Dataset Information

RNase J is required for processing of a small number of RNAs in Rhodobacter sphaeroides.

ABSTRACT: All bacteria contain multiple exoribonucleases to ensure a fast breakdown of different RNA molecules, either for maturation or for complete degradation to the level of mononucleotides. This efficient RNA degradation plays pivotal roles in the post-transcriptional gene regulation, in RNA processing and maturation as well as in RNA quality control mechanisms and global adaption to stress conditions. Besides different 3'-to-5' exoribonucleases mostly with overlapping functions in vivo many bacteria additionally possess the 5'-to-3' exoribonuclease, RNase J, to date the only known bacterial ribonuclease with this activity. An RNA-seq approach was applied to identify specific targets of RNase J in the ?-proteobacterium Rhodobacter sphaeroides. Only few transcripts were strongly affected by the lack of RNase J implying that its function is mostly required for specific processing/degradation steps in this bacterium. The accumulation of diverse RNA fragments in the RNase J deletion mutant points to RNA features that apparently cannot be targeted by the conventional 3'-exoribonucleases in Gram-negative bacteria.

SUBMITTER: Rische-Grahl T

PROVIDER: S-EPMC4179960 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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RNase J is required for processing of a small number of RNAs in Rhodobacter sphaeroides.

Rische-Grahl Tom T Weber Lennart L Remes Bernhard B Förstner Konrad U KU Klug Gabriele G

RNA biology 20140612 7

All bacteria contain multiple exoribonucleases to ensure a fast breakdown of different RNA molecules, either for maturation or for complete degradation to the level of mononucleotides. This efficient RNA degradation plays pivotal roles in the post-transcriptional gene regulation, in RNA processing and maturation as well as in RNA quality control mechanisms and global adaption to stress conditions. Besides different 3'-to-5' exoribonucleases mostly with overlapping functions in vivo many bacteria ...[more]

PMID: 24922065

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Project description:BackgroundThe polynucleotide phosphorylase (PNPase) is conserved among both Gram-positive and Gram-negative bacteria. As a core part of the Escherichia coli degradosome, PNPase is involved in maintaining proper RNA levels within the bacterial cell. It plays a major role in RNA homeostasis and decay by acting as a 3'-to-5' exoribonuclease. Furthermore, PNPase can catalyze the reverse reaction by elongating RNA molecules in 5'-to-3' end direction which has a destabilizing effect on the prolonged RNA molecule. RNA degradation is often initiated by an endonucleolytic cleavage, followed by exoribonucleolytic decay from the new 3' end.ResultsThe PNPase mutant from the facultative phototrophic Rhodobacter sphaeroides exhibits several phenotypical characteristics, including diminished adaption to low temperature, reduced resistance to organic peroxide induced stress and altered growth behavior. The transcriptome composition differs in the pnp mutant strain, resulting in a decreased abundance of most tRNAs and rRNAs. In addition, PNPase has a major influence on the half-lives of several regulatory sRNAs and can have both a stabilizing or a destabilizing effect. Moreover, we globally identified and compared differential RNA 3' ends in RNA NGS sequencing data obtained from PNPase, RNase E and RNase III mutants for the first time in a Gram-negative organism. The genome wide RNA 3' end analysis revealed that 885 3' ends are degraded by PNPase. A fair percentage of these RNA 3' ends was also identified at the same genomic position in RNase E or RNase III mutant strains.ConclusionThe PNPase has a major influence on RNA processing and maturation and thus modulates the transcriptome of R. sphaeroides. This includes sRNAs, emphasizing the role of PNPase in cellular homeostasis and its importance in regulatory networks. The global 3' end analysis indicates a sequential RNA processing: 5.9% of all RNase E-dependent and 9.7% of all RNase III-dependent RNA 3' ends are subsequently degraded by PNPase. Moreover, we provide a modular pipeline which greatly facilitates the identification of RNA 5'/3' ends. It is publicly available on GitHub and is distributed under ICS license.

Project description:UnlabelledIn bacteria, regulatory RNAs play an important role in the regulation and balancing of many cellular processes and stress responses. Among these regulatory RNAs, trans-encoded small RNAs (sRNAs) are of particular interest since one sRNA can lead to the regulation of multiple target mRNAs. In the purple bacterium Rhodobacter sphaeroides, several sRNAs are induced by oxidative stress. In this study, we focused on the functional characterization of four homologous sRNAs that are cotranscribed with the gene for the conserved hypothetical protein RSP_6037, a genetic arrangement described for only a few sRNAs until now. Each of the four sRNAs is characterized by two stem-loops that carry CCUCCUCCC motifs in their loops. They are induced under oxidative stress, as well as by various other stress conditions, and were therefore renamed here sRNAs CcsR1 to CcsR4 (CcsR1-4) for conserved CCUCCUCCC motif stress-induced RNAs 1 to 4. Increased CcsR1-4 expression decreases the expression of genes involved in C1 metabolism or encoding components of the pyruvate dehydrogenase complex either directly by binding to their target mRNAs or indirectly. One of the CcsR1-4 target mRNAs encodes the transcriptional regulator FlhR, an activator of glutathione-dependent methanol/formaldehyde metabolism. Downregulation of this glutathione-dependent pathway increases the pool of glutathione, which helps to counteract oxidative stress. The FlhR-dependent downregulation of the pyruvate dehydrogenase complex reduces a primary target of reactive oxygen species and reduces aerobic electron transport, a main source of reactive oxygen species. Our findings reveal a previously unknown strategy used by bacteria to counteract oxidative stress.ImportancePhototrophic organisms have to cope with photo-oxidative stress due to the function of chlorophylls as photosensitizers for the formation of singlet oxygen. Our study assigns an important role in photo-oxidative stress resistance to a cluster of four homologous sRNAs in the anoxygenic phototrophic bacterium Rhodobacter sphaeroides. We reveal a function of these regulatory RNAs in the fine-tuning of C1 metabolism. A model that relates oxidative stress defense to C1 metabolism is presented.

Dataset Information

RNase J is required for processing of a small number of RNAs in Rhodobacter sphaeroides.

Publications

RNase J is required for processing of a small number of RNAs in Rhodobacter sphaeroides.

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