Abortive transcription of the T7 promoter induced by elsamicin A.
The capacity of two intercalating drugs, actinomycin D and elsamicin A, to act as obstacles to transcript elongation by T7 RNA polymerase was analysed using a transcription assay in vitro. Whereas actinomycin D only delayed the production of full-length ('run-off') transcripts from the T7 promoter, elsamicin A arrested the transcription after a short transcript was made, and full-length transcripts were not obtained. The halt of transcription at the drug binding sites close to the promoter causes transcript arrest and results in the formation of a stable RNA hairpin. Actinomycin D appears to be able to detach from the template, allowing the phage polymerase to read through the pause site. In this way, full-length transcripts were obtained in a time-dependent way. With elsamicin A, a blockage of the transcription was observed which could be followed by the release of the phage RNA polymerase. The abortive transcription induced by elsamicin A might be explained considering that this DNA-binding drug drastically reduces the elongation rate and indirectly interferes with the polymerase during transcription, thus favouring its loose association with the nascent RNA and the inability of the enzyme to translocate along the DNA template. A direct interaction of the drug with the polymerase cannot be ruled out as an alternative explanation for the abortive transcription induced by elsamicin A.
Portugal
J
J
Departamento de Biología Molecular y Celular, Centro de Investigación y Desarrollo, CSIC, Barcelona, Spain.
D000617:Aminoglycosides; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology*; D017123:Bacteriophage T7 / Q000201:enzymology / Q000235:genetics*; D001483:Base Sequence; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D003609:Dactinomycin / Q000494:pharmacology; D007700:Kinetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000187:drug effects* / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics; D013698:Templates, Genetic; D014158:Transcription, Genetic / Q000187:drug effects*; D014764:Viral Proteins
D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D000903:Antibiotics, Antineoplastic; D012367:RNA, Viral; D014764:Viral Proteins; D003609:Dactinomycin; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases; C049904:elsamicin A