Growth suppression of Friend virus-transformed erythroleukemia cells by p53 protein is accompanied by hemoglobin production and is sensitive to erythropoietin

1993 ◽  
Vol 13 (3) ◽  
pp. 1456-1463
Author(s):  
P Johnson ◽  
S Chung ◽  
S Benchimol
Keyword(s):  
Cell Line ◽  
P53 Protein ◽  
Erythroid Differentiation ◽  
Erythropoietin Receptor ◽  
Temperature Sensitive ◽  
Friend Virus ◽  
Erythroleukemia Cells ◽  
Erythroleukemia Cell ◽  
Hemoglobin Production ◽  
Striking Effect

The murine allele temperature-sensitive (ts) p53Val-135 encodes a ts p53 protein that behaves as a mutant polypeptide at 37 degrees C and as a wild-type polypeptide at 32 degrees C. This ts allele was introduced into the p53 nonproducer Friend erythroleukemia cell line DP16-1. The DP16-1 cell line was derived from the spleen cells of a mouse infected with the polycythemia strain of Friend virus, and like other erythroleukemia cell lines transformed by this virus, it grows independently of erythropoietin, likely because of expression of the viral gp55 protein which binds to and activates the erythropoietin receptor. When incubated at 32 degrees C, DP16-1 cells expressing ts p53Val-135 protein, arrested in the G0/G1 phase of the cell cycle, rapidly lost viability and expressed hemoglobin, a marker of erythroid differentiation. Erythropoietin had a striking effect on p53Val-135-expressing cells at 32 degrees C by prolonging their survival and diminishing the extent of hemoglobin production. This response to erythropoietin was not accompanied by down-regulation of viral gp55 protein.

scholarly journals Growth suppression of Friend virus-transformed erythroleukemia cells by p53 protein is accompanied by hemoglobin production and is sensitive to erythropoietin.

1993 ◽  
Vol 13 (3) ◽  
pp. 1456-1463 ◽  
Author(s):  
P Johnson ◽  
S Chung ◽  
S Benchimol
Keyword(s):  
Cell Line ◽  
P53 Protein ◽  
Erythroid Differentiation ◽  
Erythropoietin Receptor ◽  
Temperature Sensitive ◽  
Friend Virus ◽  
Erythroleukemia Cells ◽  
Erythroleukemia Cell ◽  
Hemoglobin Production ◽  
Striking Effect

The murine allele temperature-sensitive (ts) p53Val-135 encodes a ts p53 protein that behaves as a mutant polypeptide at 37 degrees C and as a wild-type polypeptide at 32 degrees C. This ts allele was introduced into the p53 nonproducer Friend erythroleukemia cell line DP16-1. The DP16-1 cell line was derived from the spleen cells of a mouse infected with the polycythemia strain of Friend virus, and like other erythroleukemia cell lines transformed by this virus, it grows independently of erythropoietin, likely because of expression of the viral gp55 protein which binds to and activates the erythropoietin receptor. When incubated at 32 degrees C, DP16-1 cells expressing ts p53Val-135 protein, arrested in the G0/G1 phase of the cell cycle, rapidly lost viability and expressed hemoglobin, a marker of erythroid differentiation. Erythropoietin had a striking effect on p53Val-135-expressing cells at 32 degrees C by prolonging their survival and diminishing the extent of hemoglobin production. This response to erythropoietin was not accompanied by down-regulation of viral gp55 protein.

scholarly journals STAT5 Activation Correlates with Erythropoietin Receptor-mediated Erythroid Differentiation of an Erythroleukemia Cell Line

1997 ◽  
Vol 272 (13) ◽  
pp. 8149-8152 ◽  
Author(s):  
Ken Iwatsuki ◽  
Takaho Endo ◽  
Hiroyuki Misawa ◽  
Masahiro Yokouchi ◽  
Akira Matsumoto ◽  
...  
Keyword(s):  
Cell Line ◽  
Erythroid Differentiation ◽  
Erythropoietin Receptor ◽  
Erythroleukemia Cell

scholarly journals Putative oncogenic role of the erythropoietin receptor in murine and human erythroleukemia cells

Blood ◽  
1994 ◽  
Vol 83 (7) ◽  
pp. 1813-1821 ◽  
Author(s):  
S Chretien ◽  
F Moreau-Gachelin ◽  
F Apiou ◽  
G Courtois ◽  
P Mayeux ◽  
...  
Keyword(s):  
Long Terminal Repeat ◽  
Cell Line ◽  
Binding Sites ◽  
Genetic Alterations ◽  
Erythropoietin Receptor ◽  
R Gene ◽  
Normal Cells ◽  
Erythroleukemia Cells ◽  
Erythroleukemia Cell

