Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.10.1 (ERK)
95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Previous studies have demonstrated that SH2-containing inositol phosphatase (SHIP) is involved in the control of B cell, myeloid cell and macrophage activation and proliferation. The goal of the present study was to examine the role of SHIP during proliferation and apoptosis in cells of the erythroid lineage. Wild-type and catalytically inactive SHIP proteins were overexpressed in the erythropoietin (EPO)-dependent cell line AS-E2. Stable overexpression of catalytically inactive SHIP decreased proliferation and resulted in prolonged activation of the extracellular signal-regulated protein kinases ERK1/2 and protein kinase B (PKB), while wild-type SHIP did not affect EPO-mediated proliferation or phosphorylation of ERK and PKB. When AS-E2 cells were EPO deprived a significant increase in apoptosis was observed in clones overexpressing wild type. Mutational analysis showed that this increase in apoptosis was independent of the enzymatic activity of SHIP. The enhanced apoptosis due to overexpression of SHIP was associated with an increase in caspase-3 and -9 activity, without a distinct effect on caspase-8 activity or mitochondrial depolarization. Moreover, in cells overexpressing SHIP apoptosis could be reduced by a caspase-3 inhibitor. These data demonstrate that in the erythroid cell line AS-E2 overexpression of catalytically inactive SHIP reduced proliferation, while overexpression of wild-type SHIP had no effect. Furthermore, overexpression of SHIP enhanced apoptosis during growth factor deprivation by inducing specific caspase cascades, which are regulated independently of the 5-phosphatase activity of SHIP.
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PMID:Effects of overexpression of the SH2-containing inositol phosphatase SHIP on proliferation and apoptosis of erythroid AS-E2 cells. 1168 17

We report a method of purifying, characterizing and expanding endothelial cells (ECs) derived from CD133(+) bone marrow cells, a subset of CD34(+) haematopoietic progenitors. Isolated using immunomagnetic sorting (mean purity 90 +/- 5%), the CD133(+) bone marrow cells were grown on fibronectin-coated flasks in M199 medium supplemented with fetal bovine serum (FBS), vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and insulin growth factor (IGF-1). The CD133(+) fraction contained 95 +/- 4% CD34(+) cells, 3 +/- 2% cells expressing VEGF receptor (VEGFR-2/KDR), but did not express von Willebrand factor (VWF), VE-cadherin, P1H12 or TE-7. After 3 weeks of culture, the cells formed a monolayer with a typical EC morphology and expanded 11 +/- 5 times. The cells were further purified using Ulex europaeus agglutinin-1 (UEA-1)-fluorescein isothiocyanate (FITC) and anti-FITC microbeads, and expanded with VEGF for a further 3 weeks. All of the cells were CD45(-) and CD14(-), and expressed several endothelial markers (UEA-1, VWF, P1H12, CD105, E-selectin, VCAM-1 and VE-cadherin) and typical Weibel-Palade bodies. They had a high proliferative potential (up to a 2400-fold increase in cell number after 3 weeks of culture) and the capacity to modulate cell surface antigens upon stimulation with inflammatory cytokines. Purified ECs were also co-cultivated with CD34(+) cells, in parallel with a purified fibroblastic cell monolayer. CD34(+) cells (10 x 10(5)) gave rise to 17,951 +/- 2422 CFU-GM colonies when grown on endothelial cells, and to 12,928 +/- 4415 CFU-GM colonies on fibroblast monolayers. The ECs also supported erythroid blast-forming unit (BFU-E) colonies better. These results suggest that bone marrow CD133(+) progenitor cells can give rise to highly purified ECs, which have a high proliferative capacity, can be activated by inflammatory cytokines and are superior to fibroblasts in supporting haematopoiesis. Our data support the hypothesis that endothelial cell progenitors are present in adult bone marrow and may contribute to neo-angiogenesis.
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PMID:Differentiation and expansion of endothelial cells from human bone marrow CD133(+) cells. 1172 32

