Chloroplast DNA synthesis during the cell cycle in cultured cells of Nicotiana tabacum: inhibition by nalidixic acid and hydroxyurea

The effects of nalidixic acid and hydroxyurea on nuclear and chloroplast DNA formation in cultured cells of Nicotiana tabacum were investigated. At low concentrations (5 and 20 micrograms/ml) nalidixic acid, an inhibitor of DNA gyrase, exhibited a greater inhibitory effect on plastid DNA synthesis than on nuclear DNA formation. Since the plastid genome is a circular double-stranded DNA, this is consistent with the proven involvement of a DNA gyrase in the replication of closed circular duplex DNA genomes in procaryotic cells. At a high concentration of nalidixic acid (50 micrograms/ml), DNA synthesis in both the plastid and nuclear compartment was rapidly inhibited. Removal of the drug from the culture medium led to the resumption of DNA synthesis in 8 h. Hydroxyurea, an inhibitor of ribonucleoside diphosphate reductase, also depresses nuclear as well as plastid DNA formation. Removal of hydroxyurea from the blocked cells leads to a burst of nuclear DNA synthesis, suggesting that the cells had been synchronized at the G1/S boundary. The recovery of plastid DNA synthesis occurs within the same time frame as that of nuclear DNA. However, whereas plastid DNA formation is then maintained at a constant rate, nuclear DNA synthesis reaches a peak and subsequently declines. These results indicate that the synthesis of plastid DNA is independent of the cell cycle events governing nuclear DNA formation in cultured plant cells.