Involvement of H-ras in erythroid differentiation of TF1 and human umbilical cord blood CD34 cells.
To investigate the role of the ras gene in erythroid differentiation, a human erythroleukemic cell line, TF1, was transduced with a selectable retroviral vector carrying a mammalian wild type H-ras gene or a cytoplasmic dominant negative RAS1 gene. Transduction of TF1 cells with the wild type H-ras gene resulted in changes of cell types and up-regulation of erythroid-specific gene expression similar to that seen in differentiating erythroid cells. The number of red blood cell containing colonies derived from TF1 cells transduced with wild type H-ras cDNA was significantly increased and the cells in the colonies were more hemoglobinized as estimated by a deeper red color compared to those colony cells from mock or dominant negative RAS1 gene transduced TF1 cells, suggesting increased erythroid differentiation of TF1 cells after transduction of wild type H-ras in vitro. The mRNA levels of beta- and gamma-, but not alpha-, globin genes were significantly higher in H-ras transduced TF1 cells than those in TF1 cells transduced with mock or dominant negative RAS1 gene. Moreover, a 4kb pre-mRNA of the Erythropoietin receptor (EpoR) was highly expressed only in H-ras transduced TF1 cells. Additionally, human umbilical cord blood (CB) CD34 cells which are highly enriched for hematopoietic stem/progenitor cells were transduced with the same retroviral vectors to evaluate in normal primary cells the activities of H-ras in erythroid differentiation. Increased numbers of erythroid cell containing colonies (BFU-E and CFU-GEMM) were observed in CD34 cells transduced with the H-ras cDNA, compared to that from mock transduced cells. These data suggest a possible role for ras in erythroid differentiation.
Ge
Y
Y
Department of Microbiology/Immunology, Indiana University School of Medicine, Indianapolis 46202, USA.
Li
Z H
ZH
Marshall
M S
MS
Broxmeyer
H E
HE
Lu
L
L
D018952:Antigens, CD34 / Q000032:analysis; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D015672:Erythroid Precursor Cells / Q000166:cytology; D004920:Erythropoiesis / Q000235:genetics*; D005312:Fetal Blood / Q000166:cytology*; D011905:Genes, ras; D005914:Globins / Q000096:biosynthesis / Q000235:genetics; D016298:Hematopoietic Cell Growth Factors / Q000494:pharmacology; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D016283:Proto-Oncogene Proteins p21(ras) / Q000502:physiology*; D012333:RNA, Messenger / Q000096:biosynthesis; D017467:Receptors, Erythropoietin / Q000096:biosynthesis / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000502:physiology; D014162:Transfection
D018952:Antigens, CD34; D016298:Hematopoietic Cell Growth Factors; D012333:RNA, Messenger; D017467:Receptors, Erythropoietin; D011993:Recombinant Fusion Proteins; D005914:Globins; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras)