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Partial bacterial genome reduction by genome engineering can improve the productivity of various metabolites, possibly via deletion of non-essential genome regions involved in undesirable metabolic pathways competing with pathways for the desired end products. However, such reduction may cause growth defects. Genome reduction of Bacillus subtilis MGB874 increases the productivity of cellulases and proteases but reduces their growth rate. Here, we show that this growth defect could be restored by silencing redundant or less important genes affecting exponential growth by manipulating the global transcription factor AbrB. Comparative transcriptome analysis revealed that AbrB-regulated genes were upregulated and those involved in central metabolic pathway and synthetic pathways of amino acids and purine/pyrimidine nucleotides were downregulated in MGB874 compared with the wild-type strain, which we speculated were the cause of the growth defects. By constitutively expressing high levels of AbrB, AbrB regulon genes were repressed, while glycolytic flux increased, thereby restoring the growth rate to wild-type levels. This manipulation also enhanced the productivity of metabolites including γ-polyglutamic acid. This study provides the first evidence that undesired features induced by genome reduction can be relieved, at least partly, by manipulating a global transcription regulation system. A similar strategy could be applied to other genome engineering-based challenges aiming toward efficient material production in bacteria. [ABSTRACT FROM AUTHOR]

To construct a prototrophic Corynebacterium glutamicum strain that efficiently produces pyruvate from glucose, the effects of inactivating RamA, a global regulator responsible for activating the oxidative tricarboxylic acid (TCA) cycle, on glucose metabolism were investigated. ΔramA showed an increased specific glucose consumption rate, decreased growth, comparable pyruvate production, higher formation of lactate and acetate, and lower accumulation of succinate and 2-oxoglutarate compared to the wild type. A significant decrease in pyruvate dehydrogenase complex activity was observed for ΔramA, indicating reduced carbon flow to the TCA cycle in ΔramA. To create an efficient pyruvate producer, the ramA gene was deleted in a strain lacking the genes involved in all known lactate- and acetate-producing pathways. The resulting mutant produced 161 mM pyruvate from 222 mM glucose, which was significantly higher than that of the parent (89.3 mM; 1.80-fold). [ABSTRACT FROM AUTHOR]

Symbiotic interactions are indispensable for metazoan function, but their origin and evolution remain elusive. We use a controlled evolution experiment to demonstrate the emergence of novel commensal interactions between Pseudomonas aeruginosa, an initially pathogenic bacterium, and a metazoan host, Caenorhabditis elegans. We show that commensalism evolves through loss of virulence, because it provides bacteria with a double fitness advantage: Increased within-host fitness and a larger host population to infect. Commensalism arises irrespective of host immune status, as the adaptive path in immunocompromised C. elegans knockouts does not differ from that in wild type. Dissection of temporal dynamics of genomic adaptation for 125 bacterial populations reveals highly parallel evolution of incipient commensalism across independent biological replicates. Adaptation is mainly achieved through frame shift mutations in the global regulator lasR and nonsynonymous point mutations in the polymerase gene rpoB that arise early in evolution. Genetic knockouts of lasR not only corroborate its role in virulence attenuation but also show that further mutations are necessary for the fully commensal phenotype. The evolutionary transition from pathogenicity to commensalism as we observe here is facilitated by mutations in global regulators such as lasR, because few genetic changes cause pleiotropic effects across the genome with large phenotypic effects. Finally, we found that nucleotide diversity increased more quickly in bacteria adapting to immunocompromised hosts than in those adapting to immunocompetent hosts. Nevertheless, the outcome of evolution was comparable across host types. Commensalism can thus evolve independently of host immune state solely as a side-effect of bacterial adaptation to novel hosts. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa is one of the most intractable human pathogens that pose serious clinical challenge due to extensive prevalence of multidrug-resistant clinical isolates. Armed with abundant virulence and antibiotic resistance mechanisms, it is a major etiologic agent in a number of acute and chronic infections. A complex and intricate network of regulators dictates the expression of pathogenicity factors in P. aeruginosa. Some proteins within the network play key roles and control multiple pathways. This review discusses the role of one such protein, AmpR, which was initially recognized for its role in antibiotic resistance by regulating AmpC β-lactamase. Recent genomic, proteomic and phenotypic analyses demonstrate that AmpR regulates expression of hundreds of genes that are involved in diverse pathways such as β-lactam and non-β-lactam resistance, quorum sensing and associated virulence phenotypes, protein phosphorylation, and physiological processes. Finally, ampR mutations in clinical isolates are reviewed to shed light on important residues required for its function in antibiotic resistance. The prevalence and evolutionary implications of AmpR in pathogenic and nonpathogenic proteobacteria are also discussed. A comprehensive understanding of proteins at nodal positions in the P. aeruginosa regulatory network is crucial in understanding, and ultimately targeting, the pathogenic stratagems of this organism. [ABSTRACT FROM AUTHOR]

