scholarly journals Transcription-driven Chromatin Repression of Intragenic Promoters

2018 ◽  
Author(s):  
Mathias Nielsen ◽  
Ryan Ard ◽  
Xueyuan Leng ◽  
Maxim Ivanov ◽  
Peter Kindgren ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Positional Information ◽  
Gene Promoters ◽  
Molecular Signatures ◽  
Transcription Start Sites ◽  
Histone 3 ◽  
Isoform Diversity ◽  
Genome Wide ◽  
Chaperone Complex ◽  
Rnapii Transcription

SummaryProgression of RNA polymerase II (RNAPII) transcription relies on the appropriately positioned activities of elongation factors. The resulting profile of factors and chromatin signatures along transcription units provides a “positional information system” for transcribing RNAPII. Here, we investigate a chromatin-based mechanism that suppresses intragenic initiation of RNAPII transcription. We demonstrate that RNAPII transcription across gene promoters represses their function in plants. This repression is characterized by reduced promoter-specific molecular signatures and increased molecular signatures associated with RNAPII elongation. The FACT histone chaperone complex is required for this repression mechanism. Genome-wide mapping of Transcription Start Sites (TSSs) reveals thousands of discrete intragenic TSS positions in FACT mutants. Histone 3 lysine 4 mono-methylation poises exonic sites to initiate RNAPII transcription in FACT mutants. Uncovering the mechanism for intragenic TSS repression through the act of RNAPII elongation has important implications for understanding pervasive RNAPII transcription and the regulation of transcript isoform diversity.

scholarly journals Antisense-mediated repression of SAGA-dependent genes involves the HIR histone chaperone

2021 ◽  
Author(s):  
Julien Soudet ◽  
Nissrine Beyrouthy ◽  
Anna Marta Pastucha ◽  
Andrea Maffioletti ◽  
Zahra Bakir ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
De Novo ◽  
Antisense Transcription ◽  
Histone Chaperone ◽  
Gene Promoters ◽  
Genome Wide ◽  
Chaperone Complex ◽  
Histone Deposition ◽  
Transcription Interference

Eukaryotic genomes are pervasively transcribed by RNA polymerase II (RNAPII), and transcription of long non-coding RNAs often overlaps with coding gene promoters. This might lead to coding gene repression in a process named Transcription Interference (TI). In Saccharomyces cerevisiae (S. cerevisiae), TI is mainly driven by antisense non-coding transcription and occurs through re-shaping of promoter Nucleosome-Depleted Regions (NDRs). In this study, we developed a genetic screen to identify new players involved in Antisense-Mediated Transcription Interference (AMTI). Among the candidates, we found the HIR histone chaperone complex known to be involved in de novo histone deposition. Using genome-wide approaches, we reveal that HIR-dependent histone deposition represses the promoters of SAGA-dependent genes via antisense non-coding transcription. However, while antisense transcription is enriched at promoters of SAGA-dependent genes, this feature is not sufficient to define the mode of gene regulation. We further show that the balance between HIR-dependent nucleosome incorporation and transcription factor binding at promoters directs transcription into a SAGA- or TFIID-dependent regulation. This study sheds light on a new connection between antisense non-coding transcription and the nature of coding transcription initiation.

scholarly journals Hda1C restricts the transcription initiation frequency to limit divergent non-coding RNA transcription

2021 ◽  
Author(s):  
Uthra Gowthaman ◽  
Maxim Ivanov ◽  
Isabel Schwarz ◽  
Heta P. Patel ◽  
Niels A. Müller ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Histone Deacetylation ◽  
Gene Promoters ◽  
Genetic Screens ◽  
Fluorescence Measurements ◽  
Non Coding Rna ◽  
Genome Wide ◽  
A Genome ◽  
Initiation Frequency

ABSTRACTNucleosome-depleted regions (NDRs) at gene promoters support initiation of RNA Polymerase II transcription. Interestingly, transcription often initiates in both directions, resulting in an mRNA, and a divergent non-coding (DNC) transcript with an unclear purpose. Here, we characterized the genetic architecture and molecular mechanism of DNC transcription in budding yeast. We identified the Hda1 histone deacetylase complex (Hda1C) as a repressor of DNC in high-throughput reverse genetic screens based on quantitative single-cell fluorescence measurements. Nascent transcription profiling showed a genome-wide role of Hda1C in DNC repression. Live-cell imaging of transcription revealed that Hda1C reduced the frequency of DNC transcription. Hda1C contributed to decreased acetylation of histone H3 in DNC regions, supporting DNC repression by histone deacetylation. Our data support the interpretation that DNC results as a consequence of the NDR-based architecture of eukaryotic promoters, but that it is governed by locus-specific repression to maintain genome fidelity.

