Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1160042 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR30642400.fastq.gz | Fastqsanger.gz | |||
| SRR30642387.fastq.gz | Fastqsanger.gz | |||
| SRR30642388.fastq.gz | Fastqsanger.gz | |||
| SRR30642389.fastq.gz | Fastqsanger.gz | |||
| SRR30642390.fastq.gz | Fastqsanger.gz |
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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
Project description:In bacteria and archaea, CRISPR loci confer adaptive, sequence-based immunity against viruses and plasmids. CRISPR interference is specified by CRISPR RNAs (crRNAs) that are transcribed and processed from CRISPR spacers and repeats. Pre-crRNA processing is essential for CRISPR interference in all systems studied thus far. Here we examine crRNA biogenesis and CRISPR interference in naturally competent Neisseria spp., including the human pathogen N. meningitidis. Our studies reveal a unique crRNA maturation pathway in which crRNA transcription is driven by promoters that are embedded within each repeat, yielding crRNA 5’ ends are not formed by processing. Although crRNA 3’ end formation occurs through RNase III cleavage of a pre-crRNA/tracrRNA duplex, as in other Type II CRISPR systems, this processing event is dispensable for interference. The meningococcal pathway is the most streamlined CRISPR/cas system characterized to date. Endogenous CRISPR spacers frequently target genomic sequences of other Neisseria strains and so limit natural transformation, which is the primary source of genetic variation that contributes to immune evasion, antibiotic resistance, and virulence in N. meningitidis. dRNA-seq approach for RNA samples from cultures of N. lactamica 020-06, harvested at mid-log. Two cDNA libraries from total RNA were prepared to distinguish between transcripts with either primary orprocessed 5’ ends: one library is generated from untreated RNA, whereas the other is treated with terminator exonuclease (TEX),
2013-04-23 | E-GEOD-44582 | biostudies-arrayexpress
Project description:To gain a global view on the impact of the collateral activity on protein expression levels, mass spectrometry was used for proteomics analysis of HEK 293T cells when PspCas13b was co-expressed with non-targeted (NT) crRNA or targeting (T) crRNA, which targets the BCR-ABL1 mRNA breakpoint. There was no evidence of off-target protein degradation by T crRNA compared to the non-targeting crRNA. The only significantly repressed proteins were the target BCR-ABL1 p190 (81% reduction) and eGFP (89% reduction) that are encoded on the same mRNA expression construct and encoded mRNA. Catalytic dead Cas13 (dCas13) and crRNA only were used as negative control.
2024-05-20 | PXD051089 | Pride
Project description:The associated experiments document the production of small RNA (sRNA) during the expression of Cas13 and crRNA, crRNA alone, or controls from agrobacterium spot infiltration in Nicotiana benthamiana. We document the specific production of sRNA corresponding to the guide sequence of the targeted mRNA. In cases where a multi-guide crRNA or a hairpin were expressed, abundent sRNA are produced correspinding to the target mRNA, but outside of the corresponding guide sequence site.
2021-12-07 | GSE171980 | GEO
Project description:Purpose: RNase Y is a major enzyme responsible for mRNA degradation in Streptococcus pyogenes. The goals of this study are to understand whether RNase Y plays a role in operon transcription of S. pyogenes NZ131 by using RNA-seq analysis. Methods: S. pyogenes mRNA profiles of wild type (WT) and RNase Y mutant (∆rny) were generated by deep sequencing, in duplicate, using Illumina Hiseq 2000. The sequence reads were aligned to the S. pyogenes genome using Bowtie2. The aligned files were sorted to BAM format and indexed using Samtools. The read depth of each base was derived from BAM files using BEDtools. Operon organization of S. pyogenes WT and ∆rny strains were predicted based on base reads. Results: A total of 11 to 12 billion reads were obtained from each sample. More than 99% of these reads were mapped to the S. pyogenes genome. Predictions of operon organization using WT and ∆rny samples showed little difference between the two strains. Conclusions: Our result shows that the mutation of RNase Y does not affect the operon organization of S. pyogenes NZ131.
2017-06-02 | GSE99533 | GEO
Project description:In bacteria and archaea, CRISPR loci confer adaptive, sequence-based immunity against viruses and plasmids. CRISPR interference is specified by CRISPR RNAs (crRNAs) that are transcribed and processed from CRISPR spacers and repeats. Pre-crRNA processing is essential for CRISPR interference in all systems studied thus far. Here we examine crRNA biogenesis and CRISPR interference in naturally competent Neisseria spp., including the human pathogen N. meningitidis. Our studies reveal a unique crRNA maturation pathway in which crRNA transcription is driven by promoters that are embedded within each repeat, yielding crRNA 5’ ends are not formed by processing. Although crRNA 3’ end formation occurs through RNase III cleavage of a pre-crRNA/tracrRNA duplex, as in other Type II CRISPR systems, this processing event is dispensable for interference. The meningococcal pathway is the most streamlined CRISPR/cas system characterized to date. Endogenous CRISPR spacers frequently target genomic sequences of other Neisseria strains and so limit natural transformation, which is the primary source of genetic variation that contributes to immune evasion, antibiotic resistance, and virulence in N. meningitidis.
2013-04-23 | GSE44582 | GEO