bluuebunny/crossref_metadata_2025_split huggingface dataset:
Roles of the Intramolecular Disulfide Bridge in MotX and MotY, the Specific Proteins for Sodium-Driven Motors in
Vibrio
spp
Authors: Yagasaki Jin , Okabe Mayuko , Kurebayashi Rie , Yakushi Toshiharu , Homma Michio 
Published: June 2006

ABSTRACT

The proteins PomA, PomB, MotX, and MotY are essential for the motor function of Na
+
-driven flagella in
Vibrio
spp. Both MotY and MotX have the two cysteine residues (one of which is in a conserved tetrapeptide [CQLV]) that are inferred to form an intramolecular disulfide bond. The cysteine mutants of MotY prevented the formation of an intramolecular disulfide bond, which is presumably important for protein stability. Disruption of the disulfide bridge in MotX by site-directed mutagenesis resulted in increased instability, which did not, however, affect the motility of the cells. These lines of evidence suggest that the intramolecular disulfide bonds are involved in the stability of both proteins, but only MotY requires the intramolecular bridge for proper function.
Limited Role for the DsrA and RprA Regulatory RNAs in
rpoS
Regulation in
Salmonella enterica
Authors: Jones Amy M. , Goodwill Adam , Elliott Thomas 
Published: June 2006

ABSTRACT

RpoS, the sigma factor of enteric bacteria that responds to stress and stationary phase, is subject to complex regulation acting at multiple levels, including transcription, translation, and proteolysis. Increased translation of
rpoS
mRNA during growth at low temperature, after osmotic challenge, or with a constitutively activated Rcs phosphorelay depends on two
trans
-acting small regulatory RNAs (sRNAs) in
Escherichia coli
. The DsrA and RprA sRNAs are both highly conserved in
Salmonella enterica
, as is their target, an inhibitory antisense element within the
rpoS
untranslated leader. Analysis of
dsrA
and
rprA
deletion mutants indicates that while the increased translation of RpoS in response to osmotic challenge is conserved in
S. enterica
, dependence on these two sRNA regulators is much reduced. Furthermore, low-temperature growth or constitutive RcsC activation had only modest effects on RpoS expression, and these increases were, respectively, independent of
dsrA
or
rprA
function. This lack of conservation of sRNA function suggests surprising flexibility in RpoS regulation.
GlcNAc-6P Levels Modulate the Expression of Curli Fibers byEscherichia coli
Authors: Barnhart Michelle M. , Lynem Jaclyn , Chapman Matthew R. 
Published: June 2006

ABSTRACTCurli are extracellular surface fibers that are produced by many members of theEnterobacteriaceaeand contribute to biofilm formation. The environmental cues that promote biofilm formation are poorly understood. We found that deletion of theN-acetylglucosamine-6-phosphate (GlcNAc-6P) deacetylase gene,nagA, resulted in decreased transcription from the curli-specific promoterscsgBAandcsgDEFGand a corresponding decrease in curli production inEscherichia coli. nagAis in an operon that containsnagB,nagC,nagD, andnagE, whose products are required for utilization of GlcNAc as a carbon source. NagC is a repressor of thenagBACDandnagEgenes in the absence of intracellular GlcNAc-6P. We found thatnagCmutants were also defective in curli production. Growth of a wild-type strain on media containing additional GlcNAc reduced curli gene transcription to a level similar to the level observed whennagAwas deleted. The defect in curli production innagAornagCmutants was alleviated by deletion of the GlcNAc transporter gene,nagE. Curli-producing ΔnagAsuppressor mutants whose cells were unable to take up GlcNAc were isolated. These results suggest that elevated levels of intracellular GlcNAc-6P signal cells to down-regulate curli gene expression.
Osmotic Upshift Transiently Inhibits Uptake via ABC Transporters in Gram-Negative Bacteria
Authors: Fox M. A. , White J. P. , Hosie A. H. F. , Lodwig E. M. , Poole P. S. 
Published: June 2006

ABSTRACT

ATP-binding cassette transporters from several rhizobia and
Salmonella enterica
serovar Typhimurium, but not secondarily coupled systems, were inhibited by high concentrations (100 to 500 mM) of various osmolytes, an effect reversed by the removal of the osmolyte. ABC systems were also inactivated in isolated pea bacteroids, probably due to the obligatory use of high-osmolarity isolation media. Measurement of nutrient cycling in isolated pea bacteroids is impeded by this effect.
Autoregulation of the MisR/MisS Two-Component Signal Transduction System inNeisseria meningitidis
Authors: Tzeng Yih-Ling , Zhou Xiaoliu , Bao Shaojia , Zhao Shuming , Noble Corie , Stephens David S. 
Published: June 2006

