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Cas9 BLESS sequencing

ABSTRACT: Guide-free Cas9 from pathogenic Campylobacter jejuni bacteria causes severe damage to DNA

PROVIDER: PRJEB34300 | ENA |

REPOSITORIES: ENA

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			Action	DRS
		Other
	ERR3719571_1.fastq.gz	Fastqsanger.gz
	ERR3719571_2.fastq.gz	Fastqsanger.gz
	ERR3719563_1.fastq.gz	Fastqsanger.gz
	ERR3719563_2.fastq.gz	Fastqsanger.gz

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1 - 5 of 17

Similar Datasets

Cas9 of bacterial pathogens induces DNA damage and apoptosis of human cells

Project description:CRISPR-Cas is an RNA-based defense system that enables prokaryotes to recognize invading foreign DNA by cognate crRNA guides and destroy it by CRISPR-associated Cas nucleases 1,2 . Elucidation of the interference mechanism of the Streptococcus pyogenes Type II CRISPR- Cas9 system has allowed for the successful repurposing of SpCas9 as a generic genome editing tool, with great promise for human gene therapy 3 . However, especially for therapeutic applications, some caution seems appropriate, because Cas9 systems from some human pathogens may induce a cytotoxic response via an unknown mechanism 4 . Here we show that when released in human cells, Cas9 nucleases from the pathogenic bacteria Campylobacter jejuni and S. pyogenes have the potential to cause severe DNA damage. In the absence of a CRISPR RNA guide, native Cas9 nucleases from both pathogens enter the host nucleus, where their presence leads to promiscuous double stranded DNA breaks (DSBs) and induction of cell death. DSB induction can be reduced to background levels either by saturation of CjCas9 and SpCas9 with crRNA guides or by inactivating their nuclease activity. Our results demonstrate that guide-free Cas9 of bacterial pathogens might play an important role in pathogenicity. Furthermore, we propose that saturating Cas9 with appropriate guide RNAs is crucial for efficient and safe therapeutic applications.

2022-03-02 | PXD006712 | Pride

Campylobacter jejuni Cas9 is necessary for efficient infection of Caco-2 cells and cell stress induction

Project description:Objective: We previously showed that Campylobacter jejuni Cas9 was necessary for full virulence and efficient induction of human cell stress followed by cell death, but no virulence mechanism could be unravelled then. Therefore, we carried out global genome-wide transcriptomics, using RNA samples extracted from cells infected by a wild-type C. jejuni strain and the corresponding cas9 deletion strain. Methods: Caco-2 epithelial cells were infected with C. jejuni wild-type or cas9 deletion strains and RNA was extracted at 5 timepoints post-infection. Results: Wild-type C. jejuni induced cellular stress, DNA damage and cell death pathways in Caco-2 cells continuously from 30 min up to 4 hours after infection, eventually followed by visual cell death induction after 24 hours; the corresponding cas9 deletion strain did also induce cell stress pathways within the first two hours of infection but at later timepoints, stress pathway induction had dropped and after 4 hours of infection, expression of the corresponding stress genes had normalised to near-t=0 (uninfected) expression values; the cas9 deletion strain did not lead to visual cell death induction after 24 hours. Conclusions: Campylobacter jejuni Cas9 is necessary for full virulence of C. jejuni and is involved in the efficient induction of cellular stress pathways and eventually, cell death.

2018-09-30 | GSE89661 | GEO

CRISPR RNA-dependent binding and cleavage of endogenous RNAs by the Campylobacter jejuni Cas9

Project description:This study investigates the RNA targets and cleavage sites of endogenous Cas9 in the food-borne pathogen Campylobacter jejuni. Direct RNA binding targets of Cas9 in C. jejuni strain NCTC11168 were determined using RIP-seq. The Cleavage sites were then predicted in the RNA targets by comparing total transcriptome data from WT and deletion (cas9, crRNA3, tracrRNA, CRISPR-tracrRNA) strains. PAMs for the CjeCas9 were enriched using the PAM-SCANR platform, which operates through a GFP reporter gene. Upon GFP (and thus functional PAM) enrichment, fluorescing cells were isolated using FACS and prepared plasmid DNA was amplified and prepared for sequencing.