scholarly journals RNA polymerase II-independent recruitment of SPT6L at transcription start sites in Arabidopsis

2019 ◽  
Vol 47 (13) ◽  
pp. 6714-6725 ◽  
Author(s):  
Chen Chen ◽  
Jie Shu ◽  
Chenlong Li ◽  
Raj K Thapa ◽  
Vi Nguyen ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Elongation Factor ◽  
Proper Function ◽  
Gene Body ◽  
Transcription Start ◽  
Transcription Start Sites ◽  
Genome Wide ◽  
Yeast Mutants

Abstract SPT6 is a conserved elongation factor that is associated with phosphorylated RNA polymerase II (RNAPII) during transcription. Recent transcriptome analysis in yeast mutants revealed its potential role in the control of transcription initiation at genic promoters. However, the mechanism by which this is achieved and how this is linked to elongation remains to be elucidated. Here, we present the genome-wide occupancy of Arabidopsis SPT6-like (SPT6L) and demonstrate its conserved role in facilitating RNAPII occupancy across transcribed genes. We also further demonstrate that SPT6L enrichment is unexpectedly shifted, from gene body to transcription start site (TSS), when its association with RNAPII is disrupted. Protein domains, required for proper function and enrichment of SPT6L on chromatin, are subsequently identified. Finally, our results suggest that recruitment of SPT6L at TSS is indispensable for its spreading along the gene body during transcription. These findings provide new insights into the mechanisms underlying SPT6L recruitment in transcription and shed light on the coordination between transcription initiation and elongation.

scholarly journals NELF and GAGA Factor Are Linked to Promoter-Proximal Pausing at Many Genes in Drosophila

2008 ◽  
Vol 28 (10) ◽  
pp. 3290-3300 ◽  
Author(s):  
Chanhyo Lee ◽  
Xiaoyong Li ◽  
Aaron Hechmer ◽  
Michael Eisen ◽  
Mark D. Biggin ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Dna Sequences ◽  
Proximal Region ◽  
Gaga Factor ◽  
Pol Ii ◽  
Transcription Start Sites ◽  
Genome Wide ◽  
Highly Expressed Genes ◽  
Inactive Genes ◽  
Chip Chip

ABSTRACT Recent analyses of RNA polymerase II (Pol II) revealed that Pol II is concentrated at the promoters of many active and inactive genes. NELF causes Pol II to pause in the promoter-proximal region of the hsp70 gene in Drosophila melanogaster. In this study, genome-wide location analysis (chromatin immunoprecipitation-microarray chip [ChIP-chip] analysis) revealed that NELF is concentrated at the 5′ ends of 2,111 genes in Drosophila cells. Permanganate genomic footprinting was used to determine if paused Pol II colocalized with NELF. Forty-six of 56 genes with NELF were found to have paused Pol II. Pol II pauses 30 to 50 nucleotides downstream from transcription start sites. Analysis of DNA sequences in the vicinity of paused Pol II identified a conserved DNA sequence that probably associates with TFIID but detected no evidence of RNA secondary structures or other conserved sequences that might directly control elongation. ChIP-chip experiments indicate that GAGA factor associates with 39% of the genes that have NELF. Surprisingly, NELF associates with almost one-half of the most highly expressed genes, indicating that NELF is not necessarily a repressor of gene expression. NELF-associated pausing of Pol II might be an obligatory but sometimes transient checkpoint during the transcription cycle.

scholarly journals Native Elongation Transcript sequencing reveals temperature dependent dynamics of nascent RNAPII transcription in Arabidopsis

2019 ◽  
Author(s):  
Peter Kindgren ◽  
Maxim Ivanov ◽  
Sebastian Marquardt
Keyword(s):  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Molecular Complexes ◽  
Cold Response ◽  
Plant Environment ◽  
Genome Wide ◽  
Predisposing Genes ◽  
Nucleotide Resolution ◽  
Kinetics Of ◽  
Rnapii Transcription

