PTS-Mediated Regulation of the Transcription Activator MtlR from Different Species: Surprising Differences despite Strong Sequence Conservation

2015 ◽  
Vol 25 (2-3) ◽  
pp. 94-105 ◽  
Author(s):  
Philippe Joyet ◽  
Meriem Derkaoui ◽  
Houda Bouraoui ◽  
Josef Deutscher
Keyword(s):  
Bacillus Subtilis ◽  
Lactobacillus Casei ◽  
Sequence Conservation ◽  
Geobacillus Stearothermophilus ◽  
Transcription Activator ◽  
Phosphotransferase System ◽  
Transcription Regulator ◽  
Regulatory Domains ◽  
Genes Encoding ◽  
Enzyme I

The hexitol <smlcap>D</smlcap>-mannitol is transported by many bacteria via a phosphoenolpyruvate (PEP):carbohydrate phosphotransferase system (PTS). In most Firmicutes, the transcription activator MtlR controls the expression of the genes encoding the <smlcap>D</smlcap>-mannitol-specific PTS components and <smlcap>D</smlcap>-mannitol-1-P dehydrogenase. MtlR contains an N-terminal helix-turn-helix motif followed by an Mga-like domain, two PTS regulation domains (PRDs), an EIIB<sup>Gat</sup>- and an EIIA<sup>Mtl</sup>-like domain. The four regulatory domains are the target of phosphorylation by PTS components. Despite strong sequence conservation, the mechanisms controlling the activity of MtlR from <i>Lactobacillus casei</i>, <i>Bacillus subtilis</i> and <i>Geobacillus stearothermophilus</i> are quite different. Owing to the presence of a tyrosine in place of the second conserved histidine (His) in PRD2, <i>L. casei</i> MtlR is not phosphorylated by Enzyme I (EI) and HPr. When the corresponding His in PRD2 of MtlR from <i>B. subtilis</i> and <i>G. stearothermophilus</i> was replaced with alanine, the transcription regulator was no longer phosphorylated and remained inactive. Surprisingly, <i>L. casei</i> MtlR functions without phosphorylation in PRD2 because in a <i>ptsI</i> (EI) mutant MtlR is constitutively active. EI inactivation prevents not only phosphorylation of HPr, but also of the PTS<sup>Mtl</sup> components, which inactivate MtlR by phosphorylating its EIIB<sup>Gat</sup>- or EIIA<sup>Mtl</sup>-like domain. This explains the constitutive phenotype of the <i>ptsI</i> mutant. The absence of EIIB<sup>Mtl</sup>-mediated phosphorylation leads to induction of the <i>L. casei</i><i>mtl </i>operon. This mechanism resembles <i>mtlARFD</i> induction in <i>G. stearothermophilus</i>, but differs from EIIA<sup>Mtl</sup>-mediated induction in <i>B. subtilis</i>. In contrast to <i>B. subtilis</i> MtlR, <i>L. casei</i> MtlR activation does not require sequestration to the membrane via the unphosphorylated EIIB<sup>Mtl</sup> domain.

scholarly journals Distribution and Functions of Phosphotransferase System Genes in the Genome of the Lactic Acid Bacterium Oenococcus oeni

2013 ◽  
Vol 79 (11) ◽  
pp. 3371-3379 ◽  
Author(s):  
Zohra Jamal ◽  
Cécile Miot-Sertier ◽  
François Thibau ◽  
Lucie Dutilh ◽  
Aline Lonvaud-Funel ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Lactic Acid Bacterium ◽  
Oenococcus Oeni ◽  
Phosphotransferase System ◽  
Content Type ◽  
Low Concentrations ◽  
Perfect Adaptation ◽  
Form Part ◽  
Genes Encoding ◽  
Enzyme I

