Identification of RNase III cleavage sites in Escherichia coli
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ABSTRACT:
PROVIDER: PRJEB26394 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:In this study, we performed RNA-sequencing on an E. coli model system to confirm known sites, identify novel targets, and determine the impact of RNase III cleavage events on transcript degradation and metabolic phenotypes. To find cleavage sites, we compared the abundance of sequencing reads across the transcriptome of a wild-type E. coli and an rnc- deletion mutant. The RNA-sequencing approach provided wider coverage and unprecedented resolution of mRNA abundance at each position in the transcriptome compared to prior studies that used qPCR, Northern blots, or microarrays. In addition to data collected from exponentially growing cells, we observed the effects of RNase III cleavage on transcript degradation by collecting samples in a time course after stopping nascent transcription with rifampicin. This work was supported by the Department of Energy Grant DE-SC0010329
2017-02-24 | GSE95318 | GEO
Project description:Members of the ribonuclease (RNase) III family regulate gene expression by processing double-stranded (ds) RNA. The founding member of the family, Escherichia coli (Ec) RNase III, is the most comprehensively studied and its E38A mutant (EcE38A) is an economical reagent for the preparation of small interfering (si) RNA cocktails. Previously, it was shown that EcRNase III recognizes dsRNA with little specificity and that EcE38A mainly produces 23-nucleotide (nt) siRNAs. To characterize substrate specificity and product size, we performed in vitro cleavage of dsRNAs by bacterial RNase IIIs and delineated the cleavage products by next generation sequencing. Surprisingly, we found that RNase III cleaves dsRNA at preferred sites and most siRNAs produced by EcE38A are 22 nt long. We eliminated the sequence specificity of EcE38A through the introduction of additional mutations, thereby creating a reagent that is ideally suited for producing heterogeneous siRNA cocktails to be used in gene silencing studies.
2019-05-26 | GSE120052 | GEO
Project description:In this study we generated 5'P libraries in wt and RNAse III mutant strains, grown to exponential and stationary phases. Libraries that retain short RNA fragments were also generated in both growth phases. After sequencing by Illumina NextSeq 500 system, reads were mapped to E. coli genome NC_000913.3. By comparing the read start counts per position in the wt and mutant strain libraries we identified the cleavage sites of RNase III.
2018-09-30 | E-MTAB-6704 | biostudies-arrayexpress
Project description:In this study, we employed TIER-seq in V. cholerae to identify cleavage sites of RNase E on a genome-wide scale.
2020-05-07 | GSE148675 | GEO
Project description:Bacterial RNase III cleaves long dsRNA at preferred sites
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Project description:Global identification of RNase E sites in Vibrio cholerae
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Project description:tRNA-derived fragments (tRFs) have emerged as key players of immunoregulation. Some RNase A superfamily members participate in the shaping of tRFs population. By comparing wild-type and knock-out macrophage cell lines our previous work (Lu L, et al. CMLS, 2022, 79: 209) revealed that RNase 2 can selectively cleave tRNAs. Here, we confirm the in vitro protein cleavage pattern by screening synthetic tRNAs, single-mutant variants and anticodon-loop DNA/RNA hairpins. By sequencing the tRFs products, we identified the cleavage selectivity by recombinant RNase 2 with base specificity at B1 (U/C) and B2 (A) sites, consistent with a previous cellular study. Knowledge of RNase 2 specific tRFs generation might guide new therapeutic approaches for infectious and immune-related diseases.
2024-05-29 | GSE250347 | GEO
Project description:The Corynebacterium glutamicum R cgR_1959 gene encodes an endoribonuclease of the RNase III family. Deletion mutant of cgR_1959 (Δrnc mutant) showed an elongated cell shape, and presence of several lines on the cell surface, indicating a required of RNase III for maintaining normal cell morphology in C. glutamicum. The level of mraZ mRNA was increased, whereas cgR_1596 mRNA encoding a putative cell wall hydrolase and ftsEX mRNA were decreased in the Δrnc mutant. The half-life of mraZ mRNA was significantly prolonged in the Δrnc and the Δpnp mutant strains. This indicated that the degradation of mraZ mRNA was performed by RNase III and the 3′-to-5′ exoribonuclease, PNPase. Northern hybridization and primer extension analysis revealed that the cleavage site for mraZ mRNA by RNase III is in the coding region. Overproduction of MraZ resulted in an elongated cell shape. The expression of ftsEX decreased while that of cgR_1596 unchanged in an MraZ-overexpressing strain. An electrophoretic mobility shift assay and a transcriptional reporter assay indicate that MraZ is a transcriptional repressor of ftsEX in C. glutamicum. These results indicate that RNase III is required for efficient expression of MraZ-dependent ftsEX and MraZ-independent cgR_1596.
2016-01-17 | GSE75080 | GEO
Project description:RNase III is a ribonucleases that recognizes and cleaves double-stranded RNA. RNase III has been known be involved in rRNA processing, but has many additional roles controlling both expression and RNA turnover of specific messages. Many organisms have just one RNase III while some have both a full length RNase III and a mini-III that lacks the double-stranded RNA binding domain. The cyanobacteria Synechococcus sp. PCC 7002 has three homologs of RNase III that are unessential even when deleted in combination. We were interested what coding regions these RNase III enzymes were influencing and if they had redundant or distinct specificities. To address these questions we collected samples for RNA-sequencing from WT, the single, double, and triple RNase III mutants in triplicate. Approximately 20% of genes were differentially expressed in various mutants with some operons and regulons showing complex changes in expression levels between mutants. We describe the role of two RNase III’s in 23S rRNA maturation, and show how the third is involved in copy number regulation of one of the six plasmids (pAQ3). Purified enzymes were capable of cleaving some E. coli RNase III target sequences, highlighting the remarkably conserved substrate specificity between organisms yet complex regulation of gene expression.
2018-01-16 | GSE99279 | GEO
Project description:Our results demonstrate that RNase G controls expression levels of H-NS encoded by hns, strongly associated with the pathogenicity of S. Typhimurium. The hns mRNA abundance is mediated by RNase G, which cleaves the 5’-UTR of hns mRNA. In the upstream pathway, the induced expression of RNase G in host environment condition is attributable to reduced RNase III cleavage activity on rng mRNA. Our findings suggest the link between Salmonella pathogenicity and RNase III-RNase G pathway, underlining the importance of posttranscriptional regulation of H-NS in the downstream pathway, which controls Salmonella pathogenicity island-1 type III secretion system for the survival and virulence of S. Typhimurium in host cell.
2021-01-08 | PXD011090 | Pride