Erythropoiesis results from the proliferation and differentiation of pluripotent stem cells into immature erythroid progenitors (ie, erythroid burst-forming units (BFU-Es), whose growth, survival, and terminal differentiation depends on erythropoietin (Epo). Ineffective erythropoiesis is a common feature of myelodysplastic syndromes (MDS). We used a 2-step liquid-culture procedure to study erythropoiesis in MDS. CD34(+) cells from the marrow of patients with MDS were cultured for 10 days in serum-containing medium with Epo, stem cell factor, insulin-like growth factor 1, and steroid hormones until they reached the proerythroblast stage. The cells were then placed in medium containing Epo and insulin for terminal erythroid differentiation. Numbers of both MDS and normal control cells increased 10(3) fold by day 15. However, in semisolid culture, cells from patients with refractory anemia (RA) with ringed sideroblasts and RA or RA with excess of blasts produced significantly fewer BFU-Es than cells from controls. Fluorescence in situ hybridization analysis of interphase nuclei from patients with chromosomal defects indicated that abnormal clones were expanded in vitro. Epo-signaling pathways (STAT5, Akt, and ERK 1/2) were normally activated in MDS erythroid progenitors. In contrast, apoptosis was significantly increased in MDS cells once they differentiated, whereas it remained low in normal cells. Fas was overexpressed on freshly isolated MDS CD34(+) cells and on MDS erythroid cells throughout the culture. Apoptosis coincided with overproduction of Fas ligand during the differentiation stage and was inhibited by Fas-Fc chimeric protein. Thus, MDS CD34(+)-derived erythroid progenitors proliferated normally in our 2-step liquid culture with Epo but underwent abnormal Fas-dependent apoptosis during differentiation that could be responsible for the impaired erythropoiesis.
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PMID:In vitro proliferation and differentiation of erythroid progenitors from patients with myelodysplastic syndromes: evidence for Fas-dependent apoptosis. 1186 Dec 73

EphB4 (HTK) and its ligand, ephrinB2, are critical for angiogenesis and result in fatal abnormalities of capillary formation in null mice. EphB4 was originally identified in human bone marrow CD34(+) cells by us and has since been reported to be expressed in erythroid progenitors, whereas the ligand ephrinB2 is expressed in bone marrow stromal cells. Reasoning that the developmental relationship between angiogenesis and hematopoiesis implies common regulatory molecules, we assessed whether EphB4 signaling influences the function and phenotype of primitive human hematopoietic cells. Ectopically expressed EphB4 in cell lines of restricted differentiation potential promoted megakaryocytic differentiation, but not granulocytic or monocytic differentiation. Primary cord blood CD34(+) cells transduced with EphB4 resulted in the elevated expression of megakaryocytic and erythroid specific markers, consistent with EphB4 selectively enhancing some lineage-committed progenitors. In less mature cells, EphB4 depleted primitive cells, as measured by long-term culture-initiating cells or CD34(+)CD38(-) cell numbers, and increased progenitor cells of multiple cell types. Effects of ectopic EphB4 expression could be abrogated by either targeted mutations of select tyrosine residues or by the tyrosine kinase inhibitor, genistein. These data indicate that EphB4 accelerates the differentiation of primitive cells in a nonlineage-restricted manner but alters only select progenitor populations, influencing lineages linked by common ancestry with endothelial cells. EphB4 enforces preferential megakaryocytic and erythroid differentiation and may be a molecular bridge between angiogenesis and hematopoiesis.
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PMID:Receptor tyrosine kinase, EphB4 (HTK), accelerates differentiation of select human hematopoietic cells. 1192 61

