scholarly journals Context-Dependent Functions of the PII and GlnK Signal Transduction Proteins in Escherichia coli

2002 ◽  
Vol 184 (19) ◽  
pp. 5364-5375 ◽  
Author(s):  
Mariette R. Atkinson ◽  
Timothy A. Blauwkamp ◽  
Alexander J. Ninfa
Keyword(s):  
Escherichia Coli ◽  
Signal Transduction ◽  
Environmental Conditions ◽  
Nitrogen Assimilation ◽  
Cellular Level ◽  
Unknown Mechanism ◽  
Wide Range ◽  
Fine Control ◽  
Context Dependent ◽  
Signal Transduction Proteins

ABSTRACT Two closely related signal transduction proteins, PII and GlnK, have distinct physiological roles in the regulation of nitrogen assimilation. Here, we examined the physiological roles of PII and GlnK when these proteins were expressed from various regulated or constitutive promoters. The results indicate that the distinct functions of PII and GlnK were correlated with the timing of expression and levels of accumulation of the two proteins. GlnK was functionally converted into PII when its expression was rendered constitutive and at the appropriate level, while PII was functionally converted into GlnK by engineering its expression from the nitrogen-regulated glnK promoter. Also, the physiological roles of both proteins were altered by engineering their expression from the nitrogen-regulated glnA promoter. We hypothesize that the use of two functionally identical PII-like proteins, which have distinct patterns of expression, may allow fine control of Ntr genes over a wide range of environmental conditions. In addition, we describe results suggesting that an additional, unknown mechanism may control the cellular level of GlnK.

Carbon-source-dependent nitrogen regulation in Escherichia coli is mediated through glutamine-dependent GlnB signalling

Microbiology ◽  
2003 ◽  
Vol 149 (8) ◽  
pp. 2163-2172 ◽  
Author(s):  
Mani Maheswaran ◽  
Karl Forchhammer
Keyword(s):  
Escherichia Coli ◽  
Signal Transduction ◽  
Carbon Source ◽  
Glutamine Synthetase ◽  
Nitrogen Source ◽  
Carbon Sources ◽  
Nitrogen Status ◽  
Low Concentrations ◽  
Signal Transduction Proteins

The PII signal transduction proteins GlnB and GlnK are uridylylated/deuridylylated in response to the intracellular glutamine level, the primary signal of the cellular nitrogen status. Furthermore, GlnB was shown to be allosterically regulated by 2-oxoglutarate, and thus GlnB was suggested to integrate signals of the cellular carbon and nitrogen status. Receptors of GlnB signal transduction in Escherichia coli are the NtrB/NtrC two-component system and GlnE, an enzyme which adenylylates/deadenylylates glutamine synthetase. In this study, the authors investigated the effect of different carbon sources on the expression of the NtrC-dependent genes glnA and glnK and on the uridylylation status of GlnB and GlnK. With glutamine as nitrogen source, high levels of glnA and glnK expression were obtained when glucose was used as carbon source, but expression was strongly decreased when the cells were grown with poor carbon sources or when cAMP was present. This response correlated with the uridylylation status of GlnB, suggesting that the carbon/cAMP effect was mediated through GlnB uridylylation, a conclusion that was confirmed by mutants of the PII signalling pathway. When glutamine was replaced by low concentrations of ammonium as nitrogen source, neither glnAglnK expression nor GlnB uridylylation responded to the carbon source or to cAMP. Furthermore, glutamine synthetase could be rapidly adenylylated in vivo by the external addition of glutamine; however, this occurred only when cells were grown in the presence of cAMP, not in its absence. Together, these results suggest that poor carbon sources, through cAMP signalling, favour glutamine uptake. The cellular glutamine signal is then transduced by uridylyltransferase and GlnB to modulate NtrC-dependent gene expression.

scholarly journals Mutations Altering the N-Terminal Receiver Domain of NRI (NtrC) That Prevent Dephosphorylation by the NRII-PII Complex in Escherichia coli

2004 ◽  
Vol 186 (17) ◽  
pp. 5730-5740 ◽  
Author(s):  
Augen A. Pioszak ◽  
Alexander J. Ninfa
Keyword(s):  
Escherichia Coli ◽  
Signal Transduction ◽  
Transcriptional Activator ◽  
Conserved Residues ◽  
Receiver Domain ◽  
Phosphorylated Form ◽  
Elevated Expression ◽  
Signal Transduction Proteins ◽  
Altered Proteins

