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TitleStructural and kinetic insights into tRNA promoter engagement by yeast general transcription factor TFIIIC.
Journal, issue, pagesNucleic Acids Res, Year 2024
Publish dateDec 9, 2024
AuthorsWolfram Seifert-Dávila / Anastasiia Chaban / Florence Baudin / Mathias Girbig / Luis Hauptmann / Thomas Hoffmann / Olivier Duss / Sebastian Eustermann / Christoph W Müller /
PubMed AbstractTranscription of transfer RNA (tRNA) genes by RNA polymerase (Pol) III requires the general transcription factor IIIC (TFIIIC), which recognizes intragenic A-box and B-box DNA motifs of type II ...Transcription of transfer RNA (tRNA) genes by RNA polymerase (Pol) III requires the general transcription factor IIIC (TFIIIC), which recognizes intragenic A-box and B-box DNA motifs of type II gene promoters. However, the underlying mechanism has remained elusive, in part due to missing structural information for A-box recognition. In this study, we use single-particle cryogenic electron microscopy (cryo-EM) and single-molecule fluorescence resonance energy transfer (smFRET) to reveal structural and real-time kinetic insights into how the 520-kDa yeast TFIIIC complex engages A-box and B-box DNA motifs in the context of a tRNA gene promoter. Cryo-EM structures of τA and τB subcomplexes bound to the A-box and B-box were obtained at 3.7 and 2.5 Å resolution, respectively, while cryo-EM single-particle mapping determined the specific distance and relative orientation of the τA and τB subcomplexes revealing a fully engaged state of TFIIIC. smFRET experiments show that overall recruitment and residence times of TFIIIC on a tRNA gene are primarily governed by B-box recognition, while footprinting experiments suggest a key role of τA and the A-box in TFIIIB and Pol III recruitment following TFIIIC recognition of type II promoters.
External linksNucleic Acids Res / PubMed:39657784
MethodsEM (single particle)
Resolution2.46 - 3.7 Å
Structure data

51228

9gc3

EMDB-51228, PDB-9gc3:
yeast TFIIIC TauB subcomplex bound to a tRNA gene
Method: EM (single particle) / Resolution: 2.46 Å

51231

9gck

EMDB-51231, PDB-9gck:
yeast TFIIIC TauA subcomplex bound to a tRNA gene
Method: EM (single particle) / Resolution: 3.7 Å

Source
  • saccharomyces cerevisiae (brewer's yeast)
KeywordsDNA BINDING PROTEIN / Base-specific contacts / B-box / tRNA gene / A-box

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