Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

RNA-Seq samples of E. coli K12 MG1655, deletion of gcvB sRNA gene, temperature mutation in RNAse E.

ABSTRACT: Small RNAs (sRNAs) are major regulators of gene expression in bacteria. Traditionally, sRNAs were considered mainly as regulators of translation, exerting their regulatory function by base pairing with their target mRNAs, and through this, blocking or exposing the ribosome binding site. However, accumulating evidence suggest that sRNAs not only regulate translation, but also affect the transcript stability by assisting or interfering with endoribonuclesaes. While cooperation of sRNAs with endoribonuclesaes was demonstrated for a few genes, its extent in the bacterial cell was not assessed in large scale. Here, we take advantage of large-scale RNA- seq-based approaches to study the extent of cooperation between sRNAs with the major endoribonuclease in Escherichia coli, RNase E. As a model sRNA, we use the well-known sRNA GcvB, a sRNA that regulates genes involved in amino acid metabolism and transport. To study the cooperation between GcvB and RNase E we apply the mutant cycle approach to four strains of E. coli: wt; GcvB mutant/RNase E wt; GcvB wt/RNase E mutant; GcvB mutant/RNase E mutant. By applying RNA-seq and differential gene expression analysis, we infer different modes of cooperation between RNase E and GcvB, and show that a statistically significant number of GcvB targets are downregulated by cooperation between the two regulators. Furthermore, using the TIER-seq approach to map RNase E cleavage sites transcriptome-wide and analysing them in view of GcvB binding sites, we attempt to infer the mechanisms of cooperation between the two regulators.

INSTRUMENT(S): NextSeq 500

ORGANISM(S): Escherichia coli str. K-12 substr. MG1655

SUBMITTER: Meshi Barsheshet

PROVIDER: E-MTAB-12466 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

ACCESS DATA

Similar Datasets

Project description:Small RNAs (sRNAs) are hypothesized to contribute to hybrid vigor because they maintain genome integrity, contribute to genetic diversity, and control gene expression. We used Illumina sequencing to assess how sRNA populations vary between two maize inbred lines (B73, Mo17) and their hybrid. We sampled sRNAs from the seedling shoot apex and the developing ear, two rapidly growing tissues that program the greater growth of maize hybrids. We found that parental differences in siRNAs primarily originate from repeat regions. Although the maize genome contains greater number and complexity of repeats compared to Arabidopsis or rice, we confirmed that like these simpler plant genomes, 24-nt siRNAs whose abundance differs between maize parents also show a trend of downregulation following hybridization. Surprisingly, hybrid vigor is fully maintained when 24-nt siRNAs are globally reduced by mutation of the RNA-dependent RNA polymerase2 (RDR2) encoded by modifier of paramutation1 (mop1). We also discovered that 21-22nt siRNAs derived from a number of distinct retrotransposon families differentially accumulate between B73 and Mo17 as well as their hybrid. Thus, maize possesses a novel source of genetic variation for regulating both transposons and genes at a genomic scale, which may contribute to its high degree of observed heterosis. sRNA libraries were derived from RNA isolated from the seedling shoot apex and developing ear tissues from B73, Mo17, B73xMo17 and Mo17xB73. The shoot apex was chosen because it is enriched for meristematic tissue where cell proliferation occurs, rates of organ initiation are determined, and organ size is specified. The developing ear was examined because it is enriched in meristematic tissue and is undergoing rapid growth, and also because the mature ear shows the highest degree of heterosis. Total RNA was isolated and separated on a 15% TBE-Urea polyacrylamide gel. Using a 10-bp ladder, the sRNA fraction representing 10-40-bp was excised. sRNA libraries were prepared according to Lu et al. (2007) or manufacturer's instructitions (Illumina). A combination of Perl scripts and FASTX toolkit scripts were used to remove adapters, collapse identical sequences and count reads per sequence. Supplementary processed data text files contain the distinct sRNA sequences for all of the genotypes analyzed in that experiment. Abundance (reads per million) was calculated for each distinct sequence by dividing the number of reads of distinct sRNA in a library by the total number of sRNA reads for that library and multiplying this by 1 million. Genome builds: B73 genome, maizesequence.org release 4a.53 (October, 2009); Mo17 whole genome shotgun clones.

Project description:The rice blast fungus, Magnaporthe oryzae is a destructive pathogen of rice and other related crops, causing significant yield losses worldwide. Endogenous small RNAs (sRNAs), including small interfering RNAs (siRNAs) and microRNAs (miRNAs) are critical components of gene regulation in many eukaryotic organisms. Recently several new species of sRNAs have been identified in fungi. This fact along with the availability of genome sequence makes M. oryzae a compelling target for sRNA profiling. We have examined sRNA species and their biosynthetic genes in M. oryzae, and the degree to which these elements regulate fungal stress responses. To this end, we have characterized sRNAs under different physiological stress conditions, which had not yet been examined in this fungus. The resulting libraries are composed of more than 37 million total genome matched reads mapping to intergenic regions, coding sequences, retrotransposons, inverted, tandem, and other repeated regions of the genome with more than half of the small RNAs arising from intergenic regions. The 24 nucleotide (nt) size class of sRNAs was predominant. A comparison to transcriptional data of M. oryzae undergoing the same physiological stresses indicates that sRNAs play a role in transcriptional regulation for a small subset of genes. Support for this idea comes from generation and characterization of mutants putatively involved in sRNAs biogenesis; our results indicate that the deletion of Dicer-like genes and an RNA-Dependent RNA Polymerase gene increases the transcriptional regulation of this subset of genes, including one involved in virulence. Various physiological stressors and in planta conditions alter the small RNA profile of the rice blast fungus. Characterization of sRNA biosynthetic mutants helps to clarify the role of sRNAs in transcriptional control. Examination of small RNA populations of eight mycelial as well as In planta tissues of M. oryzae strain 70-15. Mycelia grown in complete medium (CM), minimal medium (MM), carbon starved medium (CS: MM lacking carbon source), nitrogen starved medium (NS: MM lacking nitrogen source) and a reactive oxygen species-rich environment generated by amendment of Paraquat (PQ) to the CM. In planta tissues include mock inoculated rice leaves (LMg0), 72 hours post-inoculation in rice leaves (LMg72) and 96 hours post-inoculation in rice leaves (LMg96).

Dataset Information

RNA-Seq samples of E. coli K12 MG1655, deletion of gcvB sRNA gene, temperature mutation in RNAse E.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets