Global Transcriptional Regulators Fine-Tune the Translational and Metabolic Efficiency for Optimal Growth of Escherichia coli

Abstract

Global transcriptional regulators coordinate complex genetic interactions that bestow better adaptability for an organism against external and internal perturbations. These transcriptional regulators are known to control an enormous array of genes with diverse functionalities. However, regulator-driven molecular mechanisms that underpin precisely tuned translational and metabolic processes conducive for rapid exponential growth remain obscure. Here, we comprehensively reveal the fundamental role of global transcriptional regulators FNR, ArcA, and IHF in sustaining translational and metabolic efficiency under glucose fermentative conditions in Escherichia coli. By integrating high-throughput gene expression profiles and absolute intracellular metabolite concentrations, we illustrate that these regulators are crucial in maintaining nitrogen homeostasis, govern expression of otherwise unnecessary or hedging genes, and exert tight control on metabolic bottleneck steps. Furthermore, we characterize changes in expression and activity profiles of other coregulators associated with these dysregulated metabolic pathways, determining the regulatory interactions within the transcriptional regulatory network. Such systematic findings emphasize their importance in optimizing the proteome allocation toward metabolic enzymes as well as ribosomes, facilitating condition-specific phenotypic outcomes. Consequentially, we reveal that disruption of this inherent trade-off between ribosome and metabolic proteome economy due to the loss of regulators resulted in lowered growth rates. Moreover, our findings reinforce that the accumulations of intracellular metabolites in the event of proteome repartitions negatively affects the glucose uptake. Overall, by extending the three-partition proteome allocation theory concordant with multi-omics measurements, we elucidate the physiological consequences of loss of global regulators on central carbon metabolism restraining the organism to attain maximal biomass synthesis. IMPORTANCE Cellular proteome allocation in response to environmental or internal perturbations governs their eventual phenotypic outcome. This entails striking an effective balance between amino acid biosynthesis by metabolic proteins and its consumption by ribosomes. However, the global transcriptional regulator-driven molecular mechanisms that underpin their coordination remains unexplored. Here, we emphasize that global transcriptional regulators, known to control expression of a myriad of genes, are fundamental for precisely tuning the translational and metabolic efficiencies that define the growth optimality. Towards this, we systematically characterized the single deletion effect of FNR, ArcA, and IHF regulators of Escherichia coli on exponential growth under anaerobic glucose fermentative conditions. Their absence disrupts the stringency of proteome allocation, which manifests as impairment in key metabolic processes and an accumulation of intracellular metabolites. Furthermore, by incorporating an extension to the empirical growth laws, we quantitatively demonstrate the general design principles underlying the existence of these regulators in E. coli.

Global pleiotropic effects in adaptively evolved Escherichia coli lacking CRP reveal molecular mechanisms that define the growth physiology

“…Despite the huge repository of data available for CRP, several questions are still unanswered. As changes in carbon transport rate can only in part explain the changes in growth physiology [32,33], synthesis of precursors or biomass components by metabolic enzymes and ribosomes, and their attuned efficiency coordinated by CRP, remain obscure. In addition, delineating the CRP regulation-specific effects on these molecular mechanisms from those mediated by changes in growth rate becomes imperative.…”

A systems biology approach to disentangle the direct and indirect effects of global transcription factors on gene expression in Escherichia coli

“…Here, we characterized and quantified the pleiotropic effects on gene expression coordinated by the TFs as well as growth-rate dependent global machinery (RNA polymerase and intracellular metabolites) under exponential growth of the bacteria. Overall, we found across the strains lacking FNR, ArcA, or IHF and their combinations thereof, that a greater percentage (>45%) of DEGs were associated with the direct effects of the knocked-out TF as opposed to earlier estimates (~17%) (Iyer et al, 2021). This was determined from the available TF binding data, novel genes wherein the upstream DNA binding sequence was predicted with good confidence and chemostat based gene expression analyses.…”

“…However, the gene expression of the mutants FI and AI have not been characterized so far. The gene expression data for the single mutants were obtained from our previous study (Iyer et al, 2021). We investigated the mode of interaction between each of the regulators in the double mutant, specifically, whether the expression patterns could be explained by either of the deleted TFs or by combined pleiotropic effects of both the regulators, defined hereafter as 'crosstalk'.…”

“…In our previous work, we demonstrated the systems-wide effect of the transcriptional regulators FNR, ArcA, and IHF on key metabolic pathways (Iyer et al, 2021). To decipher whether those metabolic pathways were perturbed in the double mutants as well, we carried out a KEGG pathway enrichment analysis (Proteomaps (Liebermeister et al, 2014) based gene classification) on DEGs obtained in double mutants with statistical significance analysis.…”

“…All phenotypic characterizations were performed in three biological replicates (n = 3) as described previously (Iyer et al, 2021). Extracellular supernatants collected from the entire exponential growth phase were used to determine the rates of glucose uptake as well as the rates of secretion of extracellular mixed acid fermentation metabolites such as acetate, lactate, pyruvate, succinate, formate, and ethanol, and the yields thereafter.…”