scholarly journals Autoinduction of Bacillus subtilis phoPR Operon Transcription Results from Enhanced Transcription from EσA- and EσE-Responsive Promoters by Phosphorylated PhoP

2004 ◽  
Vol 186 (13) ◽  
pp. 4262-4275 ◽  
Author(s):  
Salbi Paul ◽  
Stephanie Birkey ◽  
Wei Liu ◽  
F. Marion Hulett
Keyword(s):  
Bacillus Subtilis ◽  
Response Regulator ◽  
Phosphate Starvation ◽  
Growth Conditions ◽  
In Vitro Transcription ◽  
Phosphate Deficiency ◽  
Pho Regulon ◽  
Environmental Controls ◽  
Rna Polymerase Holoenzyme

ABSTRACT The phoPR operon encodes a response regulator, PhoP, and a histidine kinase, PhoR, which activate or repress genes of the Bacillus subtilis Pho regulon in response to an extracellular phosphate deficiency. Induction of phoPR upon phosphate starvation required activity of both PhoP and PhoR, suggesting autoregulation of the operon, a suggestion that is supported here by PhoP footprinting on the phoPR promoter. Primer extension analyses, using RNA from JH642 or isogenic sigE or sigB mutants isolated at different stages of growth and/or under different growth conditions, suggested that expression of the phoPR operon represents the sum of five promoters, each responding to a specific growth phase and environmental controls. The temporal expression of the phoPR promoters was investigated using in vitro transcription assays with RNA polymerase holoenzyme isolated at different stages of Pho induction, from JH642 or isogenic sigE or sigB mutants. In vitro transcription studies using reconstituted EσA, EσB, and EσE holoenzymes identified PA4 and PA3 as EσA promoters and PE2 as an EσE promoter. Phosphorylated PhoP (PhoP∼P) enhanced transcription from each of these promoters. EσB was sufficient for in vitro transcription of the PB1 promoter. P5 was active only in a sigB mutant strain. These studies are the first to report a role for PhoP∼P in activation of promoters that also have activity in the absence of Pho regulon induction and an activation role for PhoP∼P at an EσE promoter. Information concerning PB1 and P5 creates a basis for further exploration of the regulatory coordination or overlap of the PhoPR and SigB regulons during phosphate starvation.

scholarly journals Transcriptional Regulation of the phoPR Operon in Bacillus subtilis

2004 ◽  
Vol 186 (4) ◽  
pp. 1182-1190 ◽  
Author(s):  
Zoltán Prágai ◽  
Nicholas E. E. Allenby ◽  
Nicola O'Connor ◽  
Sarah Dubrac ◽  
Georges Rapoport ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Stationary Phase ◽  
Stress Responses ◽  
Response Regulator ◽  
Constitutive Expression ◽  
Phosphate Starvation ◽  
Pho Regulon ◽  
Dependent Manner ◽  
Low Level

ABSTRACT When Bacillus subtilis is subjected to phosphate starvation, the Pho regulon is activated by the PhoP-PhoR two-component signal transduction system to elicit specific responses to this nutrient limitation. The response regulator, PhoP, and its cognate histidine sensor kinase, PhoR, are encoded by the phoPR operon that is transcribed as a 2.7-kb bicistronic mRNA. The phoPR operon is transcribed from two σA-dependent promoters, P1 and P2. Under conditions where the Pho regulon was not induced (i.e., phosphate-replete conditions or phoR-null mutant), a low level of phoPR transcription was detected only from promoter P1. During phosphate starvation-induced transition from exponential to stationary phase, the expression of the phoPR operon was up-regulated in a phosphorylated PhoP (PhoP∼P)-dependent manner; in addition to P1, the P2 promoter becomes active. In vitro gel shift assays and DNase I footprinting experiments showed that both PhoP and PhoP∼P could bind to the control region of the phoPR operon. The data indicate that while low-level constitutive expression of phoPR is required under phosphate-replete conditions for signal perception and transduction, autoinduction is required to provide sufficient PhoP∼P to induce other members of the Pho regulon. The extent to which promoters P1 and P2 are activated appears to be influenced by the presence of other sigma factors, possibly the result of sigma factor competition. For example, phoPR is hyperinduced in a sigB mutant and, later in stationary phase, in sigH, sigF, and sigE mutants. The data point to a complex regulatory network in which other stress responses and post-exponential-phase processes influence the expression of phoPR and, thereby, the magnitude of the Pho regulon response.