ABSTRACT The phosphorylated form of NRI is the transcriptional activator of nitrogen-regulated genes in Escherichia coli. NRI∼P displays a slow autophosphatase activity and is rapidly dephosphorylated by the complex of the NRII and PII signal transduction proteins. Here we describe the isolation of two mutations, causing the alterations ΔD10 and K104Q in the receiver domain of NRI, that were selected as conferring resistance to dephosphorylation by the NRII-PII complex. The mutations, which alter highly conserved residues near the D54 site of phosphorylation in the NRI receiver domain, resulted in elevated expression of nitrogen-regulated genes under nitrogen-rich conditions. The altered NRI receiver domains were phosphorylated by NRII in vitro but were defective in dephosphorylation. The ΔD10 receiver domain retained normal autophosphatase activity but was resistant to dephosphorylation by the NRII-PII complex. The K104Q receiver domain lacked both the autophosphatase activity and the ability to be dephosphorylated by the NRII-PII complex. The properties of these altered proteins are consistent with the hypothesis that the NRII-PII complex is not a true phosphatase but rather collaborates with NRI≈P to bring about its dephosphorylation.

scholarly journals Sites of Interaction between the FecA and FecR Signal Transduction Proteins of Ferric Citrate Transport in Escherichia coli K-12

2003 ◽  
Vol 185 (13) ◽  
pp. 3745-3752 ◽  
Author(s):  
Sabine Enz ◽  
Heidi Brand ◽  
Claudia Orellana ◽  
Susanne Mahren ◽  
Volkmar Braun
Keyword(s):  
Escherichia Coli ◽  
Signal Transduction ◽  
Transcription Initiation ◽  
Ferric Citrate ◽  
Site Directed Mutagenesis ◽  
Amino Acid Residues ◽  
Mutant Proteins ◽  
Citrate Transport ◽  
Signal Transduction Proteins ◽  
K 12

ABSTRACT Transcription of the fecABCDE ferric citrate transport genes of Escherichia coli K-12 is initiated by a signaling cascade from the cell surface into the cytoplasm. FecR receives the signal in the periplasm from the outer membrane protein FecA loaded with ferric citrate, transmits the signal across the cytoplasmic membrane, and converts FecI in the cytoplasm to an active sigma factor. In this study, it was shown through the use of a bacterial two-hybrid system that, in the periplasm, the C-terminal FecR237-317 fragment interacts with the N-terminal FecA1-79 fragment. In the same C-terminal region, amino acid residues important for the interaction of FecR with FecA were identified by random and site-directed mutagenesis. They were preferentially located in and around a leucine motif (residues 247 to 268) which was found to be highly conserved in FecR-like proteins. The degree of residual binding of FecR mutant proteins to FecA was correlated with the degree of transcription initiation in response to ferric citrate in the culture medium. Three randomly generated inactive FecR mutants, FecR(L254E), FecR(L269G), and FecR(F284L), were suppressed to different degrees by the mutants FecA(G39R) and FecR(D43E). One FecR mutant, FecR (D138E, V197A), induced fecA promoter-directed transcription constitutively in the absence of ferric citrate and bound more strongly than wild-type FecR to FecA. The data showed that FecR interacts in the periplasm with FecA to confer ferric citrate-induced transcription of the fec transport genes and identified sites in FecR and FecA that are important for signal transduction.

scholarly journals Impact of Membrane Phospholipid Alterations in Escherichia coli on Cellular Function and Bacterial Stress Adaptation

2017 ◽  
Vol 199 (13) ◽  
Author(s):  
Veronica W. Rowlett ◽  
Venkata K. P. S. Mallampalli ◽  
Anja Karlstaedt ◽  
William Dowhan ◽  
Heinrich Taegtmeyer ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Environmental Conditions ◽  
Environmental Stresses ◽  
Membrane Phospholipid ◽  
Stress Adaptation ◽  
Cell Physiology ◽  
Membrane Phospholipids ◽  
Content Type ◽  
Bacterial Physiology ◽  
Wide Range

