Abstract
In eukaryotic cells, RNA polymerase I (Pol I) synthesizes precursor ribosomal RNA (pre‐rRNA) that is subsequently processed into mature rRNA. To initiate transcription, Pol I requires the assembly of a multi‐subunit pre‐initiation complex (PIC) at the ribosomal RNA promoter. In yeast, the minimal PIC includes Pol I, the transcription factor Rrn3, and Core Factor (CF) composed of subunits Rrn6, Rrn7, and Rrn11. Here, we present the cryo‐EM structure of the 18‐subunit yeast Pol I PIC bound to a transcription scaffold. The cryo‐EM map reveals an unexpected arrangement of the DNA and CF subunits relative to Pol I. The upstream DNA is positioned differently than in any previous structures of the Pol II PIC. Furthermore, the TFIIB‐related subunit Rrn7 also occupies a different location compared to the Pol II PIC although it uses similar interfaces as TFIIB to contact DNA. Our results show that although general features of eukaryotic transcription initiation are conserved, Pol I and Pol II use them differently in their respective transcription initiation complexes.
Highlights
Biogenesis of ribosomes is dependent on the high transcriptional activity of eukaryotic RNA polymerase I (Pol I)
For initiating rRNA synthesis, yeast Pol I uses a unique set of transcription factors: Upstream Activating Factor (UAF), TATAbinding protein (TBP), Core Factor (CF), and Rrn3 (Keener et al, 1998)
Rrn6 contains a b-propeller followed by a helical domain, while Rrn11 comprises a central tetratricopeptide repeat (TPR) domain preceded by an N-terminal extension (Knutson et al, 2014)
Summary
Biogenesis of ribosomes is dependent on the high transcriptional activity of eukaryotic RNA polymerase I (Pol I). Direct interactions with Pol I are precluded in the Pol I initiation complex since the corresponding position of the N-terminal cyclin domain near the wall in Pol II is occupied by upstream DNA in our model (Fig 3B). Mapping of the electrostatic potential onto the molecular surface of the Pol I PIC (Fig EV3) shows that, in addition to direct DNA contacts formed primarily by the Rrn N-terminal cyclin domain and the DBHB, other regions contribute to DNA binding. In the cryo-EM structure of the Pol I PIC, the b-propeller domain of Rrn is located at the upstream DNA end in a position where it is more likely to engage in interactions with TBPand UAF, but not Rrn (Fig 4A). In the Pol I PIC, the DNA makes several contacts to CF and plausibly influences strongly its conformation
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