DNA length is a critical parameter for eukaryotic transcription in vivo.
The organization of eukaryotic chromosomes into topological domains has led to the assumption that DNA topology and perhaps supercoiling are involved in eukaryotic nuclear processes. Xenopus oocytes provide a model system for studying the role of DNA topology in transcription. Linear plasmid templates for RNA polymerases (Pols) I and II are not transcribed in Xenopus oocytes, while circular templates are transcriptionally active. Here we show that circularity is not required for transcription of Pol I or Pol II promoters if the linear template is sufficiently long (> 17 to 19 kb). The Xenopus rRNA (Pol I) promoter is active in central positions on a long linear template but is not transcribed when located near an end. Because supercoils generated by transcription could be retained by viscous drag against the long template, these results are consistent with a supercoiling requirement for this promoter. Surprisingly, the herpes simplex virus thymidine kinase (Pol II) promoter is active even 100 bp from the end of the long template, indicating that template length fulfills a critical parameter for transcription that is not consistent with a supercoiling requirement. These results show that DNA length has unrecognized importance for transcription in vivo.
Krebs
J E
JE
Department of Molecular and Cell Biology, University of California, Berkeley 94720-3202, USA.
Dunaway
M
M
D000818:Animals; D001483:Base Sequence; D004278:DNA, Superhelical / Q000737:chemistry / Q000378:metabolism; D005260:Female; D009865:Oocytes / Q000502:physiology; D010957:Plasmids / Q000737:chemistry* / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D012318:RNA Polymerase I / Q000096:biosynthesis*; D012319:RNA Polymerase II / Q000096:biosynthesis*; D015183:Restriction Mapping; D018139:Simplexvirus / Q000201:enzymology / Q000235:genetics; D013698:Templates, Genetic; D013937:Thymidine Kinase / Q000096:biosynthesis* / Q000235:genetics; D014158:Transcription, Genetic; D014982:Xenopus laevis
D004278:DNA, Superhelical; D013937:Thymidine Kinase; D012319:RNA Polymerase II; D012318:RNA Polymerase I