DNA Topoisomerases Are Required for Preinitiation Complex Assembly during GAL Gene Activation

Roedgaard, Morten; Fredsøe, Jacob; Pedersen, Jakob Madsen; Bjergbæk, Lotte; Andersen, Anders H.

doi:10.1371/journal.pone.0132739

Cited by 13 publications

(12 citation statements)

References 40 publications

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“…To compare absolute transcription levels, we measured the number of nascent RNA molecules at the transcription site in top1 Δ CoD and HO strains using smFISH. We found a small decrease in the number of RNAs in top1 Δ strains in both orientations compared to WT cells (Figure 6D ) in accordance with the effect of topoisomerases on transcription in yeast ( 40 ).…”

Section: Resultssupporting

confidence: 81%

Transcription-replication coordination revealed in single live cells

Tsirkas

Dovrat

Thangaraj

et al. 2022

Nucleic Acids Research

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The coexistence of DNA replication and transcription during S-phase requires their tight coordination to prevent harmful conflicts. While extensive research revealed important mechanisms for minimizing these conflicts and their consequences, little is known regarding how the replication and transcription machinery are coordinated in real-time. Here, we developed a live-cell imaging approach for the real-time monitoring of replisome progression and transcription dynamics during a transcription-replication encounter. We found a wave of partial transcriptional repression ahead of the moving replication fork, which may contribute to efficient fork progression through the transcribed gene. Real-time detection of conflicts revealed their negative impact on both processes, leading to fork stalling or slowdown as well as lower transcription levels during gene replication, with different trade-offs observed in defined subpopulations of cells. Our real-time measurements of transcription-replication encounters demonstrate how these processes can proceed simultaneously while maintaining genomic stability, and how conflicts can arise when coordination is impaired.

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Section: Resultssupporting

confidence: 81%

Transcription-replication coordination revealed in single live cells

Tsirkas

Dovrat

Thangaraj

et al. 2022

Nucleic Acids Research

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“…Consistently, inhibition of DNA topoisomerase I activity reduces the density of RNA polymerase II at promoter pause sites in human cells (Khobta et al, 2006). In budding yeast, DNA topoisomerases are required for the recruitment of RNA polymerase II but are dispensable for ongoing transcriptional elongation (Roedgaard et al, 2015). These observations demonstrate a pivotal role of DNA topoisomerase I in transcriptional initiation.…”

Section: Discussionmentioning

confidence: 67%

DNA Topoisomerase Iα Affects the Floral Transition

et al. 2016

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ORCID ID: 0000-0002-9778-8855 (H.Y.).DNA topoisomerases modulate DNA topology to maintain chromosome superstructure and genome integrity, which is indispensable for DNA replication and RNA transcription. Their function in plant development still remains largely unknown. Here, we report a hitherto unidentified role of Topoisomerase Ia (TOP1a) in controlling flowering time in Arabidopsis (Arabidopsis thaliana). Loss of function of TOP1a results in early flowering under both long and short days. This is attributed mainly to a decrease in the expression of a central flowering repressor, FLOWERING LOCUS C (FLC), and its close homologs, MADS AFFECTING FLOWERING4 (MAF4) and MAF5, during the floral transition. TOP1a physically binds to the genomic regions of FLC, MAF4, and MAF5 and promotes the association of RNA polymerase II complexes to their transcriptional start sites. These correlate with the changes in histone modifications but do not directly affect nucleosome occupancy at these loci. Our results suggest that TOP1a mediates DNA topology to facilitate the recruitment of RNA polymerase II at FLC, MAF4, and MAF5 in conjunction with histone modifications, thus facilitating the expression of these key flowering repressors to prevent precocious flowering in Arabidopsis.

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“…Addressing the possible role of Top2 (or Top3) directly would not be trivial since these enzymes are essential for growth in budding yeast ( 109 , 110 ), have multiple roles in transcriptional regulation ( 15 ), and any break made by Top2 before the transfer of a conditional mutant to restrictive conditions is expected to persist even after the enzymes are down-regulated, inhibited, subjected to heat-reversal, or upon inactivation ( 111–114 ). The findings that in the bulk of gDNA Top2 contributes to the assembly of the RNAP II preinitiation complex ( 102 , 103 ), whereas the depletion of Top2 appears to unslilence RNAP II related ncRNA transcription in rDNA ( 77 ), fit the correlation between nick incidence and RNAP II promoter activity in gDNA on the one hand, and between nicking in rDNA and rDNA silencing on the other.…”

Section: Discussionmentioning

confidence: 80%

“…The promoter-proximal association of Top2 has been documented in ( 102 , 103 , 118 ). If Top2 was responsible for the generation of the nicks observed in our assays, stochasticity could arise in two ways: (i) the active promoters transiently interact with a limited pool of enzymes in a competitive manner, or (ii) each active promoter is permanently Top2-associated, but the enzyme at any particular promoter spends a small part of its catalytic cycle in the Top2cc state, when the relative duration of this time window would determine the average distance of nicks present in the DNA at any given time point.…”

Section: Discussionmentioning

confidence: 99%

Endogenous single-strand DNA breaks at RNA polymerase II promoters in Saccharomyces cerevisiae

Hegedüs

Kókai

Nánási

et al. 2018

Nucleic Acids Research

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Molecular combing and gel electrophoretic studies revealed endogenous nicks with free 3′OH ends at ∼100 kb intervals in the genomic DNA (gDNA) of unperturbed and G1-synchronized Saccharomyces cerevisiae cells. Analysis of the distribution of endogenous nicks by Nick ChIP-chip indicated that these breaks accumulated at active RNA polymerase II (RNAP II) promoters, reminiscent of the promoter-proximal transient DNA breaks of higher eukaryotes. Similar periodicity of endogenous nicks was found within the ribosomal rDNA cluster, involving every ∼10th of the tandemly repeated 9.1 kb units of identical sequence. Nicks were mapped by Southern blotting to a few narrow regions within the affected units. Three of them were overlapping the RNAP II promoters, while the ARS-containing IGS2 region was spared of nicks. By using a highly sensitive reverse-Southwestern blot method to map free DNA ends with 3′OH, nicks were shown to be distinct from other known rDNA breaks and linked to the regulation of rDNA silencing. Nicks in rDNA and the rest of the genome were typically found at the ends of combed DNA molecules, occasionally together with R-loops, comprising a major pool of vulnerable sites that are connected with transcriptional regulation.

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DNA Topoisomerases Are Required for Preinitiation Complex Assembly during GAL Gene Activation

Cited by 13 publications

References 40 publications

Transcription-replication coordination revealed in single live cells

Transcription-replication coordination revealed in single live cells

DNA Topoisomerase Iα Affects the Floral Transition

Endogenous single-strand DNA breaks at RNA polymerase II promoters in Saccharomyces cerevisiae

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