bluuebunny/crossref_metadata_2025_split huggingface dataset:
CcpA Causes Repression of the
phoPR
Promoter through a Novel Transcription Start Site, P
A6
Authors: Puri-Taneja Ankita , Paul Salbi , Chen Yinghua , Hulett F. Marion 
Published: February 2006

ABSTRACT

The
Bacillus subtilis
PhoPR two-component system is directly responsible for activation or repression of Pho regulon genes in response to phosphate deprivation. The response regulator, PhoP, and the histidine kinase, PhoR, are encoded in a single operon with a complex promoter region that contains five known transcription start sites, which respond to at least two regulatory proteins. We report here the identification of another direct regulator of
phoPR
transcription, carbon catabolite protein A, CcpA. This regulator functions in the presence of glucose or other readily metabolized carbon sources. The maximum derepression of
phoPR
expression in a
ccpA
mutant compared to a wild-type stain was observed under excess phosphate conditions with glucose either throughout growth in a high-phosphate defined medium or in a low-phosphate defined medium during exponential growth, a growth condition when
phoPR
transcription is low in a wild-type strain due to the absence of autoinduction. Either HPr or Crh were sufficient to cause CcpA dependent repression of the
phoPR
promoter in vivo. A
ptsH1
(Hpr)
crh
double mutant completely relieves
phoPR
repression during phosphate starvation but not during phosphate replete growth. In vivo and in vitro studies showed that CcpA repressed
phoPR
transcription by binding directly to the
cre
consensus sequence present in the promoter. Primer extension and in vitro transcription studies revealed that the CcpA regulation of
phoPR
transcription was due to repression of P
A6
, a previously unidentified promoter positioned immediately upstream of the
cre
box. Eσ
A
was sufficient for transcription of P
A6
, which was repressed by CcpA in vitro. These studies showed direct repression by CcpA of a newly discovered Eσ
A
-responsive
phoPR
promoter that required either Hpr or Crh in vivo for direct binding to the putative consensus
cre
sequence located between P
A6
and the five downstream promoters characterized previously.
The Bacteriophage T4 Inhibitor and Coactivator AsiA InhibitsEscherichia coliRNA Polymerase More Rapidly in the Absence of σ70Region 1.1: Evidence that Region 1.1 Stabilizes the Interaction between σ70and Core
Authors: Hinton Deborah M. , Vuthoori Srilatha , Mulamba Rebecca 
Published: February 2006

ABSTRACTThe N-terminal region (region 1.1) of σ70, the primary σ subunit ofEscherichia coliRNA polymerase, is a negatively charged domain that affects the DNA binding properties of σ70regions 2 and 4. Region 1.1 prevents the interaction of free σ70with DNA and modulates the formation of stable (open) polymerase/promoter complexes at certain promoters. The bacteriophage T4 AsiA protein is an inhibitor of σ70-dependent transcription from promoters that require an interaction between σ70region 4 and the −35 DNA element and is the coactivator of transcription at T4 MotA-dependent promoters. Like AsiA, the T4 activator MotA also interacts with σ70region 4. We have investigated the effect of region 1.1 on AsiA inhibition and MotA/AsiA activation. We show that σ70region 1.1 is not required for MotA/AsiA activation at the T4 middle promoter PuvsX. However, the rate of AsiA inhibition and of MotA/AsiA activation of polymerase is significantly increased when region 1.1 is missing. We also find that RNA polymerase reconstituted with σ70that lacks region 1.1 is less stable than polymerase with full-length σ70. Our previous work has demonstrated that the AsiA-inhibited polymerase is formed when AsiA binds to region 4 of free σ70and then the AsiA/σ70complex binds to core. Our results suggest that in the absence of region 1.1, there is a shift in the dynamic equilibrium between polymerase holoenzyme and free σ70plus core, yielding more free σ70at any given time. Thus, the rate of AsiA inhibition and AsiA/MotA activation increases when RNA polymerase lacks region 1.1 because of the increased availability of free σ70. Previous work has argued both for and against a direct interaction between regions 1.1 and 4. Using anE. colitwo-hybrid assay, we do not detect an interaction between these regions. This result supports the idea that the ability of region 1.1 to prevent DNA binding by free σ70arises through an indirect effect.
Sll1717 Affects the Redox State of the Plastoquinone Pool by Modulating Quinol Oxidase Activity in Thylakoids
Authors: Kufryk Galyna I. , Vermaas Wim F. J. 
Published: February 2006