scholarly journals Transcriptional Activation of the Bacillus subtilis spoIIG Promoter by the Response Regulator Spo0A Is Independent of the C-Terminal Domain of the RNA Polymerase Alpha Subunit

1998 ◽  
Vol 180 (17) ◽  
pp. 4760-4763 ◽  
Author(s):  
Dean A. Rowe-Magnus ◽  
Mario Mencía ◽  
Fernando Rojo ◽  
Margarita Salas ◽  
George B. Spiegelman
Keyword(s):  
Bacillus Subtilis ◽  
Rna Polymerase ◽  
Transcriptional Activation ◽  
Response Regulator ◽  
In Vitro Transcription ◽  
Alpha Subunit ◽  
Deletion Mutants ◽  
Wild Type ◽  
Initiation Mechanism

ABSTRACT In vitro transcription from the spoIIG promoter byBacillus subtilis RNA polymerase reconstituted with wild-type alpha subunits and with C-terminal deletion mutants of the alpha subunit was equally stimulated by the response regulator Spo0A. Some differences in the structure of open complexes formed by RNA polymerase containing alpha subunit mutants were noted, although the wild-type and mutant polymerases appeared to use the same initiation mechanism.

scholarly journals Definition of the Binding Architecture to a Target Promoter of HP1043, the Essential Master Regulator of Helicobacter pylori

2021 ◽  
Vol 22 (15) ◽  
pp. 7848
Author(s):  
Annamaria Zannoni ◽  
Simone Pelliciari ◽  
Francesco Musiani ◽  
Federica Chiappori ◽  
Davide Roncarati ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Response Regulator ◽  
Consensus Sequence ◽  
Binding Mode ◽  
In Vitro Transcription ◽  
Target Sequence ◽  
Computational Alanine Scanning ◽  
Definition Of ◽  
Target Promoter

HP1043 is an essential orphan response regulator of Helicobacter pylori orchestrating multiple crucial cellular processes. Classified as a member of the OmpR/PhoB family of two-component systems, HP1043 exhibits a highly degenerate receiver domain and evolved to function independently of phosphorylation. Here, we investigated the HP1043 binding mode to a target sequence in the hp1227 promoter (Php1227). Scanning mutagenesis of HP1043 DNA-binding domain and consensus sequence led to the identification of residues relevant for the interaction of the protein with a target DNA. These determinants were used as restraints to guide a data-driven protein-DNA docking. Results suggested that, differently from most other response regulators of the same family, HP1043 binds in a head-to-head conformation to the Php1227 target promoter. HP1043 interacts with DNA largely through charged residues and contacts with both major and minor grooves of the DNA are required for a stable binding. Computational alanine scanning on molecular dynamics trajectory was performed to corroborate our findings. Additionally, in vitro transcription assays confirmed that HP1043 positively stimulates the activity of RNA polymerase.

scholarly journals Transcription Regulation of ezrA and Its Effect on Cell Division of Bacillus subtilis

2004 ◽  
Vol 186 (17) ◽  
pp. 5926-5932 ◽  
Author(s):  
Kuei-Min Chung ◽  
Hsin-Hsien Hsu ◽  
Suresh Govindan ◽  
Ban-Yang Chang
Keyword(s):  
Bacillus Subtilis ◽  
Cell Division ◽  
Critical Concentration ◽  
In Vitro Transcription ◽  
Ring Formation ◽  
Negative Regulator ◽  
Cell Length ◽  
Intracellular Level ◽  
Z Ring

ABSTRACT The EzrA protein of Bacillus subtilis is a negative regulator for FtsZ (Z)-ring formation. It is able to modulate the frequency and position of Z-ring formation during cell division. The loss of this protein results in cells with multiple Z rings located at polar as well as medial sites; it also lowers the critical concentration of FtsZ required for ring formation (P. A. Levin, I. G. Kurster, and A. D. Grossman, Proc. Natl. Acad. Sci. USA 96:9642-9647, 1999). We have studied the regulation of ezrA expression during the growth of B. subtilis and its effects on the intracellular level of EzrA as well as the cell length of B. subtilis. With the aid of promoter probing, primer extension, in vitro transcription, and Western blotting analyses, two overlapping σA-type promoters, P1 and P2, located about 100 bp upstream of the initiation codon of ezrA, have been identified. P1, supposed to be an extended −10 promoter, was responsible for most of the ezrA expression during the growth of B. subtilis. Disruption of this promoter reduced the intracellular level of EzrA very significantly compared with disruption of P2. Moreover, deletion of both promoters completely abolished EzrA in B. subtilis. More importantly, the cell length and percentage of filamentous cells of B. subtilis were significantly increased by disruption of the promoter(s). Thus, EzrA is required for efficient cell division during the growth of B. subtilis, despite serving as a negative regulator for Z-ring formation.

