Regulation of DNA repair and replication in proliferating FL cells

Abstract

In this paper, we show that the rate of total DNA synthesis increases twice during the Friend erythroleukemia (FL) cell culture growth cycle. The first increase takes place at the threshold of the logarithmic (early) phase of growth, while the second peak takes place in the mid-logarithmic (late) phase. This bi-phasic pattern can be reproduced in both normal and dimethylsulfoxide (DMSO)-differentiating cells, using either [3H]-thymidine or [14Cl-methyl-L-methionine as precursors for DNA labelling. In normal cells, the non-replicating strands incorporate some radioactivity originating in [3H]-thymidine, essentially in the early phase, while their labelling is insignificant in the late phase. In UV-irradiated cells, the mechanism yielding repair patches (RPs) contained by the same non-replicating strands is highly facilitated in the earlier stage in comparison with its functioning at the later stage. In contrast, the [3H]-thymidine labelling of the semiconservatively newly replicating strands is insignificant at the early stage and more substantial later, as expected. In UV-irradiated cells, the slightly decreased [3H]-thymidine labelling of replicating strands is probably explained by the concomitant small percentage of cell death. The chronology of the optimal rates of early repair and later replication is quite well confirmed by a bi-phasic regulation of the total DNA poly-merase (pol) activity: while a peak of N-ethylmaleimide (MalNEt)-resistant pols activity (accounting for base excision-repair) is seen early on, a peak of MalNEt-sensitive polα activity (accounting for replication) is found later. DMSO strikingly inhibits both the rate of the early base excision-repair (either exploiting [3H]-thymidine or [14C]-methyl-L-methionine) and the early MalNEt-resistant pols activity. It does not, however, inhibit the late replication and the late MalNEt-sensitive polα activity, when hemoglobin (Hb) synthesis is heavily induced. The time-shift of late replication in DMSO-differentiating cells is probably explained by a delay of theS-phase, as also suggested by the net decrease in the amount of DNA per nucleus during the second half of the culture growth cycle.