DNA is more negatively supercoiled in bacterial plasmids than in minichromosomes isolated from budding yeast.
A series of circular shuttle vectors were constructed that could replicate and transcribe in the cells of both Escherichia coli and Saccharomyces cerevisiae. 2-D agarose gel electrophoresis run without or in the presence of different concentrations of chloroquine (CHL) revealed that bacterial plasmids were more negatively (-) supercoiled than minichromosomes isolated from budding yeast. Attempts to increase (-) supercoiling in S. cerevisiae or to reduce it in E. coli have deleterious biological consequences. These observations indicate that DNA supercoiling can vary in different species but cells are exquisitely sensitive to sudden changes in supercoiling. In E. coli, the observation that cell growth as well as ColE1 plasmid copy number decrease when DNA relaxes suggests that supercoiling could affect cell viability by regulating the initiation of both transcription and replication.
Mayán-Santos
María-Dolores
MD
Departamento de Biología Celular y del Desarrollo, Centro de Investigaciones Biológicas (CSIC), Madrid, Spain.
Martínez-Robles
María-Luisa
ML
Hernández
Pablo
P
Krimer
Dora
D
Schvartzman
Jorge B
JB
D002470:Cell Survival / Q000187:drug effects; D002738:Chloroquine / Q000737:chemistry* / Q000494:pharmacology; D004261:DNA Replication / Q000187:drug effects; D004269:DNA, Bacterial / Q000737:chemistry*; D004271:DNA, Fungal / Q000737:chemistry*; D004278:DNA, Superhelical / Q000737:chemistry*; D004587:Electrophoresis, Agar Gel / Q000379:methods; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D004926:Escherichia coli / Q000235:genetics*; D010957:Plasmids; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D014158:Transcription, Genetic
D004269:DNA, Bacterial; D004271:DNA, Fungal; D004278:DNA, Superhelical; D002738:Chloroquine