Grad-seq shines light on unrecognized RNA and protein complexes in the model bacterium Escherichia coli
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA641097 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:Stable protein complexes, including those formed with RNA, are major building blocks of every living cell. Escherichia coli has been the leading organism with respect to global bacterial protein-protein networks. However, there has been no global census of RNA/protein complexes in this model species of microbiology. Here, we performed Grad-seq to establish an RNA/protein complexome, reconstructing sedimentation profiles in a glycerol gradient for ~85% of all E. coli transcripts and ~49% of the proteins. In presenting use cases for this resource to be exploited, we show that its stable association with 30S ribosomes gives the seemingly noncoding RNA RyeG of prophage origin away as an mRNA of a toxic small protein. Similarly, we show that the broadly conserved uncharacterized protein YggL is a 50S subunit factor in assembled 70S ribosomes. Overall, this study complements existing interactome resources for the primary model bacterium E. coli with valuable RNA/protein complexome information and should facilitate functional discovery in this and many related species.
2020-06-30 | GSE152974 | GEO
Project description:RNA-protein interactions crucially underlie many steps of bacterial gene expression including post-transcriptional control by small regulatory RNAs (sRNAs). In stark contrast with recent progress in Gram-negative bacteria, knowledge about RNA and protein complexes in Gram-positive species remains scarce. Here, we used Grad-seq to draft a landscape of such complexes in Streptococcus pneumoniae, determining the sedimentation profiles of ~88% of the transcripts and ~62% of the proteins of this important human pathogen. Analysis of in-gradient distributions and subsequent tag-based protein capture identified interactions of the exoribonuclease Cbf1 (a.k.a. YhaM) with sRNAs that control bacterial competence. Contrary to expectation, the nucleolytic activity of Cbf1 stabilized these sRNAs, thereby promoting their function as repressors of competence. These results illustrate how this first RNA/protein complexome resource for a Gram-positive species can be utilized to identify new molecular factors in RNA-based regulation of pathways with relevance to bacterial virulence.
2021-01-27 | GSE138732 | GEO
Project description:Peripheral light harvesting (LH) antenna complexes have been studied extensively in the purple nonsulfur bacterium Rhodopseudomonas palustris because it produces different types of LH complexes under high light intensities (LH2 complex) and low light intensities (LH3 and LH4 complex). The ability of R. palustris to alter its peripheral LH complexes in response to changes in light intensity is attributed to the multiple operons that encode the a and b peptides that make up these complexes, whose expression is affected by light intensity, light quality, and oxygen tension. However, low resolution structures, amino acid similarities between the complexes, and a lack of transcriptional analysis made it difficult to determine the LH complexes composition and functions under different light intensities. It was also unclear how much diversity of the R. palustris LH complexes exists in nature.Results: To gain insight into the composition of the LH complexes, their function under high light intensities and low light intensities, and their prevalence in the environment we undertook an integrative genomics approach using 15 closely related R. palustris strains isolated from the environment and 5 R. palustris ecotypes whose genomes have been sequenced. We sequenced the genomes for the 15 closely related strains and using RNA-seq carried out transcriptomic analysis on all 20 strains grown under high light intensity and low light intensity. We were able to determine that even closely related R. palustris strains had differences in their pucBA gene content and expression, even under the same growth conditions. We also found that the LH2 complex could compensate for the lack of an LH4 complex under LL intensities but not under extremely LL intensities. Conclusions: This is the first time an integrative genomics approach has been used to study light harvesting in the environment. The variation observed in LH gene composition and expression in environmental isolates of R. palustris likely reflects how these strains have adapted to specific light conditions in the environment. We have also shown that there is redundancy between some of the LH complexes under certain light intensities, which may partially explain why multiple operons encoding LH complexes have evolved and been maintained in R. palustris. Examing the variation observed in LH gene composition and expression in various environmental isolates
2015-03-01 | E-GEOD-59544 | biostudies-arrayexpress
Project description:Peripheral light harvesting (LH) antenna complexes have been studied extensively in the purple nonsulfur bacterium Rhodopseudomonas palustris because it produces different types of LH complexes under high light intensities (LH2 complex) and low light intensities (LH3 and LH4 complex). The ability of R. palustris to alter its peripheral LH complexes in response to changes in light intensity is attributed to the multiple operons that encode the a and b peptides that make up these complexes, whose expression is affected by light intensity, light quality, and oxygen tension. However, low resolution structures, amino acid similarities between the complexes, and a lack of transcriptional analysis made it difficult to determine the LH complexes composition and functions under different light intensities. It was also unclear how much diversity of the R. palustris LH complexes exists in nature.Results: To gain insight into the composition of the LH complexes, their function under high light intensities and low light intensities, and their prevalence in the environment we undertook an integrative genomics approach using 15 closely related R. palustris strains isolated from the environment and 5 R. palustris ecotypes whose genomes have been sequenced. We sequenced the genomes for the 15 closely related strains and using RNA-seq carried out transcriptomic analysis on all 20 strains grown under high light intensity and low light intensity. We were able to determine that even closely related R. palustris strains had differences in their pucBA gene content and expression, even under the same growth conditions. We also found that the LH2 complex could compensate for the lack of an LH4 complex under LL intensities but not under extremely LL intensities. Conclusions: This is the first time an integrative genomics approach has been used to study light harvesting in the environment. The variation observed in LH gene composition and expression in environmental isolates of R. palustris likely reflects how these strains have adapted to specific light conditions in the environment. We have also shown that there is redundancy between some of the LH complexes under certain light intensities, which may partially explain why multiple operons encoding LH complexes have evolved and been maintained in R. palustris.