ABSTRACTTemperature profoundly affects the kinetics of biochemical reactions, yet how large molecular complexes such as the transcription machinery accommodate changing temperatures to maintain cellular function is poorly understood. Here, we developed plant native elongating transcripts sequencing (plaNET-seq) to profile genome-wide nascent RNA polymerase II (RNAPII) transcription during the cold-response of Arabidopsis thaliana with single-nucleotide resolution. Combined with temporal resolution, these data revealed transient genome-wide reprogramming of nascent RNAPII transcription during cold, including characteristics of RNAPII elongation and thousands of non-coding transcripts connected to gene expression. Our results suggest a role for promoter-proximal RNAPII stalling in predisposing genes for transcriptional activation during plant-environment interactions. At gene 3’-ends, cold initially facilitated transcriptional termination by limiting the distance of read-through transcription. Within gene bodies, cold reduced the kinetics of co-transcriptional splicing leading to increased intragenic stalling. Our data resolved multiple distinct mechanisms by which temperature transiently altered the dynamics of nascent RNAPII transcription and associated RNA processing, illustrating potential biotechnological solutions and future focus areas to promote food security in the context of a changing climate.

scholarly journals Ligand-induced native G-quadruplex stabilization impairs transcription initiation

2021 ◽  
Author(s):  
Conghui Li ◽  
Honghong Wang ◽  
Zhinang Yin ◽  
Pingping Fang ◽  
Ruijing Xiao ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Gene Transcription ◽  
Transcription Initiation ◽  
Regulatory Elements ◽  
Gene Promoters ◽  
Drug Induced ◽  
Transcriptional Regulatory Elements ◽  
Genome Wide ◽  
Self Association

G-quadruplexes (G4s) are noncanonical DNA secondary structures formed through the self-association of guanines, and G4s are distributed widely across the genome. G4 participates in multiple biological processes including gene transcription, and G4-targeted ligands serve as potential therapeutic agents for DNA-targeted therapies. However, genome-wide studies of the exact roles of G4s in transcriptional regulation are still lacking. Here, we establish a sensitive G4-CUT&Tag method for genome-wide profiling of native G4s with high resolution and specificity. We find that native G4 signals are cell type–specific and are associated with transcriptional regulatory elements carrying active epigenetic modifications. Drug-induced promoter-proximal RNA polymerase II pausing promotes nearby G4 formation. In contrast, G4 stabilization by G4-targeted ligands globally reduces RNA polymerase II occupancy at gene promoters as well as nascent RNA synthesis. Moreover, ligand-induced G4 stabilization modulates chromatin states and impedes transcription initiation via inhibition of general transcription factors loading to promoters. Together, our study reveals a reciprocal genome-wide regulation between native G4 dynamics and gene transcription, which will deepen our understanding of G4 biology toward therapeutically targeting G4s in human diseases.

scholarly journals RNA Polymerase II Independent Recruitment of SPT6 at Transcription Start Sites in Arabidopsis

2018 ◽  
Author(s):  
Chen Chen ◽  
Jie Shu ◽  
Chenlong Li ◽  
Raj K. Thapa ◽  
Vi Nguyen ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Potential Role ◽  
Elongation Factor ◽  
Gene Body ◽  
Transcription Start Sites ◽  
Genome Wide ◽  
Shed Light

SummarySPT6 is a conserved transcription regulator that is generally viewed as an elongation factor. However, emerging evidence show its potential role in the control of transcription initiation at genic and intragenic promoters. Here we first present the genome-wide occupancy of Arabidopsis SPT6-like (SPT6L) and demonstrate its conserved role in facilitating RNA Polymerase II (RNAPII) occupancy across transcribed genes. Further, we show that SPT6L enrichment is shifted, unexpectedly, from gene body to the transcription starting site (TSS) when its association with RNAPII is disrupted. Finally, we demonstrate that recruitment of SPT6L starts at TSS, and then spreads to the gene body during transcription. These findings refine the mechanisms underlying SPT6L recruitment in transcription and shed light on the role of SPT6L in transcription initiation.

scholarly journals Genome-Wide Association of Mediator and RNA Polymerase II in Wild-Type and Mediator Mutant Yeast

2014 ◽  
Vol 35 (1) ◽  
pp. 331-342 ◽  
Author(s):  
Emily Paul ◽  
Z. Iris Zhu ◽  
David Landsman ◽  
Randall H. Morse
Keyword(s):  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Open Reading Frames ◽  
Temperature Sensitive ◽  
Gene Promoters ◽  
Content Type ◽  
Genome Wide ◽  
A Genome ◽  
Wide Range

