Contrasting patterns of DNA strand breakage and ADP-ribosylation-dependent DNA ligation during granulocyte and...

Previous studies have shown that structural changes in DNA, including the ligation of pre-existing DNA breaks and the opening and closure of new breaks, occur shortly after exposure of granulomonocytic precursors (CFU-GM) to granulocyte-macrophage colony stimulating activity (GM-CSA). Monocytic differentiation of CFU-GM is selectively inhibited by compounds known to inhibit the nuclear enzyme ADP-ribosyl transferase (ADPRT). Since this enzyme, which transfers ADP-ribose units to chromatin proteins, is known to activate DNA ligase, we attempted to determine whether ligation of one or both types of DNA break is required for monocytic differentiation. Breaks in DNA were examined using the nucleoid sedimentation technique in which DNA breaks cause loss of DNA supercoiling in nucleoids and concomitant changes in their sedimentation through neutral sucrose gradients. We here report that two distinct patterns of DNA strand breakage and ligation are associated with differentiation to the granulocyte and monocyte lineages. Monocytic inducers (phorbolester and vitamin D3) predominantly produce closure of pre-existing strand breaks, whereas granulocytic inducers (granulocyte colony stimulating activity, G-CSA; retinoic acid) cause opening and closure of new breaks. Only ligation of the pre-existing breaks is highly sensitive to inhibition by 3-methoxybenzamide (a potent ADPRT inhibitor), and only monocytic differentiation is impaired by addition of this compound. These findings suggest that DNA structural changes may be directly involved in granulocyte-macrophage switching.