AbstractThe intracellular level of DNA supercoiling is regulated inEscherichia coliby a homeostatic control mechanism that includes DNA gyrase and topoisomerase I gene expression. Despite several biochemical and genetical evidence that supports the existence of a homeostatic regulation mechanism, there are only few studies focusinggyrA andgyrBgene expression in connection to the mechanism involved in the regulation of DNA supercoilingin vivo. To study DNA gyrase gene expression and to be able to isolate mutants with altered expression of DNA gyrase, we constructed a new chromosomal reporter system based on two translational fusions ofgyrA andgyrBtolacZ. Using this stable monitor system in a robust wild type, we simultaneously studied the influence of several inhibitors of DNA gyrase (quinolones and coumarins) ongyrA andgyrBgene expression as well as on the intracellular level of DNA supercoiling. Surprisingly, we found a delayed and differential response ofgyrA andgyrBgene expression following inhibition of DNA gyrase by quinolones or coumarins. Whereas both groups of drugs were able to increase the expression ofgyrA, thegyrBgene expression was only induced by the coumarins. Although the action of the quinolones was able to alter DNA supercoiling, we never observed any induction ofgyrBfrom the chromosome. These results revealed that the gene expression ofgyrA appears to be more sensitive to alterations in DNA supercoiling than thegyrBgene expression and suggest that probably additional regulatory mechanisms on the post‐translational level might be involved in the regulation of DNA supercoiling and DNA gyrase gene expressio