ABSTRACTOenococcus oeni, the lactic acid bacterium primarily responsible for malolactic fermentation in wine, is able to grow on a large variety of carbohydrates, but the pathways by which substrates are transported and phosphorylated in this species have been poorly studied. We show that the genes encoding the general phosphotransferase proteins, enzyme I (EI) and histidine protein (HPr), as well as 21 permease genes (3 isolated ones and 18 clustered into 6 distinct loci), are highly conserved among the strains studied and may form part of theO. oenicore genome. Additional permease genes differentiate the strains and may have been acquired or lost by horizontal gene transfer events. The coreptsgenes are expressed, and permease gene expression is modulated by the nature of the bacterial growth substrate. DecryptifiedO. oenicells are able to phosphorylate glucose, cellobiose, trehalose, and mannose at the expense of phosphoenolpyruvate. These substrates are present at low concentrations in wine at the end of alcoholic fermentation. The phosphotransferase system (PTS) may contribute to the perfect adaptation ofO. oenito its singular ecological niche.

scholarly journals The Lactobacillus casei ptsHI47T Mutation Causes Overexpression of a LevR-Regulated but RpoN-Independent Operon Encoding a Mannose Class Phosphotransferase System

2004 ◽  
Vol 186 (14) ◽  
pp. 4543-4555 ◽  
Author(s):  
Alain Mazé ◽  
Grégory Boël ◽  
Sandrine Poncet ◽  
Ivan Mijakovic ◽  
Yoann Le Breton ◽  
...  
Keyword(s):  
Catabolite Repression ◽  
Lactobacillus Casei ◽  
Constitutive Expression ◽  
Proteome Analysis ◽  
Phosphoryl Group ◽  
Transcription Activator ◽  
Phosphotransferase System ◽  
Reading Frame ◽  
Elevated Expression ◽  
Dna Fragment

ABSTRACT A proteome analysis of Lactobacillus casei mutants that are affected in carbon catabolite repression revealed that a 15-kDa protein was strongly overproduced in a ptsHI47T mutant. This protein was identified as EIIA of a mannose class phosphotransferase system (PTS). A 7.1-kb DNA fragment containing the EIIA-encoding open reading frame and five other genes was sequenced. The first gene encodes a protein resembling the RpoN (σ54)-dependent Bacillus subtilis transcription activator LevR. The following pentacistronic operon is oriented in the opposite direction and encodes four proteins with strong similarity to the proteins of the B. subtilis Lev-PTS and one protein of unknown function. The genes present on the 7.1-kb DNA fragment were therefore called levR and levABCDX. The levABCDX operon was induced by fructose and mannose. No “−12, −24” promoter typical of RpoN-dependent genes precedes the L. casei lev operon, and its expression was therefore RpoN independent but required LevR. Phosphorylation of LevR by P∼His-HPr stimulates its activity, while phosphorylation by P∼EIIBLev inhibits it. Disruption of the EIIBLev-encoding levB gene therefore led to strong constitutive expression of the lev operon, which was weaker in a strain carrying a ptsI mutation preventing phosphorylation by both P∼EIIBLev and P∼His-HPr. Expression of the L. casei lev operon is also subject to P-Ser-HPr-mediated catabolite repression. The observed slow phosphoenolpyruvate- and ATP-dependent phosphorylation of HPrI47T as well as the slow phosphoryl group transfer from the mutant P∼His-HPr to EIIALev are assumed to be responsible for the elevated expression of the lev operon in the ptsHI47T mutant.

scholarly journals Characterization of a Mannose Utilization System in Bacillus subtilis

2010 ◽  
Vol 192 (8) ◽  
pp. 2128-2139 ◽  
Author(s):  
Tianqi Sun ◽  
Josef Altenbuchner
Keyword(s):  
Bacillus Subtilis ◽  
Transcriptional Activation ◽  
Transcriptional Start Site ◽  
Regulatory Gene ◽  
Constitutive Expression ◽  
Fold Increase ◽  
Binding Motif ◽  
Transcription Activator ◽  
Phosphotransferase System ◽  
Sequence Alignments

