Heat shock response in bacteria with large genomes: lessons from rhizobia

Rhizobia are important soil bacteria due to their ability to establish nitrogen-fixing symbioses with legume plants. In this dual lifestyle, as free-living bacteria or as plant symbiont, rhizobia are often exposed to different environmental stresses. The present chapter overviews the current knowledge on the heat shock response of rhizobia, highlighting how these large genome bacteria respond to heat from a transcriptional point of view. Response to heat shock in rhizobia involves genome wide changes in the transcriptome that may affect more than 30% of the genome and involve all replicons. In addition to the expected upregulation of genes already known to be involved in stress response (dnaK, groEL, ibpA, clpB), the reports on the heat shock response in rhizobia also showed particular aspects of stress response in these resourceful bacteria. The transcriptional response to heat in rhizobia includes the overexpression of a large number of genes involved in transcription and carbohydrate transport and metabolism. Additional studies are needed in order to better understand the transcriptional regulation of stress response in bacteria with large genomes

The route to transcription initiation determines the mode of transcriptional bursting in E. coli

Transcription is fundamentally noisy, leading to significant heterogeneity across bacterial populations. Noise is often attributed to burstiness, but the underlying mechanisms and their dependence on the mode of promotor regulation remain unclear. Here, we measure E. coli single cell mRNA levels for two stress responses that depend on bacterial sigma factors with different mode of transcription initiation (σ70 and σ54). By fitting a stochastic model to the observed mRNA distributions, we show that the transition from low to high expression of the σ70-controlled stress response is regulated via the burst size, while that of the σ54-controlled stress response is regulated via the burst frequency. Therefore, transcription initiation involving σ54 differs from other bacterial systems, and yields bursting kinetics characteristic of eukaryotic systems

Scoring Targets of Transcription in Bacteria Rather than Focusing on Individual Binding Sites

Reliable identification of targets of bacterial regulators is necessary to understand bacterial gene expression regulation. These targets are commonly predicted by searching for high-scoring binding sites in the upstream genomic regions, which typically leads to a large number of false positives. In contrast to the common approach, here we propose a novel concept, where overrepresentation of the scoring distribution that corresponds to the entire searched region is assessed, as opposed to predicting individual binding sites. We explore two implementations of this concept, based on Kolmogorov–Smirnov (KS) and Anderson–Darling (AD) tests, which both provide straightforward P-value estimates for predicted targets. This approach is implemented for pleiotropic bacterial regulators, including σ70 (bacterial housekeeping σ factor) target predictions, which is a classical bioinformatics problem characterized by low specificity. We show that KS based approach is both faster and more accurate, departing from the current paradigm of AD being slower, but more accurate. Moreover, KS approach leads to a significant increase in the search accuracy compared to the standard approach, while at the same time straightforwardly assigning well established P-values to each potential target. Consequently, the new KS based method proposed here, which assigns P-values to fixed length upstream regions, provides a fast and accurate approach for predicting bacterial transcription targets

Structures of RNA Polymerase Closed and Intermediate Complexes Reveal Mechanisms of DNA Opening and Transcription Initiation.

Gene transcription is carried out by RNA polymerases (RNAPs). For transcription to occur, the closed promoter complex (RPc), where DNA is double stranded, must isomerize into an open promoter complex (RPo), where the DNA is melted out into a transcription bubble and the single-stranded template DNA is delivered to the RNAP active site. Using a bacterial RNAP containing the alternative σ54 factor and cryoelectron microscopy, we determined structures of RPc and the activator-bound intermediate complex en route to RPo at 3.8 and 5.8 Å. Our structures show how RNAP-σ54 interacts with promoter DNA to initiate the DNA distortions required for transcription bubble formation, and how the activator interacts with RPc, leading to significant conformational changes in RNAP and σ54 that promote RPo formation. We propose that DNA melting is an active process initiated in RPc and that the RNAP conformations of intermediates are significantly different from that of RPc and RPo