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BMH-21 inhibits RNA polymerase I transcription cycle and rDNA occupancy

ABSTRACT: BMH-21 inhibits RNA polymerase I transcription cycle and rDNA occupancy

PROVIDER: PRJNA732709 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR14651178.fastq.gz	Fastqsanger.gz
	SRR14651179.fastq.gz	Fastqsanger.gz
	SRR14651180.fastq.gz	Fastqsanger.gz
	SRR14651181.fastq.gz	Fastqsanger.gz
	SRR14651182.fastq.gz	Fastqsanger.gz

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BMH-21 inhibits RNA polymerase I transcription cycle and rDNA occupancy

Project description:The objective of this study was to determine the effect of a small-molecule Pol I inhibitor, BMH-21, on rRNA synthesis in vivo. NET-seq was performed to determine the Pol I occupancy after BMH-21 treatment, as compared to vehicle-treatment (phosphate buffer control). Our findings suggest that BMH-21 treatment reduces Pol I occupancy on the rDNA template. Additionally, BMH-21 induces repositioning of Pol I in AT-rich rDNA regions that are directly upstream from GC-rich regions. This study suggests that BMH-21 is a powerful inhibitor of transcription by Pol I, and gives a potential mechanism of action for this inhibitor in vivo.

2021-05-26 | GSE175553 | GEO

Decoding the occupancy of RNA polymerase I on rDNA

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Decoding the occupancy of RNA polymerase I on rDNA

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The Rio1 kinase downregulates RNA polymerase I to promote rDNA stability and segregation

Project description:To sustain growth, budding yeast actively transcribes its ribosomal gene array (rDNA) in the nucoleolus to produce ribosomes and proteins. However, intense transcription during rDNA replication may provoke collisions between RNA polymerase I (Pol I) and the replisome, may cause replication fork instability, double-strand breaks, local recombinations and rDNA instability. The latter is manifested by rDNA array expansion or reduction and the formation of extrachromosomal rDNA circles, anomalies that accelerate aging in yeast. Transcription also interferes with the resolution, condensation and segregation of the sister chromatid rDNA arrays. As a consequence, rDNA segregation lags behind the rest of the yeast genome and occurs in late anaphase when rDNA transcription is temporarily shut off. How yeast promotes the stability and transmission of its rDNA array while satisfying a constant need for ribosomes remains unclear. Here we show that the downregulation of Pol I by the conserved cell cycle kinase Rio1 spatiotemporally coordinates rDNA transcription, replication and segregation. More specifically, Rio1 activity promotes copy-number stability of the replicating rDNA array by curtailing Pol I activity and by localising the histone deacetylase Sir2, which establishes a heterochromatic state that silences rDNA transcription. At anaphase entry, Rio1 and the Cdc14 phosphatase target Pol I subunit Rpa43 to dissociate Pol I from the 35S rDNA promoter. The rDNA locus then condensates and segregates, thereby concluding the genome transmission process. Rio1 is involved in ribosome maturation in the cytoplasm of budding yeast and human cells. Additional engagements in the cytoplasm or roles in the nucleus are unknown. Our study describes its first nuclear engagement as a Pol I silencing kinase. This activity may prove highly relevant as dysregulated RNA polymerase I activity has been associated with cancer initiation and proliferation.

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Disruption of cardiac Med1 inhibits RNA polymerase-II promoter occupancy and induces chromatin remodeling

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Project description:The central goal of this project was to determine the role of the RNA polymerase I (Pol I) subunits RPA34 and RPA49 in rRNA synthesis. Previous studies by various groups have demonstrated that these two subunits may play a role in transcription initiation and elongation by on Pol I occupancy in vivo, but this is still not clearly defined. Here, we deployed a high-resolution technique, native elongating transcript sequencing (NET-seq), to test the effects of RPA34 and RPA49 on Pol I occupancy in vivo. We found that when RPA34 was deleted, there was a significant reduction in Pol I occupancy across the rDNA template. Additionally, when RPA49 was deleted, Pol I occupancy was even further reduced across the template. Collectively, our findings suggest that perhaps RPA34 acts to stabilize RPA49 and these subunits may act primarily as transcription initiation factors for Pol I.

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Paused RNA polymerase II inhibits new transcriptional initiation

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Project description:Over 2000 publicly accessible human and mouse ChIP-Seq datasets for about 250 Transcription Factors and chromatin complexes from various databases (ENCODE, GEO) were mapped to custom-made human and mouse genomes containing a reference rDNA sequence of the appropriate species (Genbank U13369.1 for human, BK000964.3 for mouse). The read mapping density across the rDNA sequence was then extracted and normalized to the median in that dataset. Unbiased clustering and analysis, followed by curation, was performed to identify high-confidence patterns of rDNA occupancy for numerous hematopoietic TFs and TF families at canonical TF motif sequences. ************************ Data processing steps: FASTQs were trimmed using Trimmomatic with the following parameters: LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:30 Reads were mapped to customized genomes (containing additional rDNA sequence) using Bowtie2 using the following parameter: -X 2000 Read density across the rDNA sequence was extracted using igvtools ************************

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Hot spots of DNA double-strand breaks and genomic contacts of human rDNA units are involved in epigenetic regulation [rDNA units]

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