Phosphorylation of Spo0A activates its stimulation of in vitro transcription from the Bacillus subtilis spollG operon

1993 ◽  
Vol 9 (4) ◽  
pp. 741-749 ◽  
Author(s):  
T. H. Bird ◽  
J. K. Grimsiey ◽  
J. A. Hoch ◽  
G. B. Spiegelman
Keyword(s):  
Bacillus Subtilis ◽  
In Vitro Transcription ◽  
Stimulation Of

scholarly journals Bacillus subtilis YngB is a UTP-glucose-1-phosphate uridylyltransferase contributing to wall teichoic acid modification and glycolipid formation during anaerobic growth

2020 ◽  
Author(s):  
Chih-Hung Wu ◽  
Jeanine Rismondo ◽  
Rhodri M. L. Morgan ◽  
Yang Shen ◽  
Martin J. Loessner ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Teichoic Acid ◽  
Glucose Production ◽  
Growth Conditions ◽  
Anaerobic Growth ◽  
Acid Modification ◽  
Wall Teichoic Acid ◽  
Native Promoter

AbstractUTP-glucose-1-phosphate uridylyltransferases (UGPases) are enzymes that produce UDP-glucose from UTP and glucose-1-phosphate. In Bacillus subtilis 168, UDP-glucose is required for the decoration of wall teichoic acid (WTA) with glucose residues and the formation of glucolipids. The B. subtilis UGPase GtaB is essential for UDP-glucose production under standard aerobic growth conditions, and gtaB mutants display severe growth and morphological defects. However, bioinformatics predictions indicate that two other UGPases, are present in B. subtilis. Here, we investigated the function of one of them named YngB. The crystal structure of YngB revealed that the protein has the typical fold and all necessary active site features of a functional UGPase. Furthermore, UGPase activity could be demonstrated in vitro using UTP and glucose-1-phosphate as substrates. Expression of YngB from a synthetic promoter in a B. subtilis gtaB mutant resulted in the reintroduction of glucose residues on WTA and production of glycolipids, demonstrating that the enzyme can function as UGPase in vivo. When wild-type and mutant B. subtilis strains were grown under anaerobic conditions, YngB-dependent glycolipid production and glucose decorations on WTA could be detected, revealing that YngB is expressed from its native promoter under anaerobic condition. Based on these findings, along with the structure of the operon containing yngB and the transcription factor thought to be required for its expression, we propose that besides WTA, potentially other cell wall components might be decorated with glucose residues during oxygen limited growth condition.

scholarly journals Analysis of Bacillus subtilis tagAB andtagDEF Expression during Phosphate Starvation Identifies a Repressor Role for PhoP∼P

1998 ◽  
Vol 180 (3) ◽  
pp. 753-758 ◽  
Author(s):  
Wei Liu ◽  
Stephen Eder ◽  
F. Marion Hulett
Keyword(s):  
Cell Wall ◽  
Bacillus Subtilis ◽  
Teichoic Acid ◽  
Phosphate Starvation ◽  
Acid Synthesis ◽  
Pho Regulon ◽  
Wall Teichoic Acid ◽  
Negative Role ◽  
First Time ◽  
Starvation Conditions

ABSTRACT The tagAB and tagDEF operons, which are adjacent and divergently transcribed, encode genes responsible for cell wall teichoic acid synthesis in Bacillus subtilis. TheBacillus data presented here suggest that PhoP and PhoR are required for direct repression of transcription of the two operons under phosphate starvation conditions but have no regulatory role under phosphate-replete conditions. These data identify for the first time that PhoP∼P has a negative role in Pho regulon gene regulation.

scholarly journals Role of PhoP∼P in Transcriptional Regulation of Genes Involved in Cell Wall Anionic Polymer Biosynthesis in Bacillus subtilis

