for enhanced cell survival through the EPO-EPOR axis. In our final model of pre-leukemia, CD34+ cord blood cells were transduced in vitro with a lentivirus capable of expressing both TEL-AML1 and GFP and ‘primed’ for pre-B lineage commitment. GFP (TEL-AML1) positive cord blood cells also showed increased levels of cell surface EPO receptor when compared to empty vector controls. Collectively, these data support the contention that TEL-AML1 directly activates ectopic expression of a functional EPO receptor and provides cell survival signals that may contribute critically to persistence of the pre-malignant clone in patients. Disclosures: No relevant conflicts of interest to declare.

Direct Targets of Epo Receptor-JAK2-pSTAT5 Signalling in Erythropoiesis

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 3881-3881
Author(s):  
Kevin R. Gillinder ◽  
Hugh Tuckey ◽  
Charles Bell ◽  
Stephen Huang ◽  
Melissa Ilsley ◽  
...  
Keyword(s):  
Target Genes ◽  
Erythropoietin Receptor ◽  
Lymphoid Cells ◽  
Erythroid Cell ◽  
Polycythaemia Vera ◽  
Erythroid Cells ◽  
Rna Seq ◽  
Feedback Controls ◽  
Erythroid Progenitor ◽  
In Erythroid Cells

Abstract Erythropoietin (EPO) acts through the dimeric erythropoietin receptor (EpoR) to stimulate proliferation, survival and differentiation of colony-forming units-erythroid (CFU-e). We undertook two complimentary approaches to find pSTAT5-depenendent and independent target genes in erythroid cells. We performed RNA-seq of newly transcribed (4sU-labelled) RNA, and ChIP-seq for pSTAT5, 30 minutes after EPO stimulation of J2E cells. This is the first time genome wide pSTAT5 ChIP-seq has been successfully undertaken in hematopoietic cells. We found ~320 robust pSTAT5 occupied sites in the erythroid genome. About 15% of these reside in promoters while the rest reside in intronic enhancers or intergenic regions, some >100kb from the nearest TSS. The majority of peaks contained a central palindromic GAS element, TTCYXRGAA, and there was significant enrichment of GATA and CACCC-box elements, suggesting co-regulation of some EPO-induced genes by GATA1 and KLF1. Using 4sU RNA-seq and CAGE data, we found just 57 genes to be immediately transcribed in response to EPO within 30 minutes while other target genes had delayed responses. We suggest this has biological relevance for feed forward and feedback controls on EPO driven erythropoiesis. Some of the DEGs (e.g. Bcl2l2 and Cish) are known direct targets of pSTAT5, but many are novel and suggest new pathways by which EPO regulates erythropoiesis. These include mRNA splicing, epigenetic regulation via histone methylation, and adaptive changes in the composition of the EpoR. This could provide increased sensitivity of erythroid progenitor cells to anaemic stress; i.e. sensitization of erythroid cells to JAK2-STAT5 signalling. We found a significant overlap between direct STAT5 target genes in erythroid cells, mammary epithelial cells and lymphoid cells, which imply conserved generic effectors of JAK-STAT5 signalling in different cell types. Our results provide new insights into how EPO co-ordinates erythroid cell proliferation, survival and differentiation. Some of these DEGs could be used as biomarkers for monitoring disease activity in polycythaemia vera (PV) and responses to JAK2 inhibitors. Disclosures Perkins: Novartis Oncology: Honoraria, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Honoraria.

scholarly journals Lyn Kinase Activity Is Required for Akt Mediated Erythroleukemia Cell Differentiation

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 24-24
Author(s):  
Janice H. C. Plani-LAM ◽  
Mhairi Maxwell ◽  
Neli Slavova-Azmanova ◽  
Nicole Kucera ◽  
Alison Louw ◽  
...  
Keyword(s):  
Tyrosine Kinases ◽  
Kinase Activity ◽  
Conflicts Of Interest ◽  
Negative Regulator ◽  
Erythroid Cell ◽  
Erythroid Cells ◽  
Wild Type ◽  
Erythroid Progenitors ◽  
Epo Receptor ◽  
Lyn Kinase