ABSTRACT The mannose operon of Bacillus subtilis consists of three genes, manP, manA, and yjdF, which are responsible for the transport and utilization of mannose. Upstream and in the same orientation as the mannose operon a regulatory gene, manR, codes for a transcription activator of the mannose operon, as shown in this study. Both mannose operon transcription and manR transcription are inducible by mannose. The presence of mannose resulted in a 4- to 7-fold increase in expression of lacZ from the manP promoter (PmanP ) and in a 3-fold increase in expression of lacZ from the manR promoter (PmanR ). The transcription start sites of manPA-yjdF and manR were determined to be a single A residue and a single G residue, respectively, preceded by −10 and −35 boxes resembling a vegetative σA promoter structure. Through deletion analysis the target sequences of ManR upstream of PmanP and PmanR were identified between bp −80 and −35 with respect to the transcriptional start site of both promoters. Deletion of manP (mannose transporter) resulted in constitutive expression from both the PmanP and PmanR promoters, indicating that the phosphotransferase system (PTS) component EIIMan has a negative effect on regulation of the mannose operon and manR. Moreover, both PmanP and PmanR are subject to carbon catabolite repression (CCR). By constructing protein sequence alignments a DNA binding motif at the N-terminal end, two PTS regulation domains (PRDs), and an EIIA- and EIIB-like domain were identified in the ManR sequence, indicating that ManR is a PRD-containing transcription activator. Like findings for other PRD regulators, the phosphoenolpyruvate (PEP)-dependent phosphorylation by the histidine protein HPr via His15 plays an essential role in transcriptional activation of PmanP and PmanR . Phosphorylation of Ser46 of HPr or of the homologous Crh protein by HPr kinase and formation of a repressor complex with CcpA are parts of the B. subtilis CCR system. Only in the double mutant with an HPr Ser46Ala mutation and a crh knockout mutation was CCR strongly reduced. In contrast, PmanR and PmanP were not inducible in a ccpA deletion mutant.

scholarly journals Bacillus subtilis Mutant LicT Antiterminators Exhibiting Enzyme I- and HPr-Independent Antitermination Affect Catabolite Repression of the bglPH Operon

2002 ◽  
Vol 184 (17) ◽  
pp. 4819-4828 ◽  
Author(s):  
Cordula Lindner ◽  
Michael Hecker ◽  
Dominique Le Coq ◽  
Josef Deutscher
Keyword(s):  
Amino Acids ◽  
Bacillus Subtilis ◽  
Catabolite Repression ◽  
Carbon Catabolite Repression ◽  
Phosphotransferase System ◽  
Terminal Domain ◽  
Target Sites ◽  
Enzyme I ◽  
In Cells

ABSTRACT The Bacillus subtilis antiterminator LicT regulates the expression of bglPH and bglS, which encode the enzymes for the metabolism of aryl-β-glucosides and the β-glucanase BglS. The N-terminal domain of LicT (first 55 amino acids) prevents the formation of ρ-independent terminators on the respective transcripts by binding to target sites overlapping these terminators. Proteins of the phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS) regulate the antitermination activity of LicT by phosphorylating histidines in its two PTS regulation domains (PRDs). Phosphorylation at His-100 in PRD-1 requires the PTS proteins enzyme I and HPr and the phosphorylated permease BglP and inactivates LicT. During transport and phosphorylation of aryl-β-glucosides, BglP is dephosphorylated, which renders LicT active and thus leads to bglPH and bglS induction. In contrast, phosphorylation at His-207 and/or His-269 in PRD-2, which requires only enzyme I and HPr, is absolutely necessary for LicT activity and bglPH and bglS expression. We isolated spontaneous licT mutants expressing bglPH even when enzyme I and HPr were absent (as indicated by the designation “Pia” [PTS-independent antitermination]). Introduced in a ptsHI + strain, two classes of licT(Pia) mutations could be distinguished. Mutants synthesizing LicT(Pia) antiterminators altered in PRD-2 still required induction by aryl-β-glucosides, whereas mutations affecting PRD-1 caused constitutive bglPH expression. One of the two carbon catabolite repression (CCR) mechanisms operative for bglPH requires the ρ-independent terminator and is probably prevented when LicT is activated by P∼His-HPr-dependent phosphorylation in PRD-2 (where the prefix “P∼” stands for “phospho”). During CCR, the small amount of P∼His-HPr present in cells growing on repressing PTS sugars probably leads to insufficient phosphorylation at PRD-2 of LicT and therefore to reduced bglPH expression. In agreement with this concept, mutants synthesizing a P∼His-HPr-independent LicT(Pia) had lost LicT-modulated CCR.