Abstract To determine whether the erythropoietin receptor (Epo-R) plays a role in the course of malignant erythropoietic disorders, this gene was studied in murine and human erythroleukemia cells. An altered Epo-R gene was found in a murine Friend erythroleukemia cell line, FCL1, due to a spleen focus-forming virus (SFFV) long terminal repeat insertion within the noncoding region of the first exon, leading to Epo-R mRNA overexpression. A similar mechanism of Epo-R activation has previously been described in the T3CL-2 Friend erythroleukemia cell line. An elevated number of Epo-binding sites has been observed in two human erythroleukemia cell lines, TF-1 and UT7. In UT7 cells, homogeneously staining region of the short arm of chromosome 19 [hsr (19)] was evidenced, which contained an amplification of the Epo-R gene. This Epo- R gene amplification was confirmed by the quantification of Southern blots in which the intensity of the Epo-R signal was compared in UT7 DNA and in DNA from normal cells. The Epo-R gene was present in UT7 at a mean number of seven to eight copies per cell. Interestingly, the Epo- R gene was rearranged; the breakpoint region was located near the 3′ end of the gene, 3 kb downstream from the end of the last exon. Taken together, these results suggest that, in both murine and human systems, genetic alterations of the Epo-R gene are not rare events and could be involved in the occurrence of the erythroleukemic process.

scholarly journals Putative oncogenic role of the erythropoietin receptor in murine and human erythroleukemia cells

Blood ◽  
1994 ◽  
Vol 83 (7) ◽  
pp. 1813-1821 ◽  
Author(s):  
S Chretien ◽  
F Moreau-Gachelin ◽  
F Apiou ◽  
G Courtois ◽  
P Mayeux ◽  
...  
Keyword(s):  
Long Terminal Repeat ◽  
Cell Line ◽  
Binding Sites ◽  
Genetic Alterations ◽  
Erythropoietin Receptor ◽  
R Gene ◽  
Normal Cells ◽  
Erythroleukemia Cells ◽  
Erythroleukemia Cell

To determine whether the erythropoietin receptor (Epo-R) plays a role in the course of malignant erythropoietic disorders, this gene was studied in murine and human erythroleukemia cells. An altered Epo-R gene was found in a murine Friend erythroleukemia cell line, FCL1, due to a spleen focus-forming virus (SFFV) long terminal repeat insertion within the noncoding region of the first exon, leading to Epo-R mRNA overexpression. A similar mechanism of Epo-R activation has previously been described in the T3CL-2 Friend erythroleukemia cell line. An elevated number of Epo-binding sites has been observed in two human erythroleukemia cell lines, TF-1 and UT7. In UT7 cells, homogeneously staining region of the short arm of chromosome 19 [hsr (19)] was evidenced, which contained an amplification of the Epo-R gene. This Epo- R gene amplification was confirmed by the quantification of Southern blots in which the intensity of the Epo-R signal was compared in UT7 DNA and in DNA from normal cells. The Epo-R gene was present in UT7 at a mean number of seven to eight copies per cell. Interestingly, the Epo- R gene was rearranged; the breakpoint region was located near the 3′ end of the gene, 3 kb downstream from the end of the last exon. Taken together, these results suggest that, in both murine and human systems, genetic alterations of the Epo-R gene are not rare events and could be involved in the occurrence of the erythroleukemic process.

scholarly journals Erythropoietin and Friend Virus gp55 Activate Different JAK/STAT Pathways through the Erythropoietin Receptor in Erythroid Cells

1998 ◽  
Vol 18 (3) ◽  
pp. 1172-1180 ◽  
Author(s):  
Yasuko Yamamura ◽  
Hisato Senda ◽  
Yukio Kageyama ◽  
Tomoko Matsuzaki ◽  
Makoto Noda ◽  
...  
Keyword(s):  
Cell Growth ◽  
Dna Sequences ◽  
Protein Tyrosine Kinase ◽  
Nuclear Translocation ◽  
Erythropoietin Receptor ◽  
Friend Virus ◽  
Erythroid Progenitor ◽  
Erythroid Progenitor Cells ◽  
Erythroleukemia Cells ◽  
Erythroleukemia Cell

