Modulation of globin gene expression in cultured erythroid precursors derived from normal individuals: transcriptional and posttranscriptional regulation by hemin.

We are interested in the genetic mechanisms whereby several classes of drugs increase fetal hemoglobin (HbF) in patients with sickle-cell anemia or beta-thalassemia. Recently, we have shown (Kollia et al., Proc. Natl. Acad. Sci. U.S.A. 93: 5693, 1996) that cultured primary human adult erythroid cells (hAEC) offer a useful model for the study of transcriptional and posttranscriptional regulation of globin gene expression. We have found also that hemin markedly increases HbF levels in these cells. We report here the effect of hemin on globin gene transcription and RNA processing in hAEC. Quantitative reverse transcriptase-polymerase chain reaction analysis showed that the gamma-globin message levels in the cytoplasm and nucleus were increased two-fold by hemin. In the untreated cells, only spliced gamma-transcripts were detected in the cytoplasm, indicating that only completely processed gamma-RNA is transported to the cytoplasm, whereas approximately half of the nuclear gamma-globin RNA transcripts were unspliced. After treatment with hemin, correctly spliced gamma-transcripts increased in the cytoplasm and nucleus, while the unprocessed gamma-transcripts decreased in number in the nucleus. We also studied epsilon-globin RNA transcripts; in the cytoplasm of untreated cells, only correctly processed transcripts were present, whereas the nuclear epsilon-globin RNA transcripts were unspliced. In hemin-induced cells, unspliced nuclear epsilon-transcripts decreased in number. In contrast to the gamma- and epsilon-globin genes, the levels of full-length, correctly spliced beta-globin message are not affected by hemin. Nuclear run-on transcription assays confirmed the increase in the rate of transcription of gamma- and epsilon-globin genes in hemin-treated versus untreated hAEC. These results indicate that hemin affects the expression of embryonic and fetal globin genes by acting both at the transcriptional and posttranscriptional levels. These results may be relevant to the action of other agents that affect the hemoglobin phenotype of human erythroid cells.