Properties of phosphorylated protein intermediates of the bacterial phosphoenolpyruvate:sugar phosphotransferase system

1992 ◽  
Vol 70 (3-4) ◽  
pp. 242-246 ◽  
Author(s):  
J. W. Anderson ◽  
E. B. Waygood ◽  
M. H. Saier Jr. ◽  
J. Reizer
Keyword(s):  
Escherichia Coli ◽  
Bacillus Subtilis ◽  
Active Site ◽  
Active Sites ◽  
Sugar Transport ◽  
Wild Type ◽  
Phosphotransferase System ◽  
E Coli ◽  
Phosphorylated Protein ◽  
Enzyme I

The phosphohydrolysis properties of the following phosphoprotein intermediates of the bacterial phosphoenolpyruvate:sugar phosphotransferase system (PTS) were investigated: enzyme I, HPr, and the IIAGlc domain of the glucose enzyme II of Bacillus subtilis; and IIAGlc (fast and slow forms) of Escherichia coli. The phosphohydrolysis properties were also studied for the site-directed mutant H68A of B. subtilis IIAGlc. Several conclusions were reached. (i) The phosphohydrolysis properties of the homologous phosphoprotein intermediates of B. subtilis and E. coli are similar. (ii) These properties deviate from those of isolated Nδ1- and Nε2-phosphohistidine indicating the participation of neighbouring residues at the active sites of these proteins. (iii) The rates of phosphohydrolysis of the H68A mutant of B. subtilis IIAGlc were reduced compared with the wild-type protein, suggesting that both His-83 and His-68 are present at the active site of wild-type IIAGlc. (iv) The removal of seven N-terminal residues of E. coli IIAGlc reduced the rates of phosphohydrolysis between pH 5 and 8.Key words: phosphoenolpyruvate:sugar phosphotransferase system, phosphoproteins, phosphohistidine, phosphorylation, sugar transport.

scholarly journals A Novel Role for Enzyme I of the Vibrio cholerae Phosphoenolpyruvate Phosphotransferase System in Regulation of Growth in a Biofilm

2007 ◽  
Vol 190 (1) ◽  
pp. 311-320 ◽  
Author(s):  
Laetitia Houot ◽  
Paula I. Watnick
Keyword(s):  
Vibrio Cholerae ◽  
Sugar Transport ◽  
Phosphotransferase System ◽  
Potent Inducer ◽  
Inducer Exclusion ◽  
Genes Encoding ◽  
Cell Components ◽  
Specific Regulation ◽  
Enzyme I

ABSTRACT Glucose is a universal energy source and a potent inducer of surface colonization for many microbial species. Highly efficient sugar assimilation pathways ensure successful competition for this preferred carbon source. One such pathway is the phosphoenolpyruvate phosphotransferase system (PTS), a multicomponent sugar transport system that phosphorylates the sugar as it enters the cell. Components required for transport of glucose through the PTS include enzyme I, histidine protein, enzyme IIAGlc, and enzyme IIBCGlc. In Escherichia coli, components of the PTS fulfill many regulatory roles, including regulation of nutrient scavenging and catabolism, chemotaxis, glycogen utilization, catabolite repression, and inducer exclusion. We previously observed that genes encoding the components of the Vibrio cholerae PTS were coregulated with the vps genes, which are required for synthesis of the biofilm matrix exopolysaccharide. In this work, we identify the PTS components required for transport of glucose and investigate the role of each of these components in regulation of biofilm formation. Our results establish a novel role for the phosphorylated form of enzyme I in specific regulation of biofilm-associated growth. As the PTS is highly conserved among bacteria, the enzyme I regulatory pathway may be relevant to a number of biofilm-based infections.

