Details

Autor(en) / Beteiligte

• Krebs, Arnaud R.
• Imanci, Dilek
• Hoerner, Leslie
• Gaidatzis, Dimos
• Burger, Lukas
• Schübeler, Dirk

Titel

Genome-wide Single-Molecule Footprinting Reveals High RNA Polymerase II Turnover at Paused Promoters

Ist Teil von

• Molecular cell, 2017-08, Vol.67 (3), p.411-422.e4

Ort / Verlag

United States: Elsevier Inc

Erscheinungsjahr

2017

Quelle

MEDLINE

Beschreibungen/Notizen

• Transcription initiation entails chromatin opening followed by pre-initiation complex formation and RNA polymerase II recruitment. Subsequent polymerase elongation requires additional signals, resulting in increased residence time downstream of the start site, a phenomenon referred to as pausing. Here, we harnessed single-molecule footprinting to quantify distinct steps of initiation in vivo throughout the Drosophila genome. This identifies the impact of promoter structure on initiation dynamics in relation to nucleosomal occupancy. Additionally, perturbation of transcriptional initiation reveals an unexpectedly high turnover of polymerases at paused promoters—an observation confirmed at the level of nascent RNAs. These observations argue that absence of elongation is largely caused by premature termination rather than by stable polymerase stalling. In support of this non-processive model, we observe that induction of the paused heat shock promoter depends on continuous initiation. Our study provides a framework to quantify protein binding at single-molecule resolution and refines concepts of transcriptional pausing. [Display omitted] •Genome-wide detection of protein-DNA contacts at single-molecule resolution•Simultaneous quantification of several transcription initiation intermediates•TATA box increases stability of the pre-initiation complex at promoters•High levels of polymerase turnover at the promoters of paused genes Krebs et al. present a methodology to probe protein binding to the genome at the resolution of single DNA molecules, enabling disentanglement of binding heterogeneity at the promoter of genes, thereby revealing unexpected dynamics of RNA polymerase II at paused genes in Drosophila.