During the initial stage of Friend virus-induced erythroleukemia in mice, interaction of the viral protein gp55 with the erythropoietin receptor, and other host factors, drives the expansion of erythroid precursor cells. Recently, we demonstrated that the Friend virus susceptibility locus, Fv2, which is required for the expansion of infected cells, encodes a naturally occurring, N-terminally truncated form of the Stk receptor tyrosine kinase (Sf-Stk). Here we show that in vitro expression of Sf-Stk confers Friend virus sensitivity to erythroid progenitor cells from Fv2(rr) mice. Furthermore, our data reveal that Sf-Stk kinase activity and Y436, but not Y429, are required for Epo-independent colony formation following Friend virus infection. Introduction of a mutation that results in failure to bind Grb2 abrogates the ability of Sf-Stk to induce colonies in response to Friend virus, while the Grb2 binding site from EGFR restores this response. Consistent with the ability of Grb2 to recruit SOS and Gab1, the Ras/MAPK and PI3K pathways are activated by Sf-Stk, and both of these pathways are required for gp55-mediated erythroblast proliferation. These data clearly demonstrate a requirement for signaling through Sf-Stk in the Epo-independent expansion of Friend virus-infected cells, and suggest a pivotal role for Grb2 in this response.
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PMID:Sf-Stk kinase activity and the Grb2 binding site are required for Epo-independent growth of primary erythroblasts infected with Friend virus. 1203 58

Two non-erythroid members of the erythrocyte Rhesus (Rh) protein family, RhBG and RhCG, have been recently cloned in the kidney. These proteins share homologies with specific NH(3)/NH(4)(+) transporters (Mep/Amt) in primitive organisms and plants. When expressed in a Mep-deficient yeast, RhCG can function as a bidirectional NH(3)/NH(4)(+) transporter. The aim of this study was to determine the intrarenal and intracellular location of RhCG in rat kidney. RT-PCR on microdissected rat nephron segments demonstrated expression of mRNAs encoding RhCG in distal convoluted tubules, connecting ducts, and cortical and outer medullary collecting ducts but not in proximal tubules and thick ascending limbs of Henle's loop. Immunolocalization studies performed on rat kidney sections with rabbit anti-human RhCG 31 to 48 antibody showed labeling of the apical pole of tubular cells within the cortex, the outer medulla, and the upper portion of the inner medulla. All cells within connecting tubules had identical apical staining. In cortical collecting ducts, a subpopulation of cells that has either apical staining (alpha-intercalated cells) or diffuse staining (beta-intercalated cells) for the beta1 subunit of the H(+)-ATPase, was heavily stained at their apical pole with the RhCG antibody while principal cells identified as H(+)-ATPase negative cells showed a faint apical staining for RhCG that was much less intense than in adjacent intercalated cells. In the outer medulla and the upper portion of the inner medulla, RhCG labeling was restricted to a subpopulation of cells within the collecting duct that apically express the beta1 subunit of the H(+)-ATPase, indicating that RhCG expression in medullary collecting ducts is restricted to intercalated cells. No labeling was seen in glomeruli, proximal tubules, and limbs of Henle's loop. Immunoblotting of apical membrane fractions from rat kidney cortex with the rabbit anti-human RhCG 31 to 48 antibody revealed a doublet band at approximatively 65 kD.
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PMID:Expression of RhCG, a new putative NH(3)/NH(4)(+) transporter, along the rat nephron. 1213 30

Two alternatively spliced stem cell factor (SCF) transcripts encode protein products, which differ in the duration of membrane presentation. One form, soluble SCF (S-SCF) gets rapidly processed to yield predominantly secreted protein. The other form, membrane-associated SCF (MA-SCF) lacks the primary proteolytic cleavage site but is cleaved slowly from an alternate site, and thus represents a more stable membrane form of SCF. Mutants of SCF that lack the expression of MA-SCF (Steel-dickie) or possess a defect in its presentation (Steel(17H)) manifest deficiencies in erythroid cell development. In this study, we have compared the consequence(s) of activating Kit, the receptor for SCF by MA-SCF with S-SCF, and an obligate membrane-restricted (MR) form of SCF (MR-SCF) on erythroid cell survival, proliferation, cell cycle progression, and the activation of p38 and ERK MAP kinase pathways. Activation of Kit by MR-SCF was associated with a significantly lower incidence of apoptosis and cell death in erythroid cells compared to either other isoform. MR- or MA-SCF-induced stimulation of erythroid cells resulted in similar and significantly greater proliferation and cell cycle progression compared to soluble SCF. The increase in proliferation and cell cycle progression via MA- or MR-SCF stimulation correlated with sustained and enhanced activation of p38 and ERK MAP kinase pathways. In addition, MR- or MA-SCF-induced proliferation was more sensitive to the inhibitory effects of ERK inhibitor compared to S-SCF-induced proliferation. In contrast, soluble SCF-induced proliferation was more sensitive to the inhibitory effects of p38 inhibitor compared with MR- or MA-SCF. These results suggest that different isoforms of SCF may use different biochemical pathways in stimulation of survival and/or proliferation of erythroid cells.
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PMID:Role of p38 and ERK MAP kinase in proliferation of erythroid progenitors in response to stimulation by soluble and membrane isoforms of stem cell factor. 1214 9

