3'-Azido-3'-deoxythymidine inhibits globin gene transcription in butyric acid-induced K-562 human leukemia cells.
We previously demonstrated that 3'-azido-3'-deoxythymidine (AZT) inhibits proliferation of human bone marrow progenitor cells in vitro and that incorporation of AZT into nuclear DNA may be one mechanism responsible for AZT-induced bone marrow toxicity [Antimicrob. Agents Chemother. 31:452-454 (1987); Mol. Pharmacol. 36:9-14 (1989)]. The present study explores possible genetic mechanisms involved in AZT-induced anemia by evaluating the effects of AZT on globin gene expression at both the transcriptional and the translational levels in butyric acid-induced K-562 human erythroleukemia cells. AZT, at concentrations ranging from 10 to 250 microM, was added to cells 25 hr after initiation of induction of hemoglobin (Hb) synthesis with 1.4 mM butyric acid. Hb synthesis, as measured by benzidine staining, was inhibited by AZT in a dose- and time-dependent manner in these cells. AZT inhibition of cell growth was not the major contributing factor in the net inhibition of Hb synthesis in K-562 cells. As assessed by Northern blot analysis, AZT inhibition of Hb synthesis was associated with a decrease in globin mRNA steady state levels without inhibition of total RNA synthesis or actin mRNA steady state levels. In particular, a decrease of globin mRNA levels of 23% by 25 microM AZT was observed, reaching a maximum inhibition of 59% in the presence of 250 microM AZT. In vitro translation experiments demonstrated that essentially all nonglobin translatable mRNAs were not inhibited by AZT concentrations as high as 250 microM, whereas globin mRNAs coding for epsilon, zeta, A gamma, G gamma, and alpha chains were substantially inhibited to similar levels by AZT, in a dose-dependent manner. Transcriptional run-on studies with isolated nuclei from AZT-treated K-562 cells demonstrated a 20 and 50% inhibition of in vitro synthesized globin transcripts from cells exposed to 25 and 100 microM AZT, respectively. 2',3'-Dideoxycytidine also inhibited K-562 cell growth in the same concentration range as AZT but, of importance, had no effects on Hb production. These data suggest that inhibition of globin gene expression may play a role in the cytotoxicity of AZT to the erythroid cell.
Weidner
D A
DA
Department of Pharmacology, Center for AIDS Research, Birmingham, Alabama.
Sommadossi
J P
JP
D000740:Anemia / Q000139:chemically induced; D002087:Butyrates / Q000494:pharmacology; D020148:Butyric Acid; D004305:Dose-Response Relationship, Drug; D005914:Globins / Q000235:genetics*; D006454:Hemoglobins / Q000096:biosynthesis; D006801:Humans; D004915:Leukemia, Erythroblastic, Acute / Q000378:metabolism / Q000473:pathology; D014176:Protein Biosynthesis / Q000187:drug effects; D014158:Transcription, Genetic / Q000187:drug effects*; D014407:Tumor Cells, Cultured; D016047:Zalcitabine / Q000494:pharmacology; D015215:Zidovudine / Q000494:pharmacology*
D002087:Butyrates; D006454:Hemoglobins; D020148:Butyric Acid; D015215:Zidovudine; D016047:Zalcitabine; D005914:Globins