Structural basis of RNA polymerase I pre-initiation complex formation and promoter melting.

Nature Communications
Michael Pilsl, Christoph Engel

Abstract

Transcription of the ribosomal RNA precursor by RNA polymerase (Pol) I is a prerequisite for the biosynthesis of ribosomes in eukaryotes. Compared to Pols II and III, the mechanisms underlying promoter recognition, initiation complex formation and DNA melting by Pol I substantially diverge. Here, we report the high-resolution cryo-EM reconstruction of a Pol I early initiation intermediate assembled on a double-stranded promoter scaffold that prevents the establishment of downstream DNA contacts. Our analyses demonstrate how efficient promoter-backbone interaction is achieved by combined re-arrangements of flexible regions in the 'core factor' subunits Rrn7 and Rrn11. Furthermore, structure-function analysis illustrates how destabilization of the melted DNA region correlates with contraction of the polymerase cleft upon transcription activation, thereby combining promoter recruitment with DNA-melting. This suggests that molecular mechanisms and structural features of Pol I initiation have co-evolved to support the efficient melting, initial transcription and promoter clearance required for high-level rRNA synthesis.

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Citations

Sep 12, 2020·Biochemical Society Transactions·Bruce A KnutsonLawrence I Rothblum
Feb 5, 2021·Nature Communications·Florian B HeissChristoph Engel
Dec 19, 2020·Nature Communications·Ewan Phillip RamsayAlessandro Vannini
Jun 1, 2021·Frontiers in Molecular Biosciences·Tomasz W Turowski, Magdalena Boguta
Jun 17, 2021·Nucleic Acids Research·Cathia RauschM Cristina Cardoso

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Methods Mentioned

BETA
size exclusion chromatography
in vitro transcription
in
electrophoresis

Software Mentioned

Gctf
Phenix
PyMOL
MolProbity
MotionCor2
serialEM
RELION

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