Molecular Topology of RNA Polymerase I Upstream Activation Factor

Knutson, Bruce A.; Smith, Marissa L.; Belkevich, Alana E.; Fakhouri, Aula M.

doi:10.1128/mcb.00056-20

Cited by 10 publications

(13 citation statements)

References 62 publications

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“…1D ). This architecture is consistent with previous predictions ( 19 , 20 ) and is reminiscent of transcription factor complex TFIID and transcriptional coactivator SAGA, which feature histone-like octamers and hexamers and perform a parallel function in support of Pol II PIC assembly (fig. S3C).…”

Section: Resultssupporting

confidence: 92%

Mechanism of RNA polymerase I selection by transcription factor UAF

et al. 2022

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Preribosomal RNA is selectively transcribed by RNA polymerase (Pol) I in eukaryotes. The yeast transcription factor upstream activating factor (UAF) represses Pol II transcription and mediates Pol I preinitiation complex (PIC) formation at the 35 S ribosomal RNA gene. To visualize the molecular intermediates toward PIC formation, we determined the structure of UAF in complex with native promoter DNA and transcription factor TATA-box-binding protein (TBP). We found that UAF recognizes DNA using a hexameric histone-like scaffold with markedly different interactions compared with the nucleosome and the histone-fold-rich transcription factor IID (TFIID). In parallel, UAF positions TBP for Core Factor binding, which leads to Pol I recruitment, while sequestering it from DNA and Pol II/III–specific transcription factors. Our work thus reveals the structural basis of RNA Pol selection by a transcription factor.

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Section: Resultssupporting

confidence: 92%

Mechanism of RNA polymerase I selection by transcription factor UAF

et al. 2022

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“…These folds combine with two H3 and one H4 to form a hexameric histone-like core within UAF (Figure 1D). This architecture is consistent with previous predictions (Knutson et al, 2020; Smith et al, 2018) and is reminiscent of transcription factor complex TFIID and transcriptional coactivator SAGA, which feature histone-like octamers and hexamers and perform a parallel function in support of Pol II PIC assembly (Figure S3C). Similar to TFIID and SAGA, nonhistone-like elements decorate the UAF histone-like core to enable specific protein-protein interactions.…”

Section: Resultssupporting

confidence: 92%

“…Completing the assembly, Uaf30 joins Rrn5 at the upstream end of the Rrn5-H3-H4 tetramer. Three helices of the predicted N-terminal winged helix domain contact Rrn5, consistent with previous observations that the domain interfaces with other UAF subunits (Knutson et al, 2020). The remainder of Uaf30 is unresolved, pointing to an inherent flexibility of the protein.…”

Section: Resultssupporting

confidence: 90%

“…Native mass spectrometry analyses revealed an unusual stoichiometry featuring two H3 proteins and one H4 protein (Smith et al, 2018). Further modelling based on crosslinking mass spectrometry and domain predictions suggests that subunit Rrn5 interacts with H3 and H4 to form a H3-H4 tetramer-like core that is surrounded by subunits Rrn9, Rrn10 and Uaf30 (Knutson et al, 2020). To elucidate the structure of UAF and understand the macromolecular context in which it functions, we reconstituted a complex between UAF, native promoter DNA and transcription factor TBP, which represents an early intermediate during Pol I PIC assembly (Keener et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

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Mechanism of RNA Polymerase I selection by transcription factor UAF

Baudin¹,

Murciano²,

Fung³

et al. 2022

Preprint

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Pre-ribosomal RNA is selectively transcribed by RNA Polymerase (Pol) I in eukaryotes. The yeast transcription factor Upstream Activating Factor (UAF) represses Pol II transcription and mediates Pol I preinitiation complex (PIC) formation during the early stages of transcription initiation at the 35S ribosomal RNA gene. To unravel the DNA recognition and Pol I selection mechanisms of UAF, we determined the structure of UAF bound to native promoter DNA and transcription factor TBP. We found that UAF recognizes DNA using a hexameric histone-like scaffold with markedly different interactions than the nucleosome and the histone-fold-rich TFIID. UAF strategically sequesters TBP from DNA and Pol II/III-specific factors, and positions it for Core Factor binding, supporting Pol I recruitment. Our findings therefore reveal the molecular basis of Pol I selection for ribosome biogenesis. As well, they reveal an unexpected potential within the histone fold as a motif for specific protein-DNA interactions inside the cell.

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“…Transcription begins at the t ranscription s tart s ite (TSS) and Pol I and Rrn3 are released from the PIC upon transcription initiation. Several structural studies gave new insights into Pol I promoter recognition and melting, and more broadly into transcription initiation by yeast Pol I ( Blattner et al, 2011 ; Engel et al, 2013 , 2016 ; Moreno-Morcillo et al, 2014 ; Neyer et al, 2016 ; Tafur et al, 2016 ; Han et al, 2017 ; Sadian et al, 2017 ; Smith et al, 2018 ; Sadian et al, 2019 ; Tafur et al, 2019 ; Knutson et al, 2020 ). These studies will not be detailed here.…”

Section: Transcription Initiation and Terminationmentioning

confidence: 99%

Coupling Between Production of Ribosomal RNA and Maturation: Just at the Beginning

Azouzi

Jaafar

Dez

et al. 2021

Front. Mol. Biosci.

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Ribosomal RNA (rRNA) production represents the most active transcription in the cell. Synthesis of the large rRNA precursors (35S/47S in yeast/human) is achieved by up to hundreds of RNA polymerase I (Pol I) enzymes simultaneously transcribing a single rRNA gene. In this review, we present recent advances in understanding the coupling between rRNA production and nascent rRNA folding. Mapping of the distribution of Pol I along ribosomal DNA at nucleotide resolution, using either native elongating transcript sequencing (NET-Seq) or crosslinking and analysis of cDNAs (CRAC), revealed frequent Pol I pausing, and CRAC results revealed a direct coupling between pausing and nascent RNA folding. High density of Pol I per gene imposes topological constraints that establish a defined pattern of polymerase distribution along the gene, with a persistent spacing between transcribing enzymes. RNA folding during transcription directly acts as an anti-pausing mechanism, implying that proper folding of the nascent rRNA favors elongation in vivo. Defects in co-transcriptional folding of rRNA are likely to induce Pol I pausing. We propose that premature termination of transcription, at defined positions, can control rRNA production in vivo.

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Molecular Topology of RNA Polymerase I Upstream Activation Factor

Cited by 10 publications

References 62 publications

Mechanism of RNA polymerase I selection by transcription factor UAF

Mechanism of RNA polymerase I selection by transcription factor UAF

Mechanism of RNA Polymerase I selection by transcription factor UAF

Coupling Between Production of Ribosomal RNA and Maturation: Just at the Beginning

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