ABSTRACTASynechocystissp. strain PCC 6803 mutant lacking CtaI, a main subunit of cytochromecoxidase, is not capable of growing at light intensities below 5 μmol photons m−2s−1, presumably due to an overreduced plastoquinone pool in the thylakoid membrane. Upon selection for growth at light intensities below 5 μmol photons m−2s−1, a secondary mutant was generated that retained the CtaI deletion and had fully assembled photosystem II complexes; in this secondary mutant (pseudorevertant), oxygen evolution and respiratory activities were similar to those in the wild type. Functional complementation of the original CtaI-less strain to low-light tolerance by transformation with restriction fragments of genomic DNA of the pseudorevertant and subsequent mapping of the pseudoreversion site showed that the point mutation led to a Ser186Cys substitution in Sll1717, a protein of as-yet-unknown function and with a predicted ATP/GTP-binding domain. This mutation caused a decrease in the plastoquinone pool reduction level of thylakoids compared to that observed for the wild type. Based on a variety of experimental evidence, the most plausible mechanism to cause this effect is an activation of plastoquinol oxidation in thylakoids by the quinol oxidase CydAB that occurs without upregulation of the corresponding gene and that may be caused by an increased CydAB activity in thylakoids, conceivably due to altered CydAB sorting between cytoplasmic and thylakoid membranes. Sll1717 appears to be unique toSynechocystissp. strain PCC 6803 and has a close homologue encoded in the genome of this organism. The transcript level of sll1717 is low, which suggests that the corresponding protein is regulatory rather than structural.
The
Brucella abortus xthA-1
Gene Product Participates in Base Excision Repair and Resistance to Oxidative Killing but Is Not Required for Wild-Type Virulence in the Mouse Model
Authors: Hornback Michael L. , Roop R. Martin 
Published: February 2006

ABSTRACT

Exonuclease III, encoded by the
xthA
gene, plays a central role in the base excision pathway of DNA repair in bacteria. Studies with
Escherichia coli xthA
mutants have also shown that exonuclease III participates in the repair of oxidative damage to DNA. An isogenic
xthA-1
mutant (designated CAM220) derived from virulent
Brucella abortus
2308 exhibited increased sensitivity to the alkylating agent methyl methanesulfonate (MMS) compared to the parent strain. In contrast, 2308 and the isogenic
xthA-1
mutant displayed similar levels of resistance to the DNA cross-linker mitomycin C. These phenotypic properties are those that would be predicted for a strain defective in base excision repair. The
B. abortus xthA-1
mutant also displayed reduced resistance to killing by H
2
O
2
and the ONOO
−
-generating compound 3-morpholinosydnonimine (SIN-1) compared to strain 2308, indicating that the
xthA-1
gene product participates in protecting
B. abortus
2308 from oxidative damage. Introducing a plasmid-borne copy of the parental
xthA-1
gene into CAM220 restored wild-type resistance of this mutant to MMS, H
2
O
2
, and SIN-1. Although the
B. abortus xthA-1
mutant exhibited increased sensitivity to oxidative killing compared to the parental strain in laboratory assays, CAM220 and 2308 displayed equivalent spleen colonization profiles in BALB/c mice through 8 weeks postinfection and equivalent intracellular survival and replication profiles in cultured murine macrophages. Thus, although the
xthA-1
gene product participates in base excision repair and resistance to oxidative killing in
B. abortus
2308, XthA-1 is not required for wild-type virulence of this strain in the mouse model.
The Innate Immune Modulators Staphylococcal Complement Inhibitor and Chemotaxis Inhibitory Protein ofStaphylococcus aureusAre Located on β-Hemolysin-Converting Bacteriophages
Authors: van Wamel Willem J. B. , Rooijakkers Suzan H. M. , Ruyken Maartje , van Kessel Kok P. M. , van Strijp Jos A. G. 
Published: February 2006

