Ribosomal insertions in Drosophila are transcribed at very low levels. The abundance of the most prominent 0.8 kb type 1 insertion transcript increased up to 60-fold when cultured cells were exposed to the DNA intercalating drug chloroquine. After injection of insertion-containing rDNA in circular form into Xenopus laevis oocytes an apparently identical 0.8 kb insertion transcript was synthesized, and its accumulation was stimulated several fold by coinjection of chloroquine or ethidium bromide. We suggest that ribosomal insertions are assembled in a chromatin conformation that lacks unconstrained torsional stress, accounting for the inactivity of these DNA regions; introduction of stress by intercalation results in activation of transcription from the insertion sequences.