1998 ◽  
Vol 180 (15) ◽  
pp. 4007-4010 ◽  
Author(s):  
Ying Qi ◽  
F. Marion Hulett
Keyword(s):  
Cell Wall ◽  
Bacillus Subtilis ◽  
Teichoic Acid ◽  
Acid Synthesis ◽  
In Vitro Transcription ◽  
Anionic Polymer ◽  
Transcription System ◽  
Teichuronic Acid

ABSTRACT tagA, tagD, and tuaA operons are responsible for the synthesis of cell wall anionic polymer, teichoic acid, and teichuronic acid, respectively, in Bacillus subtilis. Under phosphate starvation conditions, teichuronic acid is synthesized while teichoic acid synthesis is inhibited. Expression of these genes is controlled by PhoP-PhoR, a two-component system. It has been proposed that PhoP∼P plays a key role in the activation oftuaA and the repression of tagA andtagD. In this study, we demonstrated the role of PhoP∼P in the switch process from teichoic acid synthesis to teichuronic acid synthesis, by using an in vitro transcription system. The results indicate that PhoP∼P is sufficient to repress the transcription of the tagA and tagD promoters and also to activate the transcription of the tuaA promoter.

scholarly journals The Global Regulator Spx Functions in the Control of Organosulfur Metabolism in Bacillus subtilis

2006 ◽  
Vol 188 (16) ◽  
pp. 5741-5751 ◽  
Author(s):  
Soon-Yong Choi ◽  
Dindo Reyes ◽  
Montira Leelakriangsak ◽  
Peter Zuber
Keyword(s):  
Bacillus Subtilis ◽  
In Vitro Transcription ◽  
Sulfur Source ◽  
Global Regulator ◽  
Control Of Gene Expression ◽  
Α Subunit ◽  
Cysteine Synthesis ◽  
Operon Expression ◽  
State Growth

ABSTRACT Spx is a global transcriptional regulator of the oxidative stress response in Bacillus subtilis. Its target is RNA polymerase, where it contacts the α subunit C-terminal domain. Recently, evidence was presented that Spx participates in sulfate-dependent control of organosulfur utilization operons, including the ytmI, yxeI, ssu, and yrrT operons. The yrrT operon includes the genes that function in cysteine synthesis from S-adenosylmethionine through intermediates S-adenosylhomocysteine, ribosylhomocysteine, homocysteine, and cystathionine. These operons are also negatively controlled by CymR, the repressor of cysteine biosynthesis operons. All of the operons are repressed in media containing cysteine or sulfate but are derepressed in medium containing the alternative sulfur source, methionine. Spx was found to negatively control the expression of these operons in sulfate medium, in part, by stimulating the expression of the cymR gene. In addition, microarray analysis, monitoring of yrrT-lacZ fusion expression, and in vitro transcription studies indicate that Spx directly activates yrrT operon expression during growth in medium containing methionine as sole sulfur source. These experiments have uncovered additional roles for Spx in the control of gene expression during unperturbed, steady-state growth.

scholarly journals Requirement of the Receiver and Phosphotransfer Domains of ArcB for Efficient Dephosphorylation of Phosphorylated ArcA In Vivo

2005 ◽  
Vol 187 (9) ◽  
pp. 3267-3272 ◽  
Author(s):  
Gabriela R. Peña-Sandoval ◽  
Ohsuk Kwon ◽  
Dimitris Georgellis
Keyword(s):  
Response Regulator ◽  
Growth Conditions ◽  
Sensor Kinase ◽  
Aerobic Growth ◽  
Two Component System ◽  
Two Component ◽  
Necessary And Sufficient ◽  
Respiratory Conditions

ABSTRACT The Arc two-component system, comprising the ArcB sensor kinase and the ArcA response regulator, modulates the expression of numerous genes in response to the respiratory conditions of growth. Under anoxic growth conditions, ArcB autophosphorylates and transphosphorylates ArcA, which in turn represses or activates its target operons. Under aerobic growth conditions, phosphorylated ArcA (ArcA-P) dephosphorylates and its transcriptional regulation is released. The dephosphorylation of ArcA-P has been shown to occur, at least in vitro, via an ArcAAsp54-P → ArcBHis717-P → ArcBAsp576-P → Pi reverse phosphorelay. In this study, the physiological significance of this pathway was assessed. The results demonstrate that the receiver and phosphotransfer domains of the tripartite sensor kinase ArcB are necessary and sufficient for efficient ArcA-P dephosphorylation in vivo.

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