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DNA transcription and repressor binding affect deletion formation in Escherichia coli plasmids.
Author(s) -
Vilette D.,
Uzest M.,
Ehrlich S.D.,
Michel B.
Publication year - 1992
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1002/j.1460-2075.1992.tb05447.x
Subject(s) - biology , plasmid , transcription (linguistics) , pbr322 , microbiology and biotechnology , repressor , dna replication , genetics , lac repressor , origin of replication , terminator (solar) , dna , gene , transcription factor , lac operon , ionosphere , linguistics , philosophy , physics , astronomy
Chimeric plasmids containing phage M13 and plasmid pBR322 sequences undergo deletions in Escherichia coli with a high frequency. In all plasmids one deletion endpoint is the M13 replication origin nick site. We examined the effects of transcription on the position of the other deletion end‐point, by inserting in the plasmids an inducible promoter followed by a transcription terminator. Transcription dramatically affected deletions in an orientation‐dependent way, such that greater than 95% of end‐points were localized downstream from the inserted promoter when it faced the major plasmid transcripts. The end‐points were not constrained to the transcribed region and were not affected by the orientation of pBR322 DNA replication. We propose that deletion events occur preferentially in a plasmid domain which is rendered positively supercoiled by convergent transcription. We also show that interaction of LacI repressor with the cognate operator generates a localized deletion hot spot. This hot spot is dependent on pBR322 replication, and therefore probably acts by arresting progression of DNA replication.