Gene expression changes triggered by exposure of Haemophilus influenzae to novobiocin or ciprofloxacin: combined transcription and translation analysis

Abstract

The responses of Haemophilus influenzae to DNA gyrase inhibitors were analyzed at the transcriptional and the translational level. High-density microarrays based on the genomic sequence were used to monitor the expression levels of >80% of the genes in this bacterium. In parallel the proteins were analyzed by two-dimensional electrophoresis. DNA gyrase inhibitors of two different functional classes were used. Novobiocin, as a representative of one class, inhibits the ATPase activity of the enzyme, thereby indirectly changing the degree of DNA supercoiling. Ciprofloxacin, a representative of the second class, obstructs supercoiling by inhibiting the DNA cleavage-resealing reaction. Our results clearly show that different responses can be observed. Treatment with the ATPase inhibitor Novobiocin changed the expression rates of many genes, reflecting the fact that the initiation of transcription for many genes is sensitive to DNA supercoiling. Ciprofloxacin mainly stimulated the expression of DNA repair systems as a response to the DNA damage caused by the stable ternary complexes. In addition, changed expression levels were also observed for some genes coding for proteins either annotated as "unknown function" or "hypothetical" or for proteins not directly involved in DNA topology or repair.

Figure 1

(A) The correlation between the hybridization results obtained from the same RNA but independently reverse transcribed and hybridized. (B) The correlation between the hybridization results obtained from the RNA isolated, reverse transcribed, and hybridized independently. (C) The correlation between the hybridization results obtained from the RNA isolated from different cultures grown under the same conditions. Averaged intensities from 100 to 10,000 units are plotted. (Solid line) the ideal 1:1 ratio; (dashed line) a factor of two; (broken line) a factor of three between the two measurements.