Transcription-induced deletions in Escherichia coli plasmids

Characterization of functions that render DNA susceptible to rearrangement is important for a better understanding of genome instability. In a previous work, we showed that sequences located downstream of a strong promoter are particularly prone to deletion. In this paper, the parameters that influence transcription-induced deletions were studied. pBR322 derivatives carrying the M13 (+) replication origin and a PTac-dependent transcription region were used. Deletion formation was analysed in the presence of the replication protein of M13, which introduces a nick at the phage replication origin, and in a rep- strain to avoid M13-driven replication. Our study showed that: (i) 4 h after induction of transcription, a few per cent of the plasmids have experienced a deletion; (ii) these deletions result in joining of the M13 replication origin to a nucleotide located in or downstream of the transcribed region; (iii) deletion formation strongly depends on the orientation of transcription, on promoter strength and transcript length, but is independent of translation; (iv) formation of transcription-induced supercoiling domains does not induce deletion formation. We propose that deletions in the transcribed region result from collisions between converging replication and transcription machineries.