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Expression profiling reveals Spot 42 small RNA as a key regulator in the central metabolism of Aliivibrio salmonicida.

ABSTRACT: BACKGROUND: Spot 42 was discovered in Escherichia coli nearly 40 years ago as an abundant, small and unstable RNA. Its biological role has remained obscure until recently, and is today implicated in having broader roles in the central and secondary metabolism. Spot 42 is encoded by the spf gene. The gene is ubiquitous in the Vibrionaceae family of gamma-proteobacteria. One member of this family, Aliivibrio salmonicida, causes cold-water vibriosis in farmed Atlantic salmon. Its genome encodes Spot 42 with 84% identity to E. coli Spot 42. RESULTS: We generated a A. salmonicida spf deletion mutant. We then used microarray and Northern blot analyses to monitor global effects on the transcriptome in order to provide insights into the biological roles of Spot 42 in this bacterium. In the presence of glucose, we found a surprisingly large number of ? 2X differentially expressed genes, and several major cellular processes were affected. A gene encoding a pirin-like protein showed an on/off expression pattern in the presence/absence of Spot 42, which suggests that Spot 42 plays a key regulatory role in the central metabolism by regulating the switch between fermentation and respiration. Interestingly, we discovered an sRNA named VSsrna24, which is encoded immediately downstream of spf. This new sRNA has an expression pattern opposite to that of Spot 42, and its expression is repressed by glucose. CONCLUSIONS: We hypothesize that Spot 42 plays a key role in the central metabolism, in part by regulating the pyruvat dehydrogenase enzyme complex via pirin.

SUBMITTER: Hansen GA

PROVIDER: S-EPMC3295665 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Expression profiling reveals Spot 42 small RNA as a key regulator in the central metabolism of Aliivibrio salmonicida.

Hansen Geir A GA Ahmad Rafi R Hjerde Erik E Fenton Christopher G CG Willassen Nils-Peder NP Haugen Peik P

BMC genomics 20120124

<h4>Background</h4>Spot 42 was discovered in Escherichia coli nearly 40 years ago as an abundant, small and unstable RNA. Its biological role has remained obscure until recently, and is today implicated in having broader roles in the central and secondary metabolism. Spot 42 is encoded by the spf gene. The gene is ubiquitous in the Vibrionaceae family of gamma-proteobacteria. One member of this family, Aliivibrio salmonicida, causes cold-water vibriosis in farmed Atlantic salmon. Its genome enco ...[more]

PMID: 22272603

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Project description:Spot 42 was discovered in Escherichia coli nearly 40 years ago as an abundant, small and unstable RNA. Its biological role remained obscure until it was shown to cause discoordinate expression of the galactose operon (gal operon). Recently Spot 42 has also been implicated in having broader roles in the central and secondary metabolism. Spot 42 is encoded by the spf gene. The gene is ubiquitous in the Vibrionaceae family of gamma-proteobacteria, which includes a number of serious pathogens of humans and animals, including the infamous Vibrio cholerae. One member of this family, Aliivibrio salmonicida, causes cold-water vibriosis in farmed Atlantic salmon. Its genome encodes Spot 42 with 84 % identity to E. coli Spot 42 (spf). We generated a A. salmonicida spf deletion mutant. We then and used microarray and Northern blot analyses to monitor global effects on the transcriptome in order to provide insights into the biological roles of Spot 42 in this bacterium. In the presence of glucose we found a surprisingly large number of ≥2× differentially expressed genes, and several major cellular processes were affected, such as carbohydrate metabolism and transport, motility and chemotaxis, iron homeostasis and quorum sensing. A gene encoding a pirin-like protein (VSAL_I1200) showed an on/off expression pattern in the presence/absence of Spot 42, which suggests that Spot 42 plays a key regulatory role in the central metabolism by regulating the switch between fermentation and respiration. Interestingly, in a global search we discovered a sRNA, named VSsrna24, which is encoded immediately downstream of spf. This new sRNA has an expression pattern opposite to that of Spot 42, and its expression is highly dependent on glucose. Our hypothesis is that this novel sRNA works in concert with Spot 42 to regulate carbohydrate metabolism and uptake. Two-condition experiment, wild type cells (control samples) vs. spf mutants grown in A. salmonicida-specific minimal media (ASMM) (stimulated samples). Samples were prepared by culturing the A. salmonicida wild type and a spf deletion mutant strains at 12 °C in ASMM without any added carbon source to OD 600nm 0.4. At this density glucose to 44.4 mM was added. Samples were collected 15 min after stimulation. Biological replicates for each sample: 3 wild type, 3 spf mutant, independently grown and harvested. One replicate per array.

Project description:BackgroundThe coordination of group behaviors in bacteria is achieved by a cell-cell signaling process called quorum sensing (QS). QS is an intercellular communication system, which synchronously controls expression of a vast range of genes in response to changes in cell density and is mediated by autoinducers that act as extracellular signals. Aliivibrio salmonicida, the causative agent of cold-water vibrosis in marine aquacultures, uses QS to regulate several activities such as motility, biofilm formation, adhesion and rugose colony morphology. However, little is known about either genes or detailed mechanisms involved in the regulation of these phenotypes.ResultsDifferential expression profiling allowed us to define the genes involved in controlling phenotypes related to QS in A. salmonicida LFI1238. RNA sequencing data revealed that the number of expressed genes in A. salmonicida, ΔlitR and ΔrpoQ mutants were significantly altered due to changes in cell density. These included genes that were distributed among the 21 functional groups, mainly presented in cell envelope, cell processes, extrachromosomal/foreign DNA and transport-binding proteins functional groups. The comparative transcriptome of A. salmonicida wild-type at high cell density relative to low cell density revealed 1013 genes to be either up- or downregulated. Thirty-six downregulated genes were gene clusters encoding biosynthesis of the flagellar and chemotaxis genes. Additionally we identified significant expression for genes involved in acyl homoserine lactone (AHL) synthesis, adhesion and early colonization. The transcriptome profile of ΔrpoQ compared to the wild-type revealed 384 differensially expressed genes (DEGs) that allowed us to assign genes involved in regulating motility, adhesion and colony rugosity. Indicating the importance of RpoQ in controlling several QS related activities. Furthermore, the comparison of the transcriptome profiles of ΔlitR and ΔrpoQ mutants, exposed numerous overlapping DEGs that were essential for motility, exopolysaccharide production via syp operon and genes associated with tad operon.ConclusionOur findings indicate previously unexplained functional roles for LitR and RpoQ in regulation of different phenotypes related to QS. Our transcriptome data provide a better understanding of the regulation cascade of motility, wrinkling colony morphology and biofilm formation and will offer a major source for further research and analysis on this important field.

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Expression profiling reveals Spot 42 small RNA as a key regulator in the central metabolism of Aliivibrio salmonicida.

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Expression profiling reveals Spot 42 small RNA as a key regulator in the central metabolism of Aliivibrio salmonicida.

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