ABSTRACTTwo newly discovered immune modulators, chemotaxis inhibitory protein ofStaphylococcus aureus(CHIPS) and staphylococcal complement inhibitor (SCIN), cluster on the conserved 3′ end of β-hemolysin (hlb)-converting bacteriophages (βC-φs). Since these βC-φs also carry the genes for the immune evasion molecules staphylokinase (sak) and enterotoxin A (sea), this 8-kb region at the 3′ end of βC-φ represents an innate immune evasion cluster (IEC). By PCR and Southern analyses of 85 clinicalStaphylococcus aureusstrains and 5 classical laboratory strains, we show that 90% ofS. aureusstrains carry a βC-φ with an IEC. Seven IEC variants were discovered, carrying different combinations ofchp,sak, orsea(orsep), always in the same 5′-to-3′ orientation and on the 3′ end of a βC-φ. From most IEC variants we could isolate active bacteriophages by mitomycin C treatment, of which lysogens were generated inS. aureusR5 (broad phage host). All IEC-carrying bacteriophages integrated intohlb, as was measured by Southern blotting of R5 lysogens. Large quantities of the different bacteriophages were obtained by mitomycin C treatment of the lysogens, and bacteriophages were collected and used to reinfect all lysogenic R5 strains. In total, five lytic families were found. Furthermore, phage DNA was isolated and digested with EcoR1, revealing that one IEC variant can be found on different βI-φs. In conclusion, the four human-specific innate immune modulators SCIN, CHIPS, SAK, and SEA form an IEC that is easily transferred amongS. aureusstrains by a diverse group of β-hemolysin-converting bacteriophages.
The Transcriptional Antiterminator RfaH Represses Biofilm Formation inEscherichia coli
Authors: Beloin Christophe , Michaelis Kai , Lindner Karin , Landini Paolo , Hacker Jörg , Ghigo Jean-Marc , Dobrindt Ulrich 
Published: February 2006

ABSTRACTWe investigated the influence of regulatory and pathogenicity island-associated factors (Hha, RpoS, LuxS, EvgA, RfaH, and tRNA5Leu) on biofilm formation by uropathogenicEscherichia coli(UPEC) strain 536. Only inactivation ofrfaH, which encodes a transcriptional antiterminator, resulted in increased initial adhesion and biofilm formation byE. coli536.rfaHinactivation in nonpathogenicE. coliK-12 isolate MG1655 resulted in the same phenotype. Transcriptome analysis of wild-type strain 536 and anrfaHmutant of this strain revealed that deletion ofrfaHcorrelated with increased expression offluorthologs.fluencodes antigen 43 (Ag43), which mediates autoaggregation and biofilm formation. We confirmed that deletion ofrfaHleads to increased levels offluandflu-like transcripts inE. coliK-12 and UPEC. Supporting the hypothesis that RfaH represses biofilm formation through reduction of the Ag43 level, the increased-biofilm phenotype ofE. coliMG1655rfaHwas reversed upon inactivation offlu. Deletion of the twofluorthologs, however, did not modify the behavior of mutant 536rfaH. Our results demonstrate that the strong initial adhesion and biofilm formation capacities of strain MG1655rfaHare mediated by both increased steady-state production of Ag43 and likely increased Ag43 presentation due to nullrfaH-dependent lipopolysaccharide depletion. Although the roles ofrfaHin the biofilm phenotype are different in UPEC strain 536 and K-12 strain MG1655, this study shows that RfaH, in addition to affecting the expression of bacterial virulence factors, also negatively controls expression and surface presentation of Ag43 and possibly another Ag43-independent factor(s) that mediates cell-cell interactions and biofilm formation.
Characterization of the Twin-Arginine Translocase Secretion System of
Mycobacterium smegmatis
Authors: Posey James E. , Shinnick Thomas M. , Quinn Frederick D. 
Published: February 2006

