Analysis of the open region of RNA polymerase II transcription complexes in the early phase of elongation.
The RNA polymerase II (pol II) transcription complex undergoes a structural transition around registers 20-25, as indicated by ExoIII footprinting analyses. We have employed a highly purified system to prepare pol II complexes stalled at very precise positions during the initial stage of transcript elongation. Using potassium permanganate we analyzed the open region ('transcription bubble') of complexes stalled between registers 15 and 35. We found that from register 15 up to 25 the transcription bubble expands concomitantly with RNA synthesis. At registers 26 and 27 the bubble has a high tendency to retract at the leading edge. Addition of transcription elongation factor TFIIS re-extends the bubble to the stall site, resulting in complexes competent for transcript elongation. These findings are discussed in the light of the recently determined structures for RNA polymerases.
Fiedler
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U
Department of Physiological Chemistry, University Medical Center Utrecht, Stratenum Building, Universiteitsweg 100, 3584 CG Utrecht, The Netherlands.
Timmers
H T
HT
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D004247:DNA / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018983:DNA Footprinting; D004268:DNA-Binding Proteins / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D046911:Macromolecular Substances; D009154:Mutation / Q000235:genetics; D009690:Nucleic Acid Conformation; D011196:Potassium Permanganate; D021122:Protein Subunits; D012319:RNA Polymerase II / Q000737:chemistry / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics; D035181:TATA-Box Binding Protein; D013698:Templates, Genetic; D035581:Transcription Factor TFIIB; D051758:Transcription Factor TFIIH; D014157:Transcription Factors / Q000378:metabolism; D035165:Transcription Factors, General; D019473:Transcription Factors, TFII; D014158:Transcription, Genetic / Q000187:drug effects / Q000235:genetics*; D035602:Transcriptional Elongation Factors
D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D021122:Protein Subunits; D012333:RNA, Messenger; D035181:TATA-Box Binding Protein; D035581:Transcription Factor TFIIB; D014157:Transcription Factors; D035165:Transcription Factors, General; D019473:Transcription Factors, TFII; D035602:Transcriptional Elongation Factors; C018648:transcription factor S-II; C066644:transcription factor TFIIE; D011196:Potassium Permanganate; D051758:Transcription Factor TFIIH; C066816:3'-O-methylguanosine triphosphate; D006160:Guanosine Triphosphate; D004247:DNA; D012319:RNA Polymerase II; C056604:transcription factor TFIIF