2018-03-01 | GSE106849 | GEO

Crosstalk between CRISPR-Cas9 and the human transcriptome

Project description:CRISPR-Cas9 expression independent of its cognate synthetic guide RNA (gRNA) causes widespread genomic DNA damage in human cells. To investigate whether Cas9 can interact with endogenous human RNA transcripts independent of its guide, we perform eCLIP (enhanced CLIP) of Cas9 in human cells and find that Cas9 reproducibly interacts with hundreds of endogenous human RNA transcripts. This association can be partially explained by a model built on gRNA secondary structure and sequence. Critically, transcriptome-wide Cas9 binding sites do not appear to correlate with published genome-wide Cas9 DNA binding or cut-site loci under gRNA co-expression. However, even under gRNA co-expression low-affinity Cas9-human RNA interactions (which we term CRISPR crosstalk) do correlate with published elevated transcriptome-wide RNA editing. Our findings do not support the hypothesis that human RNAs can broadly guide Cas9 to bind and cleave human genomic DNA, but they illustrate a clear cellular RNA impact likely inherent to CRISPR-Cas systems.

2022-01-24 | GSE167466 | GEO

Campylobacter jejuni gene expression in germ-free mice

Project description:Expression arrays comparing Campylobacter jejuni NCTC11168 during growth in the cecum of germ-free C57 BL/6 IL-10 knockout mice to C. jejuni NCTC11168 during growth in Bolton broth.

2011-04-14 | GSE28578 | GEO

Transcriptional profile in macrolide susceptible and resistant Campylobacter jejuni

Project description:Campylobacter jejuni is a major zoonotic pathogen transmitted to humans via the food chain. C. jejuni is prevalent in chickens, a natural reservoir for this pathogenic organism. Due to the importance of macrolide antibiotics in clinical therapy of human campylobacteriosis, development of macrolide resistance in Campylobacter has become a concern for public health.To facilitate understanding the molecular basis associated with the fitness difference between Erys and Eryr Campylobacter, we compared the transcriptomes between ATCC 700819 and its isogenic Eryr transformant T.L.101 using DNA microarray.

2011-11-02 | GSE33330 | GEO

Non-canonical crRNAs derived from host transcripts enable multiplexable RNA detection by Cas9 [dRNA-seq]

Project description:Differential RNA sequencing using terminator exonuclease (TEX) to identify processed versus unprocessed transcripts in Campylobacter jejuni strain CG84-21

2021-04-27 | GSE156265 | GEO

RNAseq analysis of Campylobacter jejuni's response to glutamine addition in growth media

Project description:Campylobacter jejuni is a human pathogen which causes campylobacteriosis, one of the most widespread zoonotic enteric diseases worldwide. Growth of Campylobacter can be improved through the addition of glutamine to media which serves as the nitrogen source. RNAseq was used to identify the transcriptomic response of Campylobacter jejuni when the nitrogen source was switched from serine (poor growth) to glutamine (good growth) in chemostat cultures.

2022-12-01 | E-MTAB-11495 | biostudies-arrayexpress

Campylobacter jejuni murine adaptation: unadapted v. adapted during growth in C57 BL/6 IL-10-/- germ-free mice.

Project description:Expression arrays comparing Campylobacter jejuni 11168 before and after serial passage in C57 BL/6 IL-10 deficient mice. Transcript abundance was compared between C. jejuni variants during growth in the ceca of C57 BL/6 IL-10 deficient germ-free mice.

2010-08-28 | GSE23811 | GEO

RNAseq analysis of Campylobacter jejuni treated with hypochlorite

Project description:Campylobacter jejuni is a human pathogen which causes campylobacteriosis, one of the most widespread zoonotic enteric diseases worldwide. Most cases of sporadic C. jejuni infection occur through the handling or consumption of undercooked chicken meat, or cross-contamination of other foods with raw poultry fluid. A common practice to combat Campylobacter infection is to treat chickens with chlorine which kills the microbe. This analysis aimed to elucidate the transcriptomic response of Campylobacter jejuni treated with hypochlorite through Illumina sequencing. C. jejuni was grown and treated with hypochlorite. Samples were taken 5, 20 and 45 min after treatment for RNAseq analysis.The data generated were compared to the transcriptome pre-exposure to determine C. jejuni's response to hypochlorite.

2023-10-01 | E-MTAB-11479 | biostudies-arrayexpress

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