Recent data indicate that cell cycle transcription in many alpha-Proteobacteria is executed by at least three conserved functional modules in which pairs of antagonistic regulators act jointly, rather than in isolation, to control transcription in S-, G2- or G1-phase. Inactivation of module components often results in pleiotropic defects, ranging from cell death and impaired cell division to fairly benign deficiencies in motility. Expression of module components can follow systemic (cell cycle) or external (nutritional/cell density) cues and may be implemented by auto-regulation, ancillary regulators or other (unknown) mechanisms. Here, we highlight the recent progress in understanding the molecular events and the genetic relationships of the module components in environmental, pathogenic and/or symbiotic alpha-proteobacterial genera. Additionally, we take advantage of the recent genome-wide transcriptional analyses performed in the model alpha-Proteobacterium Caulobacter crescentus to illustrate the complexity of the interactions of the global regulators at selected cell cycle-regulated promoters and we detail the consequences of (mis-)expression when the regulators are absent. This review thus provides the first detailed mechanistic framework for understanding orthologous operational principles acting on cell cycle-regulated promoters in other alpha-Proteobacteria. [ABSTRACT FROM AUTHOR]

The vast majority of experimental data have been accumulated on the transcription regulation of individual genes within a single model prokaryote, Escherichia coli, which form the well-established on–off switch model of transcription by DNA-binding regulatory proteins. After the development of modern high-throughput experimental systems such as microarray analysis of whole genome transcription and the Genomic SELEX search for the whole set of regulation targets by transcription factors, a number of E. coli promoters are now recognized to be under the control of multiple transcription factors, as in the case of eukaryotes. The number of regulation targets of a single transcription factor has also been found to be more than hitherto recognized, ranging up to hundreds of promoters, genes or operons for several global regulators. The multifactor promoters and the multitarget transcription factors can be assembled into complex networks of transcription regulation, forming hierarchical networks. [ABSTRACT FROM AUTHOR]

In this study, we investigated the genetic organization and function of Escherichia coli yciT, a gene predicted by computational methods to belong to the DeoR-type family of transcriptional regulators. We show that transcription of yciT (here denoted deoT for deoR-Type) initiates from a promoter located upstream of a putative open reading frame (denoted deoL for deoT Leader). We also show that DeoT acts as a global regulator, repressing the expression of a number of genes involved in a variety of metabolic pathways including transport of maltose, fatty acid β-oxidation and peptide degradation. [ABSTRACT FROM AUTHOR]

We have shown that the bldD gene of Streptomyces coelicolor, while required for antibiotic production and morphological differentiation, is not essential for viability. We have also demonstrated that BldD forms a higher order complex both in solution and when bound to target DNA. Purified BldD exists in three forms in solution, as a tetramer, dimer and monomer, but only in the dimeric form when bound to its own promoter/operator. [Copyright &y& Elsevier]