Stability of Drosophila RNA polymerase II elongation complexes in vitro.
AUTOR(ES)
Kephart, D D
RESUMO
We show that nuclear extract from Drosophila Kc cells supports efficient elongation by RNA polymerase II initiated from the actin 5C promoter. The addition of 0.3% Sarkosyl, 1 mg of heparin per ml, or 250 mM KCl immediately after initiation has two effects. First, the elongation rate is reduced 80 to 90% as a result of the inhibition of elongation factors. Second, there is an increase in the amount of long runoff RNA, suggesting that there is an early block to elongation that is relieved by the disruptive reagents. Consistent with the first effect, we find that the ability of factor 5 (TFIIF) to stimulate the elongation rate is inhibited by the disruptive agents when assayed in a defined system containing pure RNA polymerase II and a dC-tailed template. The disruptive agents also inhibit the ability of DmS-II to suppress transcriptional pausing but only slightly reduce the ability of DmS-II to increase the elongation rate twofold. The pause sites encountered by RNA polymerase II after initiation at a promoter and subsequent treatment with the disruptive reagents are also recognized by pure polymerase transcribing a dC-tailed template. It has been suggested that 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole inhibits RNA polymerase II during elongation, but we find that the purine nucleoside analog has no effect on elongation complexes containing RNA over 500 nucleotides in length or on the action of factor 5 or DmS-II in the defined system.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=364378Documentos Relacionados
- RNA polymerase II ternary transcription complexes generated in vitro.
- Control of formation of two distinct classes of RNA polymerase II elongation complexes.
- Inhibition of T7 and T3 RNA polymerase directed transcription elongation in vitro.
- Nucleosomes inhibit both transcriptional initiation and elongation by RNA polymerase III in vitro.
- A role for TFIIH in controlling the activity of early RNA polymerase II elongation complexes