Embryonic erythroid differentiation in the human leukemic cell line K562.

Abstract

K562 human leukemia cells synthesize embryonic hemoglobins after culture in the presence ofhemin. We have rigorously identified these hemoglobins by globin chain analysis and peptide mapping. No adult hemoglobin could be detected, and (3-globin synthesis was less than 2 ppm of total protein synthesis. Persistent embryonic globin gene expression is known to occur as a consequence of globin gene deletions. However, restriction endonuclease mapping showed that the globin gene complexes in K562 cells are indistinguishable from normal. Hemin increased the rate ofembryonicglobin synthesis. The pattern ofhemoglobin synthesis proved to be stable when cells from different laboratories were compared. One line, however, synthesized large amounts ofHb X and very little Hb Portland in response to hemin. Hb X has been previously detected in human embryos; we show here that it has the composition Eay2 and is diagnostic of imbalanced chain synthesis or' % thalassemia."We have identified several agents that induce hemoglobin synthesis in K562 cells. Different inducers induced different patterns of embryonic hemoglobin synthesis but never any adult hemoglobin synthesis.One of the limitations to the study of the factors regulating the changes in human hemoglobin production that occur between embryonic, fetal, and adult life (1) has been the lack of a selfsustaining cell line in which human hemoglobin genes are expressed. Recently, Andersson et al. (2,3) reported that the human cell line K562, derived from a patient who had chronic myeloid leukemia in terminal blast cell crisis (4), has certain erythroid characteristics. In a preliminary report (5), we have shown that these cells can be induced to produce embryonic and fetal but not adult hemoglobin. In this paper, we report an analysis ofthe potential ofthe K562 cell line as a model for studying hemoglobin differentiation.Lozzio et al., who established the K562 cell line in culture (4) have questioned the erythroid nature of K562 cells (6). If the pattern of globin synthesis we observed is the consequence of abnormal and unstable gene expression in a leukemic cell line or the result of gene rearrangement in culture, the utility of these cells for the study of the normal control of hemoglobin synthesis would be slight.We found that the cells available to us showed a stable pattern of erythroid differentiation in response to hemin and that embryonic hemoglobins were synthesized but adult (13) globin synthesis was strictly repressed. This is the converse of the situation in normal adult erythroid tissue but closely resembles the state of the normal embryo. Restriction endonuclease analysis of the globin genes has failed to detect any gene deletion or rearrangements, and we thus propose that the presence of embryonic and the absense of adult globin synthesis is the consequence of epigenetic control analogous to that in normal embryonic erythroid cells. Nevertheless, by selection of variant cell lines and the choice of hemoglobin inducer, the varieties of hemoglobin produced c...

Development of sensitive fluorescent assays for embryonic and fetal hemoglobin inducers using the human β-globin locus in erythropoietic cells

“…Hemin-induced erythroid differentiation of K562 cells has previously been reported to result in a sharp increase in ⑀-globin and ␥-globin gene expression that is associated with cytoplasmic accumulation of embryonic Gower ( 2 ⑀ 2 ) and fetal Portland ( 2 ␥ 2 ) hemoglobin molecules. 44,45,48 In this study, we show that hemin-induced erythroid differentiation of episomally transfected K562-EBNA1 cells resulted in a significant increase in EGFP expression driven by the ⑀-, G ␥-and A ␥-globin promoters, while EGFP expression from the ␤-and ␦-globin promoters consistently produced low levels of EGFP. Importantly, we were able to demonstrate quantitatively the dependence of EGFP expression in K562-EBNA1 cells on hemin concentration.…”

“…Thus, our findings are quantitatively and qualitatively in general accordance with the sharp increase in the expression of the ⑀-and ␥-globin genes that occurs in K562 cells after induction with hemin. 44,45,48 However, the expression of EGFP under each globin promoter in the different constructs may be influenced by the deletion of downstream regulatory elements during the creation of these constructs. This was directly examined by comparing the expression from the G ␥-␤-and G ␥-A ␥-EGFP EBAC constructs (Figure 1).…”

“…Hemin can induce K562 cells to undergo erythroid differentiation, which is associated with a sharp increase in embryonic and fetal globin gene expression. 44,45,48 We therefore investigated whether hemin could enhance EGFP expression from the EGFP-modified ␤-globin EBAC constructs. K562-EBNA1 cells transfected with the EGFP-modified globin EBACs and cultured for 40 days were treated with various doses of hemin (0-100 M) for up to 5 days.…”

Induction by HMBA and DMSO of genes introduced into mouse erythroleukemia and other cell lines by transient transfection.

“…K562 cells were therefore grown with hemin, an inducer of K562 (Rutherford et al 1981), with HMBA, or with both inducers [ Fig. 81.…”

Hemoglobin Expression in Clones of K562 Cell Line

“…In particular, Rutherford et al [6,7,10] and Benz et al [8] have rigorously shown that embryonic hemoglobins (Hb Portland and Hb Gower) as well as small quantities of fetal Hb (HbF) are produced by the K562 cells after exposure to hemin. Thus, this cell line provides a new tool to study erythroid differentiation and the switch between embryonic and fetal hemoglobins.…”