Dataset Information

Proteome analysis of Escherichia coli ptsN-null strain under different nitrogen regimes

ABSTRACT: The carbohydrate-uptake phosphorelay PTS system plays a key role in metabolic regulation in Bacteria controlling the utilization of secondary carbon sources. Some Bacteria, such as Escherichia coli, encode a paralogue system named PTSNtr (nitrogen related PTS). PTSNtr is composed of EINtr (ptsP), NPr (ptsO), and EIIANtr (ptsN). These proteins act as a phosphorely system transfering phosphoryl from phosphoenolpyruvate to EINtr, NPr and them to EIIANtr. PTSNtr is not involved in carbohydrate uptake and it may be dedicated to play regulatory functions. The phosphorylation state of the EINtr is regulated by allosteric binding of glutamine and 2-oxoglutarate, metabolites whose intracellular levels reflect the nitrogen levels. Despite the denomination and nitrogen-sensory properties no major effect of PTSNtr on nitrogen regulation has been described in E. coli. Here we show that an E. coli ∆ptsN has impaired growth in minimal medium. Proteome analysis of the ∆ptsN strain under different nitrogen regimes revealed that ptsN is not involved in the regulation of the canonical nitrogen starvation response. The proteomic data support that ptsN is required for proper balance between RpoS and sigma 70 activities such that, in the absence of ptsN, RpoS-depended genes are preferentially expressed.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Escherichia Coli

SUBMITTER: Gustavo De Souza

LAB HEAD: Luciano Huergo

PROVIDER: PXD008034 | Pride | 2018-09-13

REPOSITORIES: Pride

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Proteome analysis of an Escherichia coli ptsN-null strain under different nitrogen regimes.

Gravina Fernanda F Sanchuki Heloisa S HS Rodrigues Thiago E TE Gerhardt Edileusa C M ECM Pedrosa Fábio O FO Souza Emanuel M EM Valdameri Gláucio G de Souza Gustavo A GA Huergo Luciano F LF

Journal of proteomics 20171221

The carbohydrate-uptake phosphorelay PTS system plays a key role in metabolic regulation in Bacteria controlling the utilization of secondary carbon sources. Some bacteria, such as Escherichia coli, encode a paralogous system named PTSNtr (nitrogen related PTS). PTSNtr is composed of EINtr (ptsP), NPr (ptsO), and EIIANtr (ptsN). These proteins act as a phosphorelay system from phosphoenolpyruvate to EINtr, NPr and them to EIIANtr. ...[more]

PMID: 29274402

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Project description:Nishio2008 - Design of the phosphotransferase system for enhanced glucose uptake in E. coli. This model is described in the article: Computer-aided rational design of the phosphotransferase system for enhanced glucose uptake in Escherichia coli. Nishio Y, Usuda Y, Matsui K, Kurata H. Mol. Syst. Biol. 2008; 4: 160 Abstract: The phosphotransferase system (PTS) is the sugar transportation machinery that is widely distributed in prokaryotes and is critical for enhanced production of useful metabolites. To increase the glucose uptake rate, we propose a rational strategy for designing the molecular architecture of the Escherichia coli glucose PTS by using a computer-aided design (CAD) system and verified the simulated results with biological experiments. CAD supports construction of a biochemical map, mathematical modeling, simulation, and system analysis. Assuming that the PTS aims at controlling the glucose uptake rate, the PTS was decomposed into hierarchical modules, functional and flux modules, and the effect of changes in gene expression on the glucose uptake rate was simulated to make a rational strategy of how the gene regulatory network is engineered. Such design and analysis predicted that the mlc knockout mutant with ptsI gene overexpression would greatly increase the specific glucose uptake rate. By using biological experiments, we validated the prediction and the presented strategy, thereby enhancing the specific glucose uptake rate. This model is hosted on BioModels Database and identified by: BIOMD0000000571. To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models. To the extent possible under law, all copyright and related or neighbouring rights to this encoded model have been dedicated to the public domain worldwide. Please refer to CC0 Public Domain Dedication for more information.

Project description:RpoS is a conserved stress regulator that plays a critical role in survival under stress conditions in Escherichia coli and other γ-proteobacteria. RpoS is also involved in virulence of many pathogens including Salmonella and Vibrio species. Though well characterized in non-pathogenic E. coli K12 strains, the effect of RpoS on transcriptome expression has not been examined in pathogenic isolates. E. coli O157:H7 is a serious human enteropathogen, possessing a genome 20% larger than that of E. coli K12, and many of the additional genes are required for virulence. The genomic difference may result in substantial changes in RpoS-regulated gene expression. To test this, we compared the transcriptional profile of wild type and rpoS mutants of the E. coli O157:H7 EDL933 type strain. The rpoS mutation had a pronounced effect on gene expression in stationary phase, and more than 1,000 genes were differentially expressed (two-fold, p<0.05). By contrast, we found 11 genes expressed differently in exponential phase. Western blot analysis revealed that, as expected, RpoS level was low in exponential phase and substantially increased in stationary phase. The defect in rpoS resulted in impaired expression of genes responsible for stress response (e.g., gadA, katE and osmY), arginine degradation (astCADBE), putrescine degradation (puuABCD), fatty acid oxidation (fadBA and fadE), and virulence (ler, espI and cesF). For EDL933-specific genes on O-islands, we found 50 genes expressed higher in wild type EDL933 and 49 genes expressed higher in the rpoS mutants. The protein levels of Tir and EspA, two LEE-encoded virulence factors, were elevated in the rpoS mutants under LEE induction conditions. Our results show that RpoS has a profound effect on global gene expression in the pathogenic strain O157:H7 EDL933, and the identified RpoS regulon, including many EDL933-specific genes, differs substantially from that of laboratory K12 strains. In this study, we characterized the RpoS regulon of E. coli O157:H7 strain EDL933 using microarray analysis.

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