ACTIVATION OF ERYTHROID-SPECIFIC PROMOTERS DURING ANTHRACYCLINE-INDUCED DIFFERENTIATION OF K562 CELLS

Anthracycline antitumor drugs such as aclacinomycin (ACM) and doxorubi cin (DOX) used in subtoxic concentrations induce erythroid differentia tion of the erythroleukemic cell line K562. To elucidate the possible role of erythroid genes of the erythropoietin receptor (EpoR) and the transcription factor GATA-1 in this effect, the regulatory regions of the above genes and human epsilon- and gamma-globin and porphobilinoge n deaminase (PBGD) genes were fused to the firefly luciferase gene. Th e resulting reporter constructs were tested in a transfection assay of the erythroleukemic cell line K562 stimulated to differentiate by tre atment with the anthracycline drugs ACM and DOX or hemin (HEM). The re sults showed activation of the tested promoters after cell treatment w ith ACM, but not with DOX or HEM. In contrast to the mouse EpoR gene p romoter, the activity of the human EpoR gene promoter (-659/-60) in th e reporter construct was not modified by addition of the first intron sequence. In ACM-treated K562 cells, EpoR gene promoter activity compl etely correlated with EpoR and GATA-1 mRNA levels and the degree of er ythroid maturation. In addition, ACM strongly activated the erythroid gene promoters that contain GATA binding sites. Nevertheless, less act ivation was also observed for the GATA-1 gene promoter (-312/-31) lack ing any known GATA binding sites. Insertion of the GATA-1 gene enhance r with two canonic GATA binding sites, stimulated the ACM activation e ffect for EpoR and GATA-1 promoter-containing constructs. Mutation of the enhancer GATA binding sites abolished this effect. All the regulat ory regions tested (except gamma-globin promoter) were completely inac tive in nonerythroid COS7 cells. These data indicate that (1) two stru cturally different anthracycline analogues, DOX and ACM, differ in the ir differentiation mechanisms, and (2) ACM switches on the erythroid p rogram of K562 cells, at least in part because of interaction with a f actor(s) that recognizes the GATA binding sites in the promoter region of erythroid genes leading to their activation, (C) 1996 by The Ameri can Society of Hematology.