Structural visualization of de novo transcription initiation by Saccharomyces cerevisiae RNA polymerase II

Mol Cell. 2022 Feb 3;82(3):660-676.e9. doi: 10.1016/j.molcel.2021.12.020. Epub 2022 Jan 19.

Abstract

Previous structural studies of the initiation-elongation transition of RNA polymerase II (pol II) transcription have relied on the use of synthetic oligonucleotides, often artificially discontinuous to capture pol II in the initiating state. Here, we report multiple structures of initiation complexes converted de novo from a 33-subunit yeast pre-initiation complex (PIC) through catalytic activities and subsequently stalled at different template positions. We determine that PICs in the initially transcribing complex (ITC) can synthesize a transcript of ∼26 nucleotides before transitioning to an elongation complex (EC) as determined by the loss of general transcription factors (GTFs). Unexpectedly, transition to an EC was greatly accelerated when an ITC encountered a downstream EC stalled at promoter proximal regions and resulted in a collided head-to-end dimeric EC complex. Our structural analysis reveals a dynamic state of TFIIH, the largest of GTFs, in PIC/ITC with distinct functional consequences at multiple steps on the pathway to elongation.

Keywords: RNA polymerase II; TFIIH; elongation complex; initially transcribing complex; pre-initiation complex; promoter escape; promoter opening; start-site scanning; transcription initiation; yeast.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Video-Audio Media

MeSH terms

  • Cryoelectron Microscopy
  • Gene Expression Regulation, Fungal
  • Models, Molecular
  • Promoter Regions, Genetic
  • Protein Conformation
  • RNA Polymerase II / genetics
  • RNA Polymerase II / metabolism*
  • RNA Polymerase II / ultrastructure
  • Saccharomyces cerevisiae / enzymology*
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / ultrastructure
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Saccharomyces cerevisiae Proteins / ultrastructure
  • Structure-Activity Relationship
  • Time Factors
  • Transcription Elongation, Genetic
  • Transcription Factors, TFII / genetics
  • Transcription Factors, TFII / metabolism
  • Transcription Initiation, Genetic*

Substances

  • Saccharomyces cerevisiae Proteins
  • TFB3 protein, S cerevisiae
  • Transcription Factors, TFII
  • RNA Polymerase II
  • RPB4 protein, S cerevisiae
  • RPB7 protein, S cerevisiae