Transcription-driven site-specific DNA recombination in vitro.
Transcription of a topologically relaxed, circular DNA triggers recombination between two directly repeated res sites by gamma delta resolvase in vitro. This activation of recombination depends on the res site-to-site distance and the orientation of sites with respect to the direction of RNA polymerase tracking. In addition to functioning as a site-specific recombinase, gamma delta resolvase acts as a site-specific topoisomerase and increases the topological linking number of templates during transcription. The data suggest that the link between transcription and recombination could be negative DNA supercoiling that transiently builds up on a relatively short DNA segment in the wake of an advancing RNA polymerase. Surprisingly, transcription-driven recombination is not inhibited by the presence of large amounts of eukaryotic topoisomerase type I, indicating that site-specific recombination can override relaxation by diffusible topoisomerases. This in vitro system might therefore serve as a model for some transcription-directed recombination events observed in vivo.
Dröge
P
P
Department of Biology, University of Konstanz, Germany.
D000818:Animals; D017123:Bacteriophage T7 / Q000201:enzymology / Q000235:genetics; D002417:Cattle; D003568:Cytidine Monophosphate / Q000378:metabolism; D004247:DNA / Q000235:genetics; D004264:DNA Topoisomerases, Type I / Q000378:metabolism*; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D007700:Kinetics; D009713:Nucleotidyltransferases / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011995:Recombination, Genetic; D015183:Restriction Mapping; D013698:Templates, Genetic; D013950:Thymus Gland / Q000201:enzymology; D014158:Transcription, Genetic; D019895:Transposases
D010761:Phosphorus Radioisotopes; D004247:DNA; D009713:Nucleotidyltransferases; D019895:Transposases; D012321:DNA-Directed RNA Polymerases; D004264:DNA Topoisomerases, Type I; D003568:Cytidine Monophosphate