Erythroleukemia (M6 subtype of Acute Myeloid Leukaemia) is uncommon but has a poor prognosis, with reports of successful differentiation therapy using erythropoietin (Epo). Signaling through the Epo-receptor, which involves JAK2 and Lyn tyrosine kinases, controls red blood cell progenitor development. We have highlighted the importance of Lyn for regulating downstream Akt, and feed-back inhibitory signaling of the Epo-receptor through analysis of Lyn-/-, Lynup/up (hyperactive Lyn) and Cbp-/- (Csk-binding protein, a negative regulator of Lyn) erythroid cells. However, the importance of maintaining Lyn activity as opposed to Lyn protein for erythroid cell development and signaling, has not been delineated. To address this, we generated LynKD/KD mice (expressing a kinase dead K275M mutant Lyn), and analysed their erythroid compartment and signaling in immortalized erythroid progenitors. We show that LynKD/KD mice display splenic extramedullary erythropoiesis and have evidence of elevate bone marrow erythropoiesis, similar to Lyn-/- mice but with a less severe phenotype. Immortalized erythroid progenitors from LynKD/KD mice show impaired Epo-induced differentiation and a greater dependence on Epo for viability, but unaltered proliferation, compared to wild-type cells. Epo-induced signaling of LynKD/KD cells showed enhanced pJAK2/pSTAT5, reduced pAkt/pGAB2, and substantially reduced ALAS-e levels, compared to wild-type cells. Importantly, elevating Akt signaling in LynKD/KD cells by addition of phosphatase inhibitors (okadaic acid or Calyculin A), or via expression of active Akt, restored their differentiation capacity (and ALAS-e levels) and reduced their dependence on Epo for viability. We have unveiled that Lyn kinase activity, and not just its expression, is required for correct signaling of Akt to GATA-1 to maintain ALAS-e expression in erythroid cells, enabling hemoglobin production and viability during differentiation. Disclosures No relevant conflicts of interest to declare.

Erythropoietin receptor-dependent erythroid colony-forming unit development: capacities of Y343 and phosphotyrosine-null receptor forms

Blood ◽  
2002 ◽  
Vol 99 (3) ◽  
pp. 898-904 ◽  
Author(s):  
Chris P. Miller ◽  
Destin W. Heilman ◽  
Don M. Wojchowski
Keyword(s):  
Growth Factor Receptor ◽  
Cell Development ◽  
Erythropoietin Receptor ◽  
Janus Kinase ◽  
Erythroid Cell ◽  
Compensatory Mechanisms ◽  
Erythroid Progenitor ◽  
Epo Receptor ◽  
Binding Domains ◽  
Progenitor Cell Proliferation

Abstract Red cell development depends on the binding of erythropoietin (EPO) to receptors expressed by erythroid colony-forming units (CFUe) and the subsequent activation of receptor-bound Janus kinase (Jak2). Jak2 then mediates the phosphorylation of receptor tyrosine sites and the recruitment of 25 or more Src homology 2 domain-encoding proteins and associated factors. Previous studies have shown that an EPO receptor form containing Jak2-binding domains plus a single phosphotyrosine343 (PY343)–STAT5-binding site provides all signals needed for erythroid cell development. However, roles for PY343 and STAT5 remain controversial, and findings regarding PY-null receptor activities and erythropoiesis in STAT5-deficient mice are disparate. To study activities of a PY-null EPO receptor in primary cells while avoiding compensatory mechanisms, a form retaining domains for Jak2 binding and activation, but lacking all cytoplasmic tyrosine sites, was expressed in transgenic mice from aGATA1 gene-derived vector as a human epidermal growth factor receptor- murine EPO receptor chimera (EE-T-Y343F). The bio-signaling capacities of this receptor form were investigated in CFUe from thiamphenicol-treated mice. Interestingly, this PY-null EPO receptor form supported CFUe development (in the absence of detectable STAT5 activation) at efficiencies within 3-fold of those levels mediated by either an EE-T-Y343 form or the endogenous EPO receptor. However, EE-T-Y343F–dependent Ter119+ erythroblast maturation was attenuated. In tests of cosignaling with c-Kit, EE-T-Y343F nonetheless retained full capacity to synergize with c-Kit in promoting erythroid progenitor cell proliferation. Thus, EPO receptor PY-dependent events can assist late erythropoiesis but may be nonessential for EPO receptor–c-Kit synergy.