ABSTRACT

The twin-arginine translocation (TAT) system secretes fully folded proteins that contain a twin-arginine motif within their signal sequence across the cytoplasmic membrane in bacteria. Using a green fluorescent protein fused with a TAT signal sequence, we demonstrated that
Mycobacterium smegmatis
contains a TAT system. By inactivating individual genes, we showed that three genes (
tatA
,
tatB
, and
tatC
) are required for a functional TAT system in
M. smegmatis
. The
tat
mutants exhibited a decreased growth rate and altered colony morphology compared to the parent strain. Comparison of the secreted proteins of the Δ
tatC
and parent strain by two-dimensional polyacrylamide gel electrophoresis revealed an alteration in the secretion of at least five proteins, and one of the major TAT-dependent secreted proteins was identified as β-lactamase (BlaS). The genome of
M. smegmatis
was analyzed with the TATFIND program, and 49 putative TAT substrates were identified, including the succinate transporter DctP. Because disruption of the TAT secretion system has a direct effect on the physiology of
M. smegmatis
and homologs of the TAT proteins are also present in the genome of
Mycobacterium tuberculosis
, the TAT secretion system or its substrates may be good candidates for drug or vaccine development.
Pyruvate:Quinone Oxidoreductase in
Corynebacterium glutamicum
: Molecular Analysis of the
pqo
Gene, Significance of the Enzyme, and Phylogenetic Aspects
Authors: Schreiner Mark E. , Riedel Christian , Holátko Jiři , Pátek Miroslav , Eikmanns Bernhard J. 
Published: February 2006

ABSTRACT

Corynebacterium glutamicum
recently has been shown to possess pyruvate:quinone oxidoreductase (PQO), catalyzing the oxidative decarboxylation of pyruvate to acetate and CO
2
with a quinone as the electron acceptor. Here, we analyze the expression of the
C. glutamicum pqo
gene, investigate the relevance of the PQO enzyme for growth and amino acid production, and perform phylogenetic studies. Expression analyses revealed that transcription of
pqo
is initiated 45 bp upstream of the translational start site and that it is organized in an operon together with genes encoding a putative metal-activated pyridoxal enzyme and a putative activator protein. Inactivation of the chromosomal
pqo
gene led to the absence of PQO activity; however, growth and amino acid production were not affected under either condition tested. Introduction of plasmid-bound
pqo
into a pyruvate dehydrogenase complex-negative
C. glutamicum
strain partially relieved the growth phenotype of this mutant, indicating that high PQO activity can compensate for the function of the pyruvate dehydrogenase complex. To investigate the distribution of PQO enzymes in prokaryotes and to clarify the relationship between PQO, pyruvate oxidase (POX), and acetohydroxy acid synthase enzymes, we compiled and analyzed the phylogeny of respective proteins deposited in public databases. The analyses revealed a wide distribution of PQOs among prokaryotes, corroborated the hypothesis of a common ancestry of the three enzymes, and led us to propose that the POX enzymes of
Lactobacillales
were derived from a PQO.
lbtAandlbtBAre Required for Production of theLegionella pneumophilaSiderophore Legiobactin
Authors: Allard Kimberly A. , Viswanathan V. K. , Cianciotto Nicholas P. 
Published: February 2006

