DNA origami-based single-molecule force spectroscopy elucidates RNA Polymerase III pre-initiation complex stability

Kramm, Kevin; Schröder, Tim; Gouge, Jerome; Vera, Andres Manuel; Gupta, Kapil; Heiss, Florian B.; Liedl, Tim; Engel, Christoph; Berger, Imre; Vannini, Alessandro; Tinnefeld, Philip und Grohmann, Dina (2020): DNA origami-based single-molecule force spectroscopy elucidates RNA Polymerase III pre-initiation complex stability. In: Nature Communications, Bd. 11, Nr. 1

Abstract

The TATA-binding protein (TBP) and a transcription factor (TF) IIB-like factor are important constituents of all eukaryotic initiation complexes. The reason for the emergence and strict requirement of the additional initiation factor Bdp1 in the RNA polymerase (RNAP) III system, however, remained elusive. A poorly studied aspect in this context is the effect of DNA strain arising from DNA compaction and transcriptional activity on initiation complex formation. We made use of a DNA origami-based force clamp to follow the assembly of human initiation complexes in the RNAP II and RNAP III systems at the single-molecule level under piconewton forces. We demonstrate that TBP-DNA complexes are force-sensitive and TFIIB is sufficient to stabilise TBP on a strained promoter. In contrast, Bdp1 is the pivotal component that ensures stable anchoring of initiation factors, and thus the polymerase itself, in the RNAP III system. Thereby, we offer an explanation for the crucial role of Bdp1 for the high transcriptional output of RNAP III. TATA-binding protein (TBP) and a transcription factor (TF) IIB-like factor are important constituents of all eukaryotic initiation complexes. Here, the authors use a DNA origami-based force clamp to investigate the assembly dynamics of human initiation complexes in the RNAP II and RNAP III systems at the single-molecule level under pico newton forces.

• Dokumententyp: Zeitschriftenartikel
• Fakultätsübergreifende Einrichtungen: Center for NanoScience (CENS)
• Themengebiete: 500 Naturwissenschaften und Mathematik > 500 Naturwissenschaften
• ISSN: 2041-1723
• Sprache: Englisch