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Homo sapiens

ABSTRACT: RNA:DNA hybrids in the human genome have distinctive nucleotide characteristics, chromatin composition, and transcriptional relationships (RNA-seq)

PROVIDER: PRJNA284147 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR2028250.fastq.gz	Fastqsanger.gz
	SRR2028251.fastq.gz	Fastqsanger.gz
	SRR2028252.fastq.gz	Fastqsanger.gz
	SRR2028253.fastq.gz	Fastqsanger.gz
	SRR2028254.fastq.gz	Fastqsanger.gz

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RNA:DNA hybrids in the human genome have distinctive nucleotide characteristics, chromatin composition, and transcriptional relationships

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Project description:The transcription factor IIH (TFIIH) is a key player in transcription and DNA repair by nucleotide excision repair. It is made of 10 subunits organized in core-TFIIH and CAK sub-complexes bridged by XPD. Pathogenic variants in the ERCC2/XPD gene give rise to xeroderma pigmentosum (XP) or trichothiodystrophy (TTD), two distinct clinical entities with opposite skin cancer proneness. Here, we show that TTD variants cause a partial dissociation of the CAK from the chromatin and from the core-TFIIH. Mass spectrometry analysis reveals that the chromatin-bound CAK, as a component of the entire TFIIH, participates in a protein assembly containing the RNA-binding proteins DDX1, SFPQ, NONO as well as RNA polymerase II (Pol II). Gene silencing experiments demonstrate that the protein assembly is required to process the DNA:RNA hybrids formed during Pol II extension and to protect the cell from transcriptional stress. TTD-specific variants in ERCC2/XPD result in TFIIH instability, altered interaction of the CAK with DDX1-SFPQ-NONO, and R-loop accumulation. Therefore, the limited amount of TFIIH that distinguishes TTD from XP gives rise to transcriptional stress and extensive gene expression deregulations, thus accounting for the wide spectrum of TTD clinical features.

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RNA:DNA hybrids in the human genome have distinctive nucleotide characteristics, chromatin composition, and transcriptional relationships RDIP-seq)

Project description:Mapping of RNA:DNA hybrids in human cells reveals a number of characteristics of these non-canonical nucleic acid structures. A directional sequencing approach reveals the RNA component of the RNA:DNA hybrid to be purine-rich, indicating a thermodynamic contribution to the stability of these structures. The RNA:DNA hybrids are enriched at loci with decreased DNA methylation and increased DNase hypersensitivity, and within larger domains with characteristics of heterochromatin formation. Studies of chromatin at RNA:DNA hybrids shows the presence of the ILF2 and ILF3 transcription factors, supporting a model of certain transcription factors binding preferentially to the RNA:DNA conformation. Overall, there is little to indicate a dependence for RNA:DNA hybrids forming co-transcriptionally, with results from the ribosomal DNA repeat unit instead supporting a model of RNA generating these structures in trans. The results of the study indicate heterogeneous functions of these genomic elements and new insights into their formation and stability in vivo.

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RNA:DNA hybrids in the human genome have distinctive nucleotide characteristics, chromatin composition, and transcriptional relationships (RNA-seq)

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RNA:DNA hybrids in the human genome have distinctive nucleotide characteristics, chromatin composition, and transcriptional relationships RDIP-seq)

2015-06-15 | E-GEOD-68948 | biostudies-arrayexpress

RNA:DNA hybrids in the human genome have distinctive nucleotide characteristics, chromatin composition, and transcriptional relationships (RNA-seq)

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Project description:We are interested in proteins which could preferentially interact with DNA: RNA hybrids upon DNA damage. To achieve that, we treated HEK293T with 10 Gy IR, followed by 20 min recovery. We applied S9.6 antibody, which could recognize DNA: RNA hybrids structure, to isolate their interactome upon either normal conditions or DNA damage conditions, after which those proteins sample were sent for mass spec analysis, and proteins specifically interacting with DNA: RNA hybrids were then screened and analysed.

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Project description:The biotin labeled DNA:RNA hybrids with or without RNA m5C modification were incubated with lysates of 293 cells treated with H2O2, and were pulled down by Streptavidin. The proteins pulled down by the empty beads control, unmodified or m5C-modified hybrids were analyzed by mass spectrometry

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Project description:This study shows that Integrator complex subunit 6 (INTS6) associates with the trimeric SOSS1 (comprising INTS3, INIP, and hSSB1) to form a tetrameric SOSS1 complex following DNA damage. INTS6 binds to DNA:RNA hybrids and plays a crucial role in Protein Phosphatase 2 (PP2A) recruitment to DNA double-strand breaks (DSBs), facilitating the dephosphorylation of RNAPII. Furthermore, INTS6 prevents the accumulation of damage-induced RNA transcripts and stabilizes DNA:RNA hybrids at DSB sites. INTS6 interacts with, and promotes the recruitment of Senataxin (SETX) to DSBs, facilitating the resolution of DNA:RNA hybrids/R-loops. These data demonstrate the significance of the SOSS1 complex in autoregulating DNA:RNA dynamics and the promotion of efficient DNA repair.

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