GATA-1 and Erythropoietin Cooperate to Promote Erythroid Cell Survival by Regulating bcl-xL Expression

Blood ◽  
1999 ◽  
Vol 94 (1) ◽  
pp. 87-96 ◽  
Author(s):  
Todd Gregory ◽  
Channing Yu ◽  
Averil Ma ◽  
Stuart H. Orkin ◽  
Gerd A. Blobel ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Cell Survival ◽  
Embryonic Stem ◽  
Erythroid Cell ◽  
Erythroid Cells ◽  
Downstream Effector ◽  
Apoptotic Protein ◽  
Related Proteins

The transcription factor GATA-1 is essential for normal erythropoiesis. By examining in vitro–differentiated embryonic stem cells, we showed previously that in the absence of GATA-1, committed erythroid precursors fail to complete maturation and instead undergo apoptosis. The mechanisms by which GATA-1 controls cell survival are unknown. Here we report that in erythroid cells, GATA-1 strongly induces the expression of the anti-apoptotic protein bcl-xL, but not the related proteins bcl-2 and mcl-1. Consistent with a role for bcl-xL in mediating GATA-1–induced erythroid cell survival, in vitro–differentiated bcl-xL−/− embryonic stem cells fail to generate viable mature definitive erythroid cells, a phenotype resembling that of GATA-1 gene disruption. In addition, we show that erythropoietin, which is also required for erythroid cell survival, cooperates with GATA-1 to stimulate bcl-xL gene expression and to maintain erythroid cell viability during terminal maturation. Together, our data show that bcl-xL is essential for normal erythroid development and suggest a regulatory hierarchy in which bcl-xL is a critical downstream effector of GATA-1 and erythropoietin-mediated signals.

scholarly journals AKT induces erythroid-cell maturation of JAK2-deficient fetal liver progenitor cells and is required for Epo regulation of erythroid-cell differentiation

Blood ◽  
2006 ◽  
Vol 107 (5) ◽  
pp. 1888-1891 ◽  
Author(s):  
Saghi Ghaffari ◽  
Claire Kitidis ◽  
Wei Zhao ◽  
Dragan Marinkovic ◽  
Mark D. Fleming ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
Cell Survival ◽  
Fetal Liver ◽  
Dominant Negative ◽  
Cell Maturation ◽  
Erythroid Cell ◽  
Erythroid Cells ◽  
Serine Threonine Kinase ◽  
Regulation Of Expression ◽  
Erythroid Cell Differentiation

AKT serine threonine kinase of the protein kinase B (PKB) family plays essential roles in cell survival, growth, metabolism, and differentiation. In the erythroid system, AKT is known to be rapidly phosphorylated and activated in response to erythropoietin (Epo) engagement of Epo receptor (EpoR) and to sustain survival signals in cultured erythroid cells. Here we demonstrate that activated AKT complements EpoR signaling and supports erythroid-cell differentiation in wild-type and JAK2-deficient fetal liver cells. We show that erythroid maturation of AKT-transduced cells is not solely dependent on AKT-induced cell survival or proliferation signals, suggesting that AKT transduces also a differentiation-specific signal downstream of EpoR in erythroid cells. Down-regulation of expression of AKT kinase by RNA interference, or AKT activity by expression of dominant negative forms, inhibits significantly fetal liver–derived erythroid-cell colony formation and gene expression, demonstrating that AKT is required for Epo regulation of erythroid-cell maturation.

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