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TitleStructural and kinetic insights into tRNA promoter engagement by yeast general transcription factor TFIIIC.
Journal, issue, pagesNucleic Acids Res, Vol. 53, Issue 1, Year 2025
Publish dateJan 7, 2025
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 / PubMed Central
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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