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RNA-Seq of A375 cells treated with tetracycline-based crosslinking probes to identify binding sites for the tetracycline analogs Col-3 and doxycycline on the human ribosome

ABSTRACT: RNA-Seq of A375 cells treated with tetracycline-based crosslinking probes to identify binding sites for the tetracycline analogs Col-3 and doxycycline on the human ribosome

PROVIDER: PRJEB28264 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	ERR2743200.fastq.gz	Fastqsanger.gz
	ERR2743210.fastq.gz	Fastqsanger.gz
	ERR2743201.fastq.gz	Fastqsanger.gz
	ERR2743202.fastq.gz	Fastqsanger.gz
	ERR2743193.fastq.gz	Fastqsanger.gz

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RNA-Seq of A375 cells treated with tetracycline-based crosslinking probes to identify binding sites for the tetracycline analogs Col-3 and doxycycline on the human ribosome

Project description:Following the identification of the 80S ribosome as a putative target of the tetracycline analogs Col3 and doxycycline, we next sought to identify the precise binding sites for these tetracyclines within the ribonucleoprotein complex. We incorporated dual bioorthogonal handles into tetracycline-based probes, containing both a photoactive diazirine to enable direct probe crosslinking to the human ribosome and an azide handle to allow selective enrichment of crosslinked biomolecules via copper-free click chemistry. The Col-3 and doxycycline probes were each incubated with A375 cells, followed by irradiation at 365 nm to induce photolysis of the diazirine moiety and subsequent crosslinking to adjacent ribosomal components. Pulldown and RNA-Seq of the crosslinked RNAs from our experiments were used to identify enrichment of reverse transcription (RT) stops at ribosomal RNA sites caused by local crosslinking of our probes. This RNA-Seq based RT stop enrichment analysis was compared to results using an non-specific aniline control probe and untreated controls.

2018-08-30 | E-MTAB-7147 | biostudies-arrayexpress

RNA-Seq of A375 cells treated with tetracycline-based crosslinking probes, with and without competition by parent tetracyclines, to show specific binding of the tetracycline analogs Col-3 and doxycycline on rRNA substructures of the human ribosome

Project description:Upon finding that the tetracyclines COL-3 and doxycycline target unique rRNA substructures of the 80S ribosome (E-MTAB-7147), we confirmed these putative ribosomal binding sites by showing that tetracycline-based crosslinking probes are specifically competed from these rRNA structures by their parent drugs. In short, we incorporated dual bioorthogonal handles into tetracycline-based probes, containing both a photoactive diazirine to enable direct probe crosslinking to the human ribosome and an azide handle to allow selective enrichment of crosslinked biomolecules via copper-free ‘click’ chemistry. The COL-3 and doxycycline probes were each incubated with A375 cells, followed by irradiation at 365 nm to induce photolysis of the diazirine moiety and subsequent crosslinking to adjacent ribosomal components. Pulldown and RNA-Seq of the crosslinked RNAs from our experiments were used to identify enrichment of reverse transcription (RT) stops at ribosomal RNA sites caused by local crosslinking of our probes. In these experiments, UV crosslinking was preformed with the COL-3 and doxycycline probe compounds in the presence and absence of free COL-3 and doxycycline (i.e., parent tetracycline molecules lacking the bifunctional diazirine-azide moiety), which were used as competitors to specifically diminish crosslinking to the ribosomal RNA. Crosslinking to ribosomal RNA was determined using RNA-Seq based RT stop enrichment, which was was also compared to inactive tetracycline probes containing a modified bifunctional linker, a non-specific aniline probe, and untreated controls.

2018-08-31 | E-MTAB-7161 | biostudies-arrayexpress

Project description:RNA-Seq of A375 cells treated with tetracycline-based crosslinking probes, with and without competition by parent tetracyclines, to show specific binding of the tetracycline analogs Col-3 and doxycycline on rRNA substructures of the human ribosome

| PRJEB28296 | ENA

RNA-seq of human melanoma cell line A375 treated with the tetracycline analogs Col-3 or doxycycline against untreated controls

Project description:RNA-seq of human melanoma cell line A375 treated with the tetracycline analogs Col-3 or doxycycline against untreated controls

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Genome Wide Effect of Tetracycline, Doxycycline and 4-epidoxycycline on gene expression in Saccharomyces cerevisiae

Project description:Investigating Saccharomyces cerevisiae genome-wide gene expression response to tetracycline family chemicals.

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RNA-seq of human melanoma cell line A375 treated with the tetracycline analogs Col-3 or doxycycline against untreated controls

Project description:Apart from their antibacterial activity, tetracyclines have demonstrated diverse and beneficial “off-target” biological activities in human disease, spurring interest in their development as non-antimicrobial therapeutics in diseases such as cancer. To better define biological pathways modified by the tetracyclines in human cancer, we sought to determine differential gene expression in the A375 cancer cell line treated with the tetracyclines, Col-3 and doxycycline, relative to control (vehicle-treated) cells. A375 cells were treated with either 10 μM Col-3, 10 μM doxycycline, or vehicle (DMSO) for 6 h, and changes to gene expression were assessed by RNA-Seq.

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Soil tetracycline resistant genes

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Project description:We isolated a strain that efficiently degrades tetracycline and compared its proteomic data in the presence and absence of tetracycline to analyze the molecular mechanism of tetracycline biodegradation

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Acinetobacter oleivorans DR1 : Wild type treated with tetracycline vs. harboring plasmid (pAST2) treated with tetracycline

Project description:Transcriptional profiling of A. oleivorans DR1 cells comparing tetracylcline-treated wild type cells with tetracycline-treated DR1 cell harboring pAST2. To identify genes important for tetracycline stress response in A. oleivorans DR1 and A. oleivorans DR1 harboring pAST2, the cells were grown to exponential phase (OD600 ~0.4) and treated with tetracycline (1μg/ml) over a period of 15 min.

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