scholarly journals Coordinated patterns of cytochrome bd and lactate dehydrogenase expression in Bacillus subtilis

Microbiology ◽  
2005 ◽  
Vol 151 (10) ◽  
pp. 3323-3335 ◽  
Author(s):  
Jonas T. Larsson ◽  
Annika Rogstam ◽  
Claes von Wachenfeldt
Keyword(s):  
Bacillus Subtilis ◽  
Lactate Dehydrogenase ◽  
Core Promoter ◽  
Streptomyces Coelicolor ◽  
Regulatory System ◽  
Upstream Region ◽  
Low Oxygen ◽  
Transcription Activator ◽  
Gene Encoding ◽  
Genes Encoding

A variety of pathways for electron and carbon flow in the soil bacterium Bacillus subtilis are differentially expressed depending on whether oxygen is present in the cell environment. This study characterizes the regulation of the respiratory oxidase cytochrome bd and the NADH-linked fermentative lactate dehydrogenase (LDH). Transcription of the cydABCD operon, encoding cytochrome bd, is highly regulated and only becomes activated at low oxygen availability. This induction is not dependent on the gene encoding the redox regulator Fnr or the genes encoding the ResDE two-component regulatory system. The DNA-binding protein YdiH was found to be a principal regulator that controls cydABCD expression. Transcription from the cyd promoter is stimulated 15-fold by a region located upstream of the core promoter. The upstream region may constitute a binding site for an unidentified transcription activator that is likely to influence the level of transcription but not its timing, which is negatively controlled by YdiH. This report provides evidence that YdiH also functions as a repressor of the ldh gene encoding LDH and of a gene, ywcJ, which encodes a putative formate-nitrite transporter. Based on the similarity between YdiH and the Rex protein of Streptomyces coelicolor, it is proposed that YdiH serves as a redox sensor, the activity of which is regulated by cellular differences in the free levels of NAD+ and NADH. It is suggested that ydiH be renamed as rex.

scholarly journals Cloning and Expression of the Listeria monocytogenes Scott A ptsH and ptsI Genes, Coding for HPr and Enzyme I, Respectively, of the Phosphotransferase System

1998 ◽  
Vol 64 (9) ◽  
pp. 3147-3152 ◽  
Author(s):  
Douglas P. Christensen ◽  
Andrew K. Benson ◽  
Robert W. Hutkins
Keyword(s):  
Listeria Monocytogenes ◽  
High Energy ◽  
Transcription Unit ◽  
Cloning And Expression ◽  
Nucleotide Sequence Analysis ◽  
Phosphotransferase System ◽  
Genes Encoding ◽  
And Control ◽  
Enzyme I

ABSTRACT The phosphoenolpyruvate (PEP)-dependent phosphotransferase system (PTS) utilizes high-energy phosphate present in PEP to drive the uptake of several different carbohydrates in bacteria. In order to examine the role of the PTS in the physiology of Listeria monocytogenes, we identified the ptsH andptsI genes encoding the HPr and enzyme I proteins, respectively, of the PTS. Nucleotide sequence analysis indicated that the predicted proteins are nearly 70% similar to HPr and enzyme I proteins from other organisms. Purified L. monocytogenesHPr overexpressed in Escherichia coli was also capable of complementing an HPr defect in heterologous extracts ofStaphylococcus aureus ptsH mutants. Additional studies of the transcriptional organization and control indicated that theptsH and ptsI genes are organized into a transcription unit that is under the control of a consensus-like promoter and that expression of these genes is mediated by glucose availability and pH or by by-products of glucose metabolism.

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