Valproate has been found to stimulate fetal hemoglobin (HbF) synthesis in patients with sickle cell disease. In accordance with these clinical observations, we found a moderate induction of HbF synthesis in K562 erythroid cells in vitro. Investigation of the role of the mitogen-activated protein kinase (MAPK) pathways by Western blot analysis and use of specific kinase inhibitors suggests that inhibition of ERK pathway and activation of the p38 pathway may contribute to the HbF-inducing activity of valproate.
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PMID:Induction of fetal hemoglobin synthesis by valproate: modulation of MAP kinase pathways. 1222 74

The expression/function of vascular endothelial growth factor (VEGF) receptors (VEGFR1/Flt1 and VEGFR2/KDR/Flk1) in hematopoiesis is under scrutiny. We have investigated the expression of Flt1 and kinase domain receptor (KDR) on hematopoietic precursors, as evaluated in liquid culture of CD34(+) hematopoietic progenitor cells (HPCs) induced to unilineage differentiation/maturation through the erythroid (E), megakaryocytic (Mk), granulocytic (G), or monocytic (Mo) lineage. KDR, expressed on 0.5% to 1.5% CD34(+) cells, is rapidly downmodulated on induction of differentiation. Similarly, Flt1 is present at very low levels in HPCs and is downmodulated in E and G lineages; however, Flt1 is induced in the precursors of both Mo and Mk series; ie, its level progressively increases during Mo maturation, and it peaks at the initial-intermediate culture stages in the Mk lineage. Functional experiments indicate that Mk and E, but not G and Mo, precursors release significant amounts of VEGF in the culture medium, particularly at low O(2) levels. The functional role of VEGF release on Mk maturation is indicated by 2 series of observations. (1) Molecules preventing the VEGF-Flt1 interaction on the precursor membrane (eg, soluble Flt1 receptors) significantly inhibit Mk polyploidization. (2) Addition of exogenous VEGF or placenta growth factor (PlGF) markedly potentiates Mk maturation. Conversely, VEGF does not modify Mo differentiation/maturation. Altogether, our results suggest that in the hematopoietic microenvironment an autocrine VEGF loop contributes to optimal Mk maturation through Flt1. A paracrine loop involving VEGF release by E precursors may also operate. Similarly, recent studies indicate that an autocrine loop involving VEGF and Flt1/Flk1 receptors mediates hematopoietic stem cell survival and differentiation.
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PMID:Autocrine-paracrine VEGF loops potentiate the maturation of megakaryocytic precursors through Flt1 receptor. 1240 76

We studied the effects of cyclosporin A (CsA) on the erythroid differentiation of human erythroid leukemia cell line K562. After K562 was treated with CsA for 4 days, the percentage of hemoglobinized cells was increased by 3.3 times. Because it was reported p38 MAPK (p38) and ERK are involved in erythropoietin-induced erythroid differentiation, we studied their roles using specific inhibitors. p38 inhibitor (SB203580) prevented CsA-induced hemoglobin synthesis in K562 cells, although MEK/ERK inhibitor (U0126) enhanced it by 3.3 times in K562 cells. These results indicate activation of p38 and inactivation of ERK are involved in CsA-induced erythroid differentiation of K562 cells.
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PMID:Cyclosporin A induces erythroid differentiation of K562 cells through p38 MAPK and ERK pathways. 1250 71


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