Mediator is a large, multisubunit complex that is required for essentially all mRNA transcription in eukaryotes. In spite of the importance of Mediator, the range of its targets and how it is recruited to these is not well understood. Previous work showed that inSaccharomyces cerevisiae, Mediator contributes to transcriptional activation by two distinct mechanisms, one depending on the tail module triad and favoring SAGA-regulated genes, and the second occurring independently of the tail module and favoring TFIID-regulated genes. Here, we use chromatin immunoprecipitation sequencing (ChIP-seq) to show that dependence on tail module subunits for Mediator recruitment and polymerase II (Pol II) association occurs preferentially at SAGA-regulated over TFIID-regulated genes on a genome-wide scale. We also show that recruitment of tail module subunits to active gene promoters continues genome-wide when Mediator integrity is compromised inmed17temperature-sensitive (ts) yeast, demonstrating the modular nature of the Mediator complexin vivo. In addition, our data indicate that promoters exhibiting strong and stable occupancy by Mediator have a wide range of activity and are enriched for targets of the Tup1-Cyc8 repressor complex. We also identify a number of strong Mediator occupancy peaks that overlap dubious open reading frames (ORFs) and are likely to include previously unrecognized upstream activator sequences.

scholarly journals Frequent lack of repressive capacity of promoter DNA methylation identified through genome-wide epigenomic manipulation

2017 ◽  
Author(s):  
Ethan Ford ◽  
Matthew R. Grimmer ◽  
Sabine Stolzenburg ◽  
Ozren Bogdanovic ◽  
Alex de Mendoza ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Fusion Protein ◽  
Rna Polymerase Ii ◽  
Zinc Finger ◽  
Gene Promoters ◽  
Natural Conditions ◽  
Promoter Regions ◽  
Genome Wide ◽  
Promoter Dna Methylation ◽  
Promoter Dna

AbstractIt is widely assumed that the addition of DNA methylation at CpG rich gene promoters silences gene transcription. However, this conclusion is largely drawn from the observation that promoter DNA methylation inversely correlates with gene expression in natural conditions. The effect of induced DNA methylation on endogenous promoters has yet to be comprehensively assessed. Here, we induced the simultaneous methylation of thousands of promoters in the genome of human cells using an engineered zinc finger-DNMT3A fusion protein, enabling assessment of the effect of forced DNA methylation upon transcription, histone modifications, and DNA methylation persistence after the removal of the fusion protein. We find that DNA methylation is frequently insufficient to transcriptionally repress promoters. Furthermore, DNA methylation deposited at promoter regions associated with H3K4me3 is rapidly erased after removal of the zinc finger-DNMT3A fusion protein. Finally, we demonstrate that induced DNA methylation can exist simultaneously on promoter nucleosomes that possess the active histone modification H3K4me3, or DNA bound by the initiated form of RNA polymerase II. These findings suggest that promoter DNA methylation is not generally sufficient for transcriptional inactivation, with implications for the emerging field of epigenome engineering.One Sentence SummaryGenome-wide epigenomic manipulation of thousands of human promoters reveals that induced promoter DNA methylation is unstable and frequently does not function as a primary instructive biochemical signal for gene silencing and chromatin reconfiguration.

scholarly journals Role of the pre-initiation complex in Mediator recruitment and dynamics

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Elisabeth R Knoll ◽  
Z Iris Zhu ◽  
Debasish Sarkar ◽  
David Landsman ◽  
Randall H Morse
Keyword(s):  
Rna Polymerase Ii ◽  
Gene Activation ◽  
Mediator Complex ◽  
Gene Promoters ◽  
Initiation Complex ◽  
Yeast Saccharomyces Cerevisiae ◽  
Pol Ii ◽  
Genome Wide ◽  
Cooperative Assembly

The Mediator complex stimulates the cooperative assembly of a pre-initiation complex (PIC) and recruitment of RNA Polymerase II (Pol II) for gene activation. The core Mediator complex is organized into head, middle, and tail modules, and in budding yeast (Saccharomyces cerevisiae), Mediator recruitment has generally been ascribed to sequence-specific activators engaging the tail module triad of Med2-Med3-Med15 at upstream activating sequences (UASs). We show that yeast lacking Med2-Med3-Med15 are viable and that Mediator and PolII are recruited to promoters genome-wide in these cells, albeit at reduced levels. To test whether Mediator might alternatively be recruited via interactions with the PIC, we examined Mediator association genome-wide after depleting PIC components. We found that depletion of Taf1, Rpb3, and TBP profoundly affected Mediator association at active gene promoters, with TBP being critical for transit of Mediator from UAS to promoter, while Pol II and Taf1 stabilize Mediator association at proximal promoters.

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