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Structural basis for sigma factor mimicry in the general stress response of Alphaproteobacteria.

ABSTRACT: Reprogramming gene expression is an essential component of adaptation to changing environmental conditions. In bacteria, a widespread mechanism involves alternative sigma factors that redirect transcription toward specific regulons. The activity of sigma factors is often regulated through sequestration by cognate anti-sigma factors; however, for most systems, it is not known how the activity of the anti-sigma factor is controlled to release the sigma factor. Recently, the general stress response sigma factor in Alphaproteobacteria, ?(EcfG), was identified. ?(EcfG) is inactivated by the anti-sigma factor NepR, which is itself regulated by the response regulator PhyR. This key regulator sequesters NepR upon phosphorylation of its PhyR receiver domain via its ?(EcfG) sigma factor-like output domain (PhyR(SL)). To understand the molecular basis of the PhyR-mediated partner-switching mechanism, we solved the structure of the PhyR(SL)-NepR complex using NMR. The complex reveals an unprecedented anti-sigma factor binding mode: upon PhyR(SL) binding, NepR forms two helices that extend over the surface of the PhyR(SL) subdomains. Homology modeling and comparative analysis of NepR, PhyR(SL), and ?(EcfG) mutants indicate that NepR contacts both proteins with the same determinants, showing sigma factor mimicry at the atomic level. A lower density of hydrophobic interactions, together with the absence of specific polar contacts in the ?(EcfG)-NepR complex model, is consistent with the higher affinity of NepR for PhyR compared with ?(EcfG). Finally, by reconstituting the partner switch in vitro, we demonstrate that the difference in affinity of NepR for its partners is sufficient for the switch to occur.

SUBMITTER: Campagne S

PROVIDER: S-EPMC3361459 | biostudies-literature | 2012 May

REPOSITORIES: biostudies-literature

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Structural basis for sigma factor mimicry in the general stress response of Alphaproteobacteria.

Campagne Sébastien S Damberger Fred F FF Kaczmarczyk Andreas A Francez-Charlot Anne A Allain Frédéric H-T FH Vorholt Julia A JA

Proceedings of the National Academy of Sciences of the United States of America 20120501 21

Reprogramming gene expression is an essential component of adaptation to changing environmental conditions. In bacteria, a widespread mechanism involves alternative sigma factors that redirect transcription toward specific regulons. The activity of sigma factors is often regulated through sequestration by cognate anti-sigma factors; however, for most systems, it is not known how the activity of the anti-sigma factor is controlled to release the sigma factor. Recently, the general stress response ...[more]

PMID: 22550171

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Project description:A core component of the α-proteobacterial general stress response is the extracytoplasmic function (ECF) sigma factor EcfG, exclusively present in this taxonomic class. Half of the completed α-proteobacterial genome sequences contain two or more copies of genes encoding σEcfG-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 σEcfG copies has not satisfactorily been explained. This work explores the hierarchical relation between Rhizobium etli σEcfG1 and σEcfG2, canonical and non-canonical σEcfG proteins, respectively. Contrary to reports in other species, we find that σEcfG1 and σEcfG2 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. σEcfG1 drives expression of rpoH2, explaining the increased heat sensitivity of an ecfG1 mutant, while katG is under control of σEcfG2, accounting for reduced oxidative stress resistance of an ecfG2 mutant. We also identify non-coding RNA genes as novel σEcfG targets. We propose a modified model for general stress response regulation in R. etli, in which σEcfG1 and σEcfG2 function largely independently. Based on a phylogenetic analysis and considering the prevalence of α-proteobacterial genomes with multiple σEcfG 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.

Dataset Information

Structural basis for sigma factor mimicry in the general stress response of Alphaproteobacteria.

Publications

Structural basis for sigma factor mimicry in the general stress response of Alphaproteobacteria.

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