ABSTRACT Abnormal erythropoietin (EPO)-independent cell growth is induced after infection of erythroid progenitor cells with a polycythemic strain of Friend virus (FVp). Binding of its Env-related glycoprotein (gp55) to the EPO receptor (EPOR) mimics the activation of the EPOR with EPO. We investigated the gp55-EPOR signaling in erythroblastoid cells from mice infected with FVp and in cells of FVp-induced or gp55-transgenic-mouse-derived erythroleukemia cell lines, comparing it with the EPO-EPOR signaling in EPO-responsive erythroblastoid cells. While the Janus protein tyrosine kinase JAK2 and the transcription factor STAT5 became tyrosine phosphorylated with the EPO stimulation in EPO-responsive erythroblastoid cells from anemic mice, JAK1 and STAT5 were constitutively tyrosine phosphorylated in all of these FVpgp55-induced erythroblastoid or erythroleukemic cells. Moreover, this constitutively tyrosine-phosphorylated STAT5 was unable to bind to its specific DNA sequences and did not translocate to the nucleus. Nuclear translocation and DNA binding of this STAT5 species required EPO stimulation. These findings clearly indicate that the FVpgp55-EPOR signaling is distinct from the EPO-EPOR signaling and suggest that STAT5 may not play an essential role in the transmission of the cell growth signals in FVp gp55-induced erythroleukemia cells.

scholarly journals Stem cell factor influences the proliferation and erythroid differentiation of the MB-02 human erythroleukemia cell line by binding to a high-affinity c-kit receptor

Blood ◽  
1993 ◽  
Vol 82 (2) ◽  
pp. 436-444 ◽  
Author(s):  
VC Broudy ◽  
DA Morgan ◽  
N Lin ◽  
KM Zsebo ◽  
FW Jacobsen ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Stem Cell ◽  
Growth Factors ◽  
Cell Line ◽  
Stem Cell Factor ◽  
Erythroid Differentiation ◽  
Gm Csf ◽  
High Affinity ◽  
Kit Receptor ◽  
Erythroleukemia Cell

Abstract Stem cell factor (SCF) acts in synergy with other growth factors such as erythropoietin (Epo), granulocyte-macrophage colony-stimulating factor (GM-CSF), or interleukin-3 (IL-3), to stimulate the growth of primitive hematopoietic cells. Because of the prominent role of CSF in the maintenance of normal erythropoiesis in vivo, we examined the effects of SCF on the Epo-inducible human erythroleukemia cell line MB- 02, and characterized the c-kit receptor in these cells. MB-02 cells cultured in serum-containing media do not survive in the absence of exogenous growth factors, but the addition of SCF, Epo, or IL-3 as a single factor enhanced MB-02 survival. Furthermore, in the presence of Epo, SCF (5 to 25 ng/mL) enhanced MB-02 proliferation in a dose- dependent manner, and increased the relative and absolute number of benzidine-positive cells generated. SCF also enhanced cell proliferation in the presence of either IL-3 or low concentrations of GM-CSF. A neutralizing anti-c-kit receptor monoclonal antibody (SR-1) blocked binding of 125I-SCF to MB-02 cells by 98%, and the effect of SCF on MB-02 growth, c-kit receptor-binding parameters were quantitated by equilibrium-binding experiments with 125I-SCF. MB-02 cells display a single class of high-affinity (50 pmol/L) c-kit receptors, with approximately 8,000 receptors per cell. The molecular weight of the c- kit receptor was determined by affinity cross-linking 125I-SCF to MB-02 cells. 125I-SCF-c-kit receptor complexes of approximately 155,000 and approximately 310,000 daltons were found, likely representing the monomeric and dimeric forms of the c-kit receptor. The binding affinity and molecular weight of the c-kit receptor on MB-02 cells are similar to those of normal human marrow cells. These results suggest that SCF synergizes with Epo to influence not only the proliferation but the erythroid differentiation of MB-02 cells. Thus, the MB-02 cell line may be a useful model in which to investigate the molecular mechanisms of SCF action.

Truncated erythropoietin receptor in a murine erythroleukemia cell line

1996 ◽  
Vol 28 (2) ◽  
pp. 175-181 ◽  
Author(s):  
Thomas Bittorf ◽  
Samantha J. Busfield ◽  
S. Peter Klinken ◽  
Peta A. Tilbrook
Keyword(s):  
Cell Line ◽  
Erythropoietin Receptor ◽  
Erythroleukemia Cell ◽  
Murine Erythroleukemia ◽  
Murine Erythroleukemia Cell

AHSP Knockdown in Human Erythroleukemia Cell Line and Human Hematopoietic Stem Cells Results in Alpha Hemoglobin Chain Precipitation, Decreased Hemoglobin Levels and Increased Cell Death.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1778-1778
Author(s):  
Flavia O. Pinho ◽  
Dulcineia M. Albuquerque ◽  
Sara T.O Saad ◽  
Fernando F. Costa
Keyword(s):  
Stem Cells ◽  
Cell Death ◽  
Hematopoietic Stem Cells ◽  
Cell Line ◽  
K562 Cells ◽  
Erythroid Differentiation ◽  
Hematopoietic Stem ◽  
Cd34 Cells ◽  
Erythroleukemia Cell ◽  
Human Red Cells

