The effects of oxolinic acid and novobiocin -which respectively inhibit the A and B subunits of DNA gyrase, and therefore inhibit DNA synthesis -have been examined in sporulating cultures of Bacillus subtilis. Although both inhibitors prevent sporulation, this is not due to inhibition of DNA synthesis. Instead, they affect protein synthesis generally, though weakly, but have very marked effects on the formation of individual enzymes. These effects are reproducible, but the type of enzyme that will be affected is not predictable. The results point to an involvement of DNA gyrase in the transcription of some genes. This is suggested as the reason for the effect of the inhibitors on spore formation, which they block mainly at Stage 0-1.
I N T R O D U C T I O NThe experiments of Mandelstam et al. (1971) established a relationship between DNA replication and induction of sporulation in Bacillus subtilis. They showed that thymine deprivation in thymine auxotrophs resulted in inhibition of sporulation and of the associated biochemical events such as production of extracellular proteases, ribonuclease and alkaline phosphatase. Later, Dunn et al. (1978) showed that 6-(p-hydroxyphenylazo)uracil (HPUra), a specific inhibitor of DNA polymerase I11 (Gass et al., 1973) stopped sporulation if the inhibitor was added at the time when sporulation was induced, and that the cell population escaped progressively from the effect of the inhibitor as the cells completed chromosome replication.Other inhibitors have recently been shown to have a preferential effect on DNA synthesis, e.g. novobiocin, oxolinic acid and nalidixic acid. All these affect the action of the enzyme DNA gyrase (Gellert et al., 1977), which controls the extent of supercoiling of DNA and which is necessary for DNA replication ( In this paper the effects of DNA gyrase inhibitors on sporulation are compared with those produced by HPUra. The results show that inhibition of sporulation by gyrase inhibitors cannot be accounted for by their effect on DNA synthesis and is more likely to result from an effect on RNA synthesis.
M E T H O D SOrganisms. Bacillus subtilis NiI, a derivative of strain 168trpC2, was used in most of the experiments. It is auxotrophic for thymine and tryptophan and carries a defective PBSX phage in its genome which does not induce lysis of the cells (Karamata, 1968). Bacillus subtilis 168 trpC2 (Marburg) was used for electron microscopy experiments because it sporulates better than strain NiI. It requires tryptophan for growth.