ABSTRACTUnder iron stress,Legionella pneumophilasecretes legiobactin, a nonclassical siderophore that is reactive in the chrome azurol S (CAS) assay. Here, we have optimized conditions for legiobactin expression, shown its biological activity, and identified two genes,lbtAandlbtB, which are involved in legiobactin production.lbtAappears to be iron repressed and encodes a protein that has significant homology with siderophore synthetases, and FrgA, a previously described iron-regulated protein ofL. pneumophila. lbtBencodes a protein homologous with members of the major facilitator superfamily of multidrug efflux pumps. Mutants lackinglbtAorlbtBwere defective for legiobactin, producing 40 to 70% less CAS reactivity in deferrated chemically defined medium (CDM). In bioassays, mutant CDM culture supernatants, unlike those of the wild type, did not support growth of iron-limited wild-type bacteria in 2′,2′-dipyridyl-containing buffered charcoal yeast extract (BCYE) agar and a ferrous iron transport mutant on BCYE agar without added iron. ThelbtAmutant was modestly defective for growth in deferrated CDM containing the iron chelator citrate, indicating that legiobactin is required in conditions of severe iron limitation. Complementation of thelbtmutants restored both siderophore expression, as measured by the CAS assay and bioassays, and bacterial growth in deferrated, citrate-containing media. ThelbtAmutant replicated as the wild type did in macrophages, amoebae, and the lungs of mice. However,L. pneumophilaexpresseslbtAin the macrophage, suggesting that legiobactin, though not required, may play a dispensable role in intracellular growth. The discovery oflbtABrepresents the first identification of genes required forL. pneumophilasiderophore expression.
Disruption of the Operon Encoding Ehb Hydrogenase Limits Anabolic CO
2
Assimilation in the Archaeon
Methanococcus maripaludis
Authors: Porat Iris , Kim Wonduck , Hendrickson Erik L. , Xia Qiangwei , Zhang Yi , Wang Tiansong , Taub Fred , Moore Brian C. , Anderson Iain J. , Hackett Murray , Leigh John A. , Whitman William B. 
Published: February 2006

ABSTRACT

Methanococcus maripaludis
is a mesophilic archaeon that reduces CO
2
to methane with H
2
or formate as an energy source. It contains two membrane-bound energy-conserving hydrogenases, Eha and Ehb. To determine the role of Ehb, a deletion in the
ehb
operon was constructed to yield the mutant, strain S40. Growth of S40 was severely impaired in minimal medium. Both acetate and yeast extract were necessary to restore growth to nearly wild-type levels, suggesting that Ehb was involved in multiple steps in carbon assimilation. However, no differences in the total hydrogenase specific activities were found between the wild type and mutant in either cell extracts or membrane-purified fractions. Methanogenesis by resting cells with pyruvate as the electron donor was also reduced by 30% in S40, suggesting a defect in pyruvate oxidation. CO dehydrogenase/acetyl coenzyme A (CoA) synthase and pyruvate oxidoreductase had higher specific activities in the mutant, and genes encoding these enzymes, as well as AMP-forming acetyl-CoA synthetase, were expressed at increased levels. These observations support a role for Ehb in anabolic CO
2
assimilation in methanococci.
Characterization of Late Acyltransferase Genes ofYersinia pestisand Their Role in Temperature-Dependent Lipid A Variation
Authors: Rebeil Roberto , Ernst Robert K. , Jarrett Clayton O. , Adams Kristin N. , Miller Samuel I. , Hinnebusch B. Joseph 
Published: February 2006

