Influence of Glucose Availability and CRP Acetylation on the Genome-Wide Transcriptional Response of Escherichia coli: Assessment by an Optimized Factorial Microarray Analysis.

Daniel V Guebel,Néstor V Torres

doi:10.3389/fmicb.2018.00941

Abstract

Background: While in eukaryotes acetylation/deacetylation regulation exerts multiple pleiotropic effects, in Escherichia coli it seems to be more limited and less known. Hence, we aimed to progress in the characterization of this regulation by dealing with three convergent aspects: the effector enzymes involved, the master regulator CRP, and the dependence on glucose availability.Methods: The transcriptional response of E. coli BW25113 was analyzed across 14 relevant scenarios. These conditions arise when the wild type and four isogenic mutants (defective in deacetylase CobB, defective in N(ε)-lysine acetyl transferase PatZ, Q- and R-type mutants of protein CRP) are studied under three levels of glucose availability (glucose-limited chemostat and glucose-excess or glucose-exhausted in batch culture). The Q-type emulates a permanent stage of CRPacetylated, whereas the R-type emulates a permanent stage of CRPdeacetylated. The data were analyzed by an optimized factorial microarray method (Q-GDEMAR).Results: (a) By analyzing one mutant against the other, we were able to unravel the true genes that participate in the interaction between ΔcobB/ΔpatZ mutations and glucose availability; (b) Increasing stages of glucose limitation appear to be associated with the up-regulation of specific sets of target genes rather than with the loss of genes present when glucose is in excess; (c) Both CRPdeacetylated and CRPacetylated produce extensive changes in specific subsets of genes, but their number and identity depend on the glucose availability; (d) In other sub-sets of genes, the transcriptional effect of CRP seems to be independent of its acetylation or deacetylation; (e) Some specific ontology functions can be associated with each of the different sets of genes detected herein.Conclusions: CRP cannot be thought of only as an effector of catabolite repression, because it acts along all the glucose conditions tested (excess, limited, and exhausted), exerting both positive and negative effects through different sets of genes. Acetylation of CRP does not seem to be a binary form of regulation, as there is not a univocal relationship between its activation/inhibitory effect and its acetylation/deacetylation stage. All the combinatorial possibilities are observed. During the exponential growth phase, CRP also exerts a very significant transcriptional effect, mainly on flagellar assembly and chemotaxis (FDR = 7.2 × 10−44).

Highlights

It is well known that microbial cells continuously sense their external and internal environment, with this information being filtered and integrated to trigger responses that allow them to best adapt to these stimuli (Snitkin and Segrè, 2008; Zhao et al, 2016)
A ratio value significantly
We note the low values of false discovery rate (FDR)

Summary

Introduction

It is well known that microbial cells continuously sense their external and internal environment, with this information being filtered and integrated to trigger responses that allow them to best adapt to these stimuli (Snitkin and Segrè, 2008; Zhao et al, 2016). Beyond the “short-term adaptations” provided by the allosteric regulations working alongside the extensive set of protein post-translational modifications, changes in the transcriptional responses are the main mechanisms for achieving the “long-term adjustments” required (Shimizu, 2014). There are good reasons to analyse changes in the global transcriptome of microorganisms. To this end, one technique that is widely used is the high-density array of oligonucleotides (microarrays). Transcriptome analysis provides information about the non-coding RNAs (anti-sense RNAs and microRNAs), which have increasing importance as additional layer of regulation at post-transcriptional level, in eukaryotes and in bacteria (Delihas, 2015; Cech et al, 2016; Tronnet et al, 2016). We aimed to progress in the characterization of this regulation by dealing with three convergent aspects: the effector enzymes involved, the master regulator CRP, and the dependence on glucose availability

Objectives

Methods

Results

Conclusion