It has been known for a long time that supercoiling can affect gene expression at the level of promoter activity. Moreover, the results of a genome-wide analysis have recently led to the proposal that supercoiling could play a role in the regulation of gene expression at this level by acting as a second messenger, relaying environmental signals to regulatory networks. Although evidence is lacking for a regulatory role of supercoiling following transcription initiation, recent results from both yeast and bacteria suggest that the effect of supercoiling on gene expression can be considerably more dramatic after this initiation step. Transcription-induced supercoiling and its associated R-loops seem to be involved in this effect. In this context, one major function of topoisomerases would be to prevent the generation of excess negative supercoiling by transcription elongation, to inhibit R-loop formation and allow gene expression. This function would be especially evident when substantial and rapid gene expression is required for stress resistance, and it may explain, at least in part, why topoisomerase I synthesis is directed from stress-induced promoters in Escherichia coli. Growth inhibition mediated by excess negative supercoiling might be related to this interplay between transcription elongation and supercoiling.