Gyrase-dependent stabilization of pSC101 plasmid inheritance by transcriptionally active promoters.
The pSC101 plasmid encodes a cis-acting genetic locus termed par that ensures the stable inheritance of plasmids in a population of dividing cells. In the absence of selection, par-defective plasmids are lost rapidly from the bacterial population. We report here that the stability of par-deleted pSC101 derivatives is restored by introducing certain adventitious bacterial promoters onto the plasmid. Stabilization requires active transcription from the inserted promoter and is affected by the site and orientation of the insertion, the length of the nascent transcript and DNA gyrase activity. While a promotor-associated overall increase in negative superhelicity of plasmid DNA was observed, stabilized inheritance appeared to be dependent on localized rather than generalized supercoiling. Our demonstration that promoter-induced DNA supercoiling can mimic the effects of the pSC101 par locus provides evidence that the previously reported superhelicity-generating effects of par are intrinsic to its function.
Beaucage
S L
SL
Department of Genetics, Stanford University School of Medicine, CA 94305.
Miller
C A
CA
Cohen
S N
SN
D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D004278:DNA, Superhelical / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D004926:Escherichia coli / Q000235:genetics; D010641:Phenotype; D010957:Plasmids; D011401:Promoter Regions, Genetic; D015183:Restriction Mapping; D014158:Transcription, Genetic
D004278:DNA, Superhelical; D004250:DNA Topoisomerases, Type II