ABSTRACT Bacteria have evolved multiple strategies to sense and rapidly adapt to challenging and ever-changing environmental conditions. The ability to alter membrane lipid composition, a key component of the cellular envelope, is crucial for bacterial survival and adaptation in response to environmental stress. However, the precise roles played by membrane phospholipids in bacterial physiology and stress adaptation are not fully elucidated. The goal of this study was to define the role of membrane phospholipids in adaptation to stress and maintenance of bacterial cell fitness. By using genetically modified strains in which the membrane phospholipid composition can be systematically manipulated, we show that alterations in major Escherichia coli phospholipids transform these cells globally. We found that alterations in phospholipids impair the cellular envelope structure and function, the ability to form biofilms, and bacterial fitness and cause phospholipid-dependent susceptibility to environmental stresses. This study provides an unprecedented view of the structural, signaling, and metabolic pathways in which bacterial phospholipids participate, allowing the design of new approaches in the investigation of lipid-dependent processes involved in bacterial physiology and adaptation. IMPORTANCE In order to cope with and adapt to a wide range of environmental conditions, bacteria have to sense and quickly respond to fluctuating conditions. In this study, we investigated the effects of systematic and controlled alterations in bacterial phospholipids on cell shape, physiology, and stress adaptation. We provide new evidence that alterations of specific phospholipids in Escherichia coli have detrimental effects on cellular shape, envelope integrity, and cell physiology that impair biofilm formation, cellular envelope remodeling, and adaptability to environmental stresses. These findings hold promise for future antibacterial therapies that target bacterial lipid biosynthesis.

The role of effector molecules in signal transduction by PII proteins

2011 ◽  
Vol 39 (1) ◽  
pp. 189-194 ◽  
Author(s):  
Martha Radchenko ◽  
Mike Merrick
Keyword(s):  
Signal Transduction ◽  
Great Majority ◽  
Complex Structures ◽  
Effector Molecules ◽  
Protein Protein Interaction ◽  
Wide Range ◽  
Pii Proteins ◽  
Regulatory Effects ◽  
Signal Transduction Proteins

PII proteins are one of the most widely distributed signal transduction proteins in Nature, being ubiquitous in bacteria, archaea and plants. They act by protein–protein interaction to control the activities of a wide range of enzymes, transcription factors and transport proteins, the great majority of which are involved in cellular nitrogen metabolism. The regulatory activities of PII proteins are mediated through their ability to bind the key effector metabolites 2-OG (2-oxoglutarate), ATP and ADP. However, the molecular basis of these regulatory effects remains unclear. Recent advances in the solution of the crystal structures of PII proteins complexed with some of their target proteins, as well as the identification of the ATP/ADP- and 2-OG-binding sites, have improved our understanding of their mode of action. In all of the complex structures solved to date, the flexible T-loops of PII facilitate interaction with the target protein. The effector molecules appear to play a key role in modulating the conformation of the T-loops and thereby regulating the interactions between PII and its targets.

scholarly journals Sites of Interaction between the FecA and FecR Signal Transduction Proteins of Ferric Citrate Transport in Escherichia coli K-12

2003 ◽  
Vol 185 (21) ◽  
pp. 6494-6494
Author(s):  
Sabine Enz ◽  
Heidi Brand ◽  
Claudia Orellana ◽  
Susanne Mahren ◽  
Uwe H. Stroeher ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Signal Transduction ◽  
Ferric Citrate ◽  
Citrate Transport ◽  
Signal Transduction Proteins ◽  
K 12

An Improved Membrane Filtration Method for Enumeration of Faecal Coliforms and E. Coli by a Fluorogenic β-Glucuronidase Assay

1993 ◽  
Vol 27 (3-4) ◽  
pp. 267-270 ◽  
Author(s):  
M. T. Augoustinos ◽  
N. A. Grabow ◽  
B. Genthe ◽  
R. Kfir
Keyword(s):  
Escherichia Coli ◽  
Water Samples ◽  
Membrane Filtration ◽  
Faecal Coliforms ◽  
Environmental Waters ◽  
Filtration Method ◽  
E Coli ◽  
Wide Range ◽  
Pure Cultures ◽  
Glucuronidase Assay

A fluorogenic β-glucuronidase assay comprising membrane filtration followed by selective enumeration on m-FC agar at 44.5°C and further confirmation using tlie 4-metliylumbelliferyl-β-D-glucuronide (MUG) containing medium was evaluated for the detection of Escherichia coli in water. A total of 200 typical blue and non-typical blue colonies were isolated from sea and fresh water samples using initial selective enumeration on m-FC agar. Pure cultures of the selected colonies were further tested using the MUG assay and identified using the API 20E method. Of the colonies tested which were shown to be positive using the MUG assay 99.4% were Escherichia coli. The results of this study indicate the combination of the m-FC method followed by the MUG assay to be highly efficient for the selection and confirmation of E. coli from a wide range of environmental waters.

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