Cell death initiated by the adenosine analog 3-deazaadenosine (c3 Ado) was studied in human promyelocytic leukemia HL-60 cells. A rapid decrease in cell number was seen after 4-hr exposure to 50-100 microM c3 Ado. The dominating mode of cell death was apoptosis as demonstrated by condensation and fragmentation of the nucleus, formation of apoptotic bodies and endonucleolytic degradation of DNA. Four hour treatment with 100 microM c3 Ado resulted in a reduction of early S-phase cells, and appearance of cells with a lower DNA and protein content than that of the G1 population. Whereas 25 and 50 microM c3 Ado only initiated apoptosis in S-phase cells, 75 and 100 microM c3 Ado also initiated apoptosis in G1- and G2 + M-phase cells, suggesting different mechanisms for cell death at different concentrations. Apoptosis initiated by 100 microM c3 Ado was completely inhibited by 1 mM ZnCl2. Addition of homocysteine thiolactone (Hcy) partly inhibited cell death by c3 Ado. Light microscopic examination of cultures treated with 100 microM c3 Ado and 1 mM Hcy showed nuclear condensation and fragmentation consistent with the first stage in apoptosis, however, only a minor formation of apoptotic bodies took place in these cultures compared to that observed in cultures treated with 100 microM c3 Ado alone. The modifying action of Hcy on c3 Ado initiated apoptosis in HL-60 cells and this suggests that c3 Ado and 3-deazaadenosylhomocysteine (c3 AdoHcy) interact with different targets during initiation and progression of cell death in this cell line.