Ge Y, Li Z H, Marshall M S, Broxmeyer H E, Lu L
Department of Microbiology/Immunology, Indiana University School of Medicine, Indianapolis 46202, USA.
Blood Cells Mol Dis. 1998 Jun;24(2):124-36; discussion 137. doi: 10.1006/bcmd.1998.0179.
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.
为了研究ras基因在红系分化中的作用，用携带哺乳动物野生型H-ras基因或细胞质显性负性RAS1基因的可选择逆转录病毒载体转导人红白血病细胞系TF1。用野生型H-ras基因转导TF1细胞导致细胞类型改变和红系特异性基因表达上调，类似于在分化的红系细胞中所见。与用空载体或显性负性RAS1基因转导的TF1细胞形成的集落细胞相比，用野生型H-ras cDNA转导的TF1细胞形成的含红细胞集落数量显著增加，且集落中的细胞血红蛋白化程度更高，这可通过更深的红色来估计，表明体外转导野生型H-ras后TF1细胞的红系分化增加。在H-ras转导的TF1细胞中，β-和γ-珠蛋白基因（而非α-珠蛋白基因）的mRNA水平显著高于用空载体或显性负性RAS1基因转导的TF1细胞。此外，促红细胞生成素受体（EpoR）的4kb前体mRNA仅在H-ras转导的TF1细胞中高表达。另外，用相同的逆转录病毒载体转导高度富集造血干/祖细胞的人脐带血（CB）CD34细胞，以评估正常原代细胞中H-ras在红系分化中的活性。与空载体转导的细胞相比，在用H-ras cDNA转导的CD34细胞中观察到含红系细胞集落（BFU-E和CFU-GEMM）数量增加。这些数据表明ras在红系分化中可能发挥作用。