ABSTRACTYersinia pestisis an important human pathogen that is maintained in flea-rodent enzootic cycles in many parts of the world. During its life cycle,Y. pestissenses host-specific environmental cues such as temperature and regulates gene expression appropriately to adapt to the insect or mammalian host. For example,Y. pestissynthesizes different forms of lipid A when grown at temperatures corresponding to the in vivo environments of the mammalian host and the flea vector. At 37°C, tetra-acylated lipid A is the major form; but at 26°C or below, hexa-acylated lipid A predominates. In this study, we show that theY. pestis msbB(lpxM) andlpxPhomologs encode the acyltransferases that add C12and C16:1groups, respectively, to lipid IVAto generate the hexa-acylated form, and that their expression is upregulated at 21°C in vitro and in the flea midgut. AY. pestis ΔmsbB ΔlpxPdouble mutant that did not produce hexa-acylated lipid A was more sensitive to cecropin A, but not to polymyxin B. This mutant was able to infect and block fleas as well as the parental wild-type strain, indicating that the low-temperature-dependent change to hexa-acylated lipid A synthesis is not required for survival in the flea gut.
Inhibition of Cell Division Suppresses Heterocyst Development in
Anabaena
sp. Strain PCC 7120
Authors: Sakr Samer , Jeanjean Robert , Zhang Cheng-Cai , Arcondeguy Tania 
Published: February 2006

ABSTRACT

When the filamentous cyanobacterium
Anabaena
PCC 7120 is exposed to combined nitrogen starvation, 5 to 10% of the cells along each filament at semiregular intervals differentiate into heterocysts specialized in nitrogen fixation. Heterocysts are terminally differentiated cells in which the major cell division protein FtsZ is undetectable. In this report, we provide molecular evidence indicating that cell division is necessary for heterocyst development. FtsZ, which is translationally fused to the green fluorescent protein (GFP) as a reporter, is found to form a ring structure at the mid-cell position. SulA from
Escherichia coli
inhibits the GTPase activity of FtsZ in vitro and prevents the formation of FtsZ rings when expressed in
Anabaena
PCC 7120. The expression of
sulA
arrests cell division and suppresses heterocyst differentiation completely. The antibiotic aztreonam, which is targeted to the FtsI protein necessary for septum formation, has similar effects on both cell division and heterocyst differentiation, although in this case, the FtsZ ring is still formed. Therefore, heterocyst differentiation is coupled to cell division but independent of the formation of the FtsZ ring. Consistently, once the inhibitory pressure of cell division is removed, cell division should take place first before heterocyst differentiation resumes at a normal frequency. The arrest of cell division does not affect the accumulation of 2-oxoglutarate, which triggers heterocyst differentiation. Consistently, a nonmetabolizable analogue of 2-oxoglutarate does not rescue the failure of heterocyst differentiation when cell division is blocked. These results suggest that the control of heterocyst differentiation by cell division is independent of the 2-oxoglutarate signal.
Spo0A-Dependent Activation of an Extended −10 Region Promoter in
Bacillus subtilis
Authors: Chen Guangnan , Kumar Amrita , Wyman Travis H. , Moran Charles P. 
Published: February 2006

ABSTRACT

At the onset of endospore formation in
Bacillus subtilis
the DNA-binding protein Spo0A directly activates transcription from promoters of about 40 genes. One of these promoters, P
skf
, controls expression of an operon encoding a killing factor that acts on sibling cells. AbrB-mediated repression of P
skf
provides one level of security ensuring that this promoter is not activated prematurely. However, Spo0A also appears to activate the promoter directly, since Spo0A is required for P
skf
activity in a Δ
abrB
strain. Here we investigate the mechanism of P
skf
activation. DNase I footprinting was used to determine the locations at which Spo0A bound to the promoter, and mutations in these sites were found to significantly reduce promoter activity. The sequence near the −10 region of the promoter was found to be similar to those of extended −10 region promoters, which contain a TRTGn motif. Mutational analysis showed that this extended −10 region, as well as other base pairs in the −10 region, is required for Spo0A-dependent activation of the promoter. We found that a substitution of the consensus base pair for the nonconsensus base pair at position −9 of P
skf
produced a promoter that was active constitutively in both Δ
abrB
and Δ
spo0A
Δ
abrB
strains. Therefore, the base pair at position −9 of P
skf
makes its activity dependent on Spo0A binding, and the extended −10 region motif of the promoter contributes to its high level of activity.