2015-03-01 | GSE59544 | GEO
Project description:New methods for the global identification of RNA-protein interactions have led to greater recognition of the abundance and importance of RNA-binding proteins (RBPs) in bacteria. Here, we expand this tool kit by developing SEC-seq, a method based on a similar concept as the established Grad-seq approach. In Grad-seq, cellular RNA and protein complexes of a bacterium of interest are separated in a glycerol gradient, followed by high-throughput RNA-sequencing and mass spectrometry analyses of individual gradient fractions. New RNA-protein complexes are predicted based on the similarity of their elution profiles. In SEC-seq, we have replaced the glycerol gradient with separation by size exclusion chromatography, which shortens operation times and offers greater potential for automation. Applying SEC-seq to Escherichia coli, we find that the method provides a higher resolution than Grad-seq in the lower molecular weight range up to ~500 kDa. This is illustrated by the ability of SEC-seq to resolve two distinct, but similarly sized complexes of the global translational repressor CsrA with either of its antagonistic small RNAs, CsrB and CsrC. We also characterized changes in the SEC-seq profiles of the small RNA MicA upon deletion of its RNA chaperones Hfq and ProQ and investigated the redistribution of these two proteins upon RNase treatment. Overall, we demonstrate that SEC-seq is a tractable and reproducible method for the global profiling of bacterial RNA-protein complexes that offers the potential to discover yet-unrecognized associations between bacterial RNAs and proteins.
2022-10-27 | GSE212408 | GEO
Project description:A Grad-seq resource of RNA and protein complexes of Pseudomonas aeruginosa including transcripts of phage ΦKZ
| PRJNA662513 | ENA
Project description:RNA-protein interactions crucially underlie many steps of bacterial gene expression including post-transcriptional control by small regulatory RNAs (sRNAs). In stark contrast with recent progress in Gram-negative bacteria, knowledge about RNA and protein complexes in Gram-positive species remains scarce. Here, we used Grad-seq to draft a landscape of such complexes in Streptococcus pneumoniae, determining the sedimentation profiles of ~88% of the transcripts and ~62% of the proteins of this important human pathogen. Analysis of in-gradient distributions and subsequent tag-based protein capture identified interactions of the exoribonuclease Cbf1 (a.k.a. YhaM) with sRNAs that control bacterial competence. Contrary to expectation, the nucleolytic activity of Cbf1 stabilized these sRNAs, thereby promoting their function as repressors of competence. These results illustrate how this first RNA/protein complexome resource for a Gram-positive species can be utilized to identify new molecular factors in RNA-based regulation of pathways with relevance to bacterial virulence.
2020-03-20 | GSE145604 | GEO
Project description:Grad-seq of Pseudomonas aerugionosa PAO1 at OD 0.3 with and without infection with ΦKZ bacteriophage
2021-01-06 | GSE157708 | GEO
Project description: Not available
| S-EPMC7498339 | biostudies-literature
Project description:A census of RNA/protein complexes in a model Gram-positive bacterium reveals exonuclease-mediated sRNA activation in competence regulation
| PRJNA576919 | ENA