Reporter-ChIP-nexus reveals strong contribution of the Drosophila initiator sequence to RNA polymerase pausing

Shao, Wanqing; Alcantara, Sergio G-M; Zeitlinger, Julia

doi:10.7554/elife.41461

Cited by 23 publications

(36 citation statements)

References 78 publications

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“…Very recently, Shao et al. ( 65 ) reported a correlation of efficient escape into productive elongation in Drosophila with features of the Inr, particularly a preference for G at +2. We also see G as the preferred +2 base, especially for the strongest promoters (Figure 3A ), but we have not ranked the HeLa promoters based on relative efficiency of escape from pausing.…”

Section: Discussionmentioning

confidence: 99%

A unified view of the sequence and functional organization of the human RNA polymerase II promoter

Luse

Parida

Spector

et al. 2020

Nucleic Acids Research

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To better understand human RNA polymerase II (Pol II) promoters in the context of promoter-proximal pausing and local chromatin organization, 5′ and 3′ ends of nascent capped transcripts and the locations of nearby nucleosomes were accurately identified through sequencing at exceptional depth. High-quality visualization tools revealed a preferred sequence that defines over 177 000 core promoters with strengths varying by >10 000-fold. This sequence signature encompasses and better defines the binding site for TFIID and is surprisingly invariant over a wide range of promoter strength. We identified a sequence motif associated with promoter-proximal pausing and demonstrated that cap methylation only begins once transcripts are about 30 nt long. Mapping also revealed a ∼150 bp periodic downstream sequence element (PDE) following the typical pause location, strongly suggestive of a +1 nucleosome positioning element. A nuclear run-off assay utilizing the unique properties of the DNA fragmentation factor (DFF) coupled with sequencing of DFF protected fragments demonstrated that a +1 nucleosome is present downstream of paused Pol II. Our data more clearly define the human Pol II promoter: a TFIID binding site with built-in downstream information directing ubiquitous promoter-proximal pausing and downstream nucleosome location.

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Section: Discussionmentioning

confidence: 99%

A unified view of the sequence and functional organization of the human RNA polymerase II promoter

Luse

Parida

Spector

et al. 2020

Nucleic Acids Research

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“…Core promoter sequence features might help distinguish enhancers from promoters, particularly if RNAPII itself reads a regulatory code during pausing and/or early elongation. For example, RNAPII pausing is sequence dependent 19,42 , and is substantially longer-lived at promoters than enhancers 10 . Stable RNAPII pausing at promoters may provide time to recruit distal regulatory complexes by co-localization with the unstable RNAPII pausing seen at enhancers.…”

Section: Discussionmentioning

confidence: 99%

Transcription imparts architecture, function, and logic to enhancer units

Tippens

Liang

Leung

et al. 2019

Preprint

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Distal enhancers remain one of the least understood regulatory elements with pivotal roles in development and disease. We used massively parallel reporter assays to perform functional comparisons of two leading enhancer models and find that gene-distal transcription start sites (TSSs) are robust predictors of enhancer activity with higher resolution and specificity than histone modifications. We show that active enhancer units are precisely delineated by active TSSs, validate that these boundaries are sufficient to capture enhancer function, and confirm that core promoter sequences are required for this activity. Finally, we assay pairs of adjacent units and find that their cumulative activity is best predicted by the strongest unit within the pair.Synthetic fusions of enhancer units demonstrate that adjacency imposes winner-takes-all logic, revealing a simple design for a maximum-activity filter of enhancer unit outputs. Together, our results define fundamental enhancer units and a principle of non-cooperativity between adjacent units.

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“…After initiation, Pol II transcribes a short stretch of 30-80 nucleotides before pausing 7 . This step is regulated by the TFIID complex, as directly demonstrated in vitro 8 and inferred from the over-representation of TFIID-bound core promoter motifs (INR, DPE, pause button) in highly paused genes [9][10][11][12][13] . Pausing durations are highly variable among genes [14][15][16] .…”

Section: Introductionmentioning

confidence: 73%

Quantitative imaging of transcription in livingDrosophilaembryos reveals the impact of core promoter motifs on promoter state dynamics

Pimmett

Dejean

Fernandez

et al. 2021

Preprint

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Genes are expressed in stochastic transcriptional bursts linked to alternating active and inactive promoter states. A major challenge in transcription is understanding how promoter composition dictates bursting, particularly in multicellular organisms. We investigate two key Drosophila developmental promoter motifs, the TATA box (TATA) and the Initiator (INR). Using live imaging in Drosophila embryos and new computational methods, we demonstrate that bursting occurs on multiple timescales ranging from seconds to minutes. TATA-containing promoters and INR-containing promoters exhibit distinct dynamics, with one or two separate rate-limiting steps respectively. A TATA box is associated with long active states, high rates of polymerase initiation, and short-lived, infrequent inactive states. In contrast, the INR motif leads to two inactive states, one of which relates to promoter-proximal polymerase pausing. Surprisingly, the model suggests pausing is not obligatory, but occurs stochastically for a subset of polymerases. Overall, our results provide a rationale for promoter switching during zygotic genome activation.

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Reporter-ChIP-nexus reveals strong contribution of the Drosophila initiator sequence to RNA polymerase pausing

Cited by 23 publications

References 78 publications

A unified view of the sequence and functional organization of the human RNA polymerase II promoter

A unified view of the sequence and functional organization of the human RNA polymerase II promoter

Transcription imparts architecture, function, and logic to enhancer units

Quantitative imaging of transcription in livingDrosophilaembryos reveals the impact of core promoter motifs on promoter state dynamics

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