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    DNA origami-based single-molecule forcespectroscopy elucidates RNA Polymerase IIIpre-initiation complex stability

    Kramm, K. and Schroder, T. and Gouge, Jerome and Vera, A.M. and Gupta, K. and Heiss, F.B. and Engel, C. and Berger, I. and Vannini, A. and Tinnefeld, P. and Grohmann, D. (2020) DNA origami-based single-molecule forcespectroscopy elucidates RNA Polymerase IIIpre-initiation complex stability. Nature Communications 11 (2828), ISSN 2041-1723.

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    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.

    Metadata

    Item Type: Article
    Keyword(s) / Subject(s): Biochemistry, Biophysics, Nanoscale biophysics, Single-molecule biophysics, Transcription
    School: Birkbeck Schools and Departments > School of Science > Biological Sciences
    Research Centres and Institutes: Structural Molecular Biology, Institute of (ISMB)
    Depositing User: Jerome Gouge
    Date Deposited: 10 Jun 2020 12:56
    Last Modified: 07 Jul 2020 00:15
    URI: http://eprints.bbk.ac.uk/id/eprint/32200

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