Abstract Alpha Hemoglobin Stabilizing Protein (AHSP) binds alpha hemoglobin chain (αHb), avoiding its precipitation and its pro-oxidant activity. In the presence of beta hemoglobin chain (βHb), the αHb-AHSP complex is dismembered and βHb displaces AHSP to generate the quaternary structure of hemoglobin. These data have been obtained in vitro and in mouse cells, but strongly suggest the importance of AHSP for normal hemoglobin synthesis in humans. To the best of our knowledge, the relationship between hemoglobin formation and alterations in AHSP expression has not yet been described in human red cells. Hence, to investigate the consequences of a reduced AHSP synthesis in human red cells, we established the RNA interference-mediated knockdown of AHSP expression in human erythroleukemia cell line (K562 cells) and human hematopoietic stem cells (CD34+ cells) induced to erythroid differentiation, and analyzed the consequent cellular and molecular aspects of AHSP knockdown in these cells. shRNA expression vectors, aimed at the AHSP mRNA target sequence, were cloned and transfected into K562 and CD34+ cells using a non-liposomal lipid reagent. Following transfection, K562 cells that stably expressed AHSP-shRNA and CD34+ cells that transiently expressed AHSP-shRNA were selected. K562 and CD34+ cells were stimulated to erythroid differentiation by hemin and erythropoietin (EPO) respectively. The cells were examined in terms of gene expression using quantitative real-time PCR; production of reactive oxygen species (ROS), apoptosis and hemoglobin production through flow cytometry assays; and immunofluorescence assays for globin chains. AHSP-shRNA hemin-induced K562 cells and AHSP-shRNA EPO-induced CD34+ cells presented 71% and 75% decreases in AHSP expression levels, respectively. The RNAi-mediated knockdown of AHSP expression resulted in a considerable αHb precipitation, as well as in a significant decrease in fetal hemoglobin formation. In addition, AHSP-knockdown cells demonstrated an increased ROS production and increased rate of apoptosis. These findings strengthen the hypothesis that AHSP stabilizes the alpha hemoglobin chain, avoiding its precipitation and its ability to generate ROS which implicate in cell death. Moreover, data indicate that AHSP may be highly significant for human hemoglobin formation and suggest that AHSP is a key chaperone protein during human erythropoiesis.

scholarly journals Induction and inhibition of Friend erythroleukemia cell differentiation by pyrimidine analogs: analysis of the requirement for intracellular accumulation and incorporation into DNA.

1982 ◽  
Vol 2 (8) ◽  
pp. 1020-1024 ◽  
Author(s):  
J A Bilello ◽  
K K Gauri ◽  
J Kühne ◽  
G Warnecke ◽  
G Koch
Keyword(s):  
Cell Differentiation ◽  
Dimethyl Sulfoxide ◽  
Dna Analysis ◽  
Erythroid Differentiation ◽  
Intracellular Accumulation ◽  
Induced Differentiation ◽  
Alkyl Chains ◽  
Erythroleukemia Cells ◽  
Erythroleukemia Cell ◽  
Pyrimidine Analogs

Alkyldeoxyuridines which differ from thymidine by a C5 substitution of straight or branched alkyl chains of two to six carbon atoms have been tested for their ability to be taken up, phosphorylated, and incorporated into DNA. Analysis of the uptake of 5-ethyl-2'-deoxyuridine and 5-propyl-2'-deoxyuridine (n-PrdU)--similar to both thymidine and 5-bromo-2'-deoxyuridine--indicates that transport is dependent upon a functional cellular thymidine kinase. All of the aforementioned pyrimidines with the exception of n-PrdU are phosphorylated to the triphosphate and incorporated into DNA. The homologs 5-iso-propyl-2'-deoxyuridine (iso-PrdU) and 5-hexyl-2'-deoxyuridine are neither transported into the cell, phosphorylated, nor incorporated into DNA. These analogs were tested (i) for their ability to induce in the absence of dimethyl sulfoxide and (ii) to determine whether they enhance or inhibit dimethyl sulfoxide-induced differentiation of Friend erythroleukemia cells. Inhibition of erythroid differentiation appears to require the incorporation of thymidine analogs into DNA, and thus only 5-ethyl-2'-deoxyuridine and 5-bromo-2'-deoxyuridine were effective in inhibiting dimethyl sulfoxide-induced differentiation. The observation that iso-PrdU, and to a lesser extent n-PrdU and 5-propyldeoxyuridine monophosphate, induce differentiation under conditions in which they are not detectable intracellularly is strong evidence that this class of inducer acts at the cell membrane.

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