Unknown,Transcriptomics,Genomics,Proteomics

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Stress response regulators identified through genome-wide transcriptome analysis of the stringent response in Rhizobium etli

ABSTRACT: The impact on gene expression by the alarmone (p)ppGpp during growth and non-growth was determined by comparing the transcriptome of R. etli CFN42 wild type and rel mutant. This allowed us to better understand the pleiotropic stress phenotype of the rel mutant as well as identifying specific (p)ppGpp-dependent stress regulators. In order to study expression, R. etli CFN42 wild type and rel mutant was grown at 30˚C in acid minimal salts medium supplied with 10 mM NH4Cl and 10 mM succinate while monitoring the optical density (OD) of the culture. Samples were taken at OD600 = 0.3, 0.7 and 6 hours after reaching the maximum OD, representing early/late exponential and stationary phase, respectively.

ORGANISM(S): Rhizobium etli CFN 42

SUBMITTER: Kristof Engelen

PROVIDER: E-GEOD-23961 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:A core component of the M-NM-1-proteobacterial general stress response is the extracytoplasmic function (ECF) sigma factor EcfG, exclusively present in this taxonomic class. Half of the completed M-NM-1-proteobacterial genome sequences contain two or more copies of genes encoding M-OM-^CEcfG-like sigma factors, with the primary copy typically located adjacent to genes coding for a cognate anti-sigma factor (NepR) and two-component response regulator (PhyR). So far, the widespread occurrence of additional, non-canonical M-OM-^CEcfG copies has not satisfactorily been explained. This work explores the hierarchical relation between Rhizobium etli M-OM-^CEcfG1 and M-OM-^CEcfG2, canonical and non-canonical M-OM-^CEcfG proteins, respectively. Contrary to reports in other species, we find that M-OM-^CEcfG1 and M-OM-^CEcfG2 act in parallel, as nodes of a complex regulatory network, rather than in series, as elements of a linear regulatory cascade. We demonstrate that both sigma factors control unique yet also shared target genes, corroborating phenotypic evidence. M-OM-^CEcfG1 drives expression of rpoH2, explaining the increased heat sensitivity of an ecfG1 mutant, while katG is under control of M-OM-^CEcfG2, accounting for reduced oxidative stress resistance of an ecfG2 mutant. We also identify non-coding RNA genes as novel M-OM-^CEcfG targets. We propose a modified model for general stress response regulation in R. etli, in which M-OM-^CEcfG1 and M-OM-^CEcfG2 function largely independently. Based on a phylogenetic analysis and considering the prevalence of M-NM-1-proteobacterial genomes with multiple M-OM-^CEcfG copies, this model may also be applicable to numerous other species. In this study, we investigate to which degree R. etli EcfG1 and EcfG2 regulation is interdependent. Furthermore, we demonstrate that both sigma factors control distinct regulons and pinpoint unique EcfG2 target genes. We also identify non-coding RNAs (ncRNAs) as novel targets of EcfG1 and EcfG2 and show that expression of at least one of these ncRNAs is under direct EcfG control. The presence of ncRNAs in the regulon of an EcfG type sigma factor has not been described previously and adds an extra level of complexity to the regulation of the general stress response in Alpha-proteobacteria. Moreover, considering the widespread existence of Alpha-proteobacterial genomes with multiple EcfG copies, the here presented results not only lead to a better understanding of the general stress response in R. etli, but also contribute to a conceptual paradigm for general stress response regulation by multiple EcfG proteins in Alpha-proteobacteria. Comparative transcriptome analyses were carried out on four samples: the parental strain (WT) and mutants in either sigma factor gene ecfG1, sigma factor gene ecfG2, and both genes combined.

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Stress response regulators identified through genome-wide transcriptome analysis of the stringent response in Rhizobium etli

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