ABSTRACTTwo-component regulatory systems are involved in processes important for bacterial pathogenesis. The proposedmisR/misS(orphoP/phoQ) system is one of four two-component systems of the obligate human pathogenNeisseria meningitidis. Inactivation of this system results in loss of phosphorylation of the lipooligosaccharide inner core and causes attenuation in a mouse model of meningococcal infection. MisR and the cytoplasmic domain of MisS were purified as His6and maltose binding protein fusion proteins, respectively. The MisS fusion was shown to be autophosphorylated in the presence of ATP, and the phosphoryl group was subsequently transferred to MisR. The phosphotransfer reaction was halted with a MisR/D52A mutation, while a MisS/H246A mutation prevented autophosphorylation. Specific interaction of phosphorylated MisR (MisR∼P) and MisR with themisRpromoter was demonstrated by gel mobility shift assays, where MisR∼P exhibited higher affinity than did the nonphosphorylated protein. The transcriptional start site of themisRSoperon was mapped, and DNase I protection assays revealed that MisR interacted with a 15-bp region upstream of the transcriptional start site that shared no similarity to binding motifs of other two-component systems. Transcriptional reporter studies suggested that MisR phosphorylation is critical for the autoinduction of themisRSoperon. Limited Mg2+concentration failed to induce expression of themisRSoperon, which is the only operon now proven to be under the direct control of the MisRS two-component system. Thus, these results indicate that the meningococcal MisRS system constitutes a functional signal transduction circuit and that both components are critical in the autoregulation of their expression.
Peptidoglycan Structure Analysis of
Lactococcus lactis
Reveals the Presence of an
l
,
d
-Carboxypeptidase Involved in Peptidoglycan Maturation
Authors: Courtin Pascal , Miranda Guy , Guillot Alain , Wessner Françoise , Mézange Christine , Domakova Elena , Kulakauskas Saulius , Chapot-Chartier Marie-Pierre 
Published: June 2006

ABSTRACT

Detailed structural analysis of
Lactococcus lactis
peptidoglycan was achieved by identification of its constituent muropeptides separated by reverse phase high-performance liquid chromatography. Modification of the classical elution buffer allowed direct and sensitive analysis of the purified muropeptides by matrix-assisted laser desorption ionization-time of flight mass spectrometry. The structures of 45 muropeptides were assigned for
L. lactis
strain MG1363. Analysis of the muropeptide composition of an MG1363
dacB
mutant showed that the
dacB
-encoded protein has
l
,
d
-carboxypeptidase activity and is involved in peptidoglycan maturation.
σ
28
-Dependent Transcription in
Salmonella enterica
Is Independent of Flagellar Shearing
Authors: Rosu Valentina , Hughes Kelly T. 
Published: June 2006

ABSTRACT

The FlgM anti-σ
28
factor is secreted in response to flagellar hook-basal body completion to allow σ
28
-dependent transcription of genes needed late in flagellar assembly, such as the flagellin structural gene,
fliC
. A long-standing hypothesis was that one role of FlgM secretion was to allow rapid expression of flagellin in response to shearing. We tested this hypothesis by following FlgM secretion and
fliC
transcription in response to flagellar shearing. Experiments showed that the level of FlgM inside the cell was unchanged after shearing whereas the extracellular FlgM levels increased in the growth medium as time passed. Identical results were obtained with cells that were not exposed to shear forces: internal FlgM levels remained constant while external FlgM levels rose with time at rates similar to those for the sheared culture. Consistent with this find, FlgM/σ
28
-dependent class 3 gene expression was unaffected by flagellar shearing but was affected by the growth phase of the cell. Regardless of exposure to shear forces, flagellar class 3 transcription rose sharply and then declined. These results demonstrate that flagellar regrowth following shearing is independent of FlgM secretion.
DevA, a GntR-Like Transcriptional Regulator Required for Development in
Streptomyces coelicolor
Authors: Hoskisson Paul A. , Rigali Sebastien , Fowler Kay , Findlay Kim C. , Buttner Mark J. 
Published: June 2006

ABSTRACT

The gram-positive filamentous bacterium
Streptomyces coelicolor
has a complex developmental cycle with three distinct phases: growth of the substrate mycelium, development of reproductive structures called aerial hyphae, and differentiation of these aerial filaments into long chains of exospores. During a transposon mutagenesis screen, we identified a novel gene (
devA
) required for proper development. The
devA
mutant produced only rare aerial hyphae, and those that were produced developed aberrant spore chains that were much shorter than wild-type chains and had misplaced septa.
devA
encodes a member of the GntR superfamily, a class of transcriptional regulators that typically respond to metabolite effector molecules.
devA
forms an operon with the downstream gene
devB
, which encodes a putative hydrolase that is also required for aerial mycelium formation on R5 medium. S1 nuclease protection analysis showed that transcription from the single
devA
promoter was temporally associated with vegetative growth, and enhanced green fluorescent protein transcriptional fusions showed that transcription was spatially confined to the substrate hyphae in the wild type. In contrast,
devAB
transcript levels were dramatically upregulated in a
devA
mutant and the
devA
promoter was also active in aerial hyphae and spores in this background, suggesting that DevA might negatively regulate its own production. This suggestion was confirmed by gel mobility shift assays that showed that DevA binds its own promoter region in vitro.
Regulation of the
Myxococcus xanthus
C-Signal-Dependent Ω4400 Promoter by the Essential Developmental Protein FruA
Authors: Yoder-Himes Deborah R. , Kroos Lee 
Published: June 2006

ABSTRACT

The bacterium
Myxococcus xanthus
employs extracellular signals to coordinate aggregation and sporulation during multicellular development. Extracellular, contact-dependent signaling that involves the CsgA protein (called C-signaling) activates FruA, a putative response regulator that governs a branched signaling pathway inside cells. One branch regulates cell movement, leading to aggregation. The other branch regulates gene expression, leading to sporulation. C-signaling is required for full expression of most genes induced after 6 h into development, including the gene identified by Tn
5 lac
insertion Ω4400. To determine if FruA is a direct regulator of Ω4400 transcription, a combination of in vivo and in vitro experiments was performed. Ω4400 expression was abolished in a
fruA
mutant. The DNA-binding domain of FruA bound specifically to DNA upstream of the promoter −35 region in vitro. Mutations between bp −86 and −77 greatly reduced binding. One of these mutations had been shown previously to reduce Ω4400 expression in vivo and make it independent of C-signaling. For the first time, chromatin immunoprecipitation (ChIP) experiments were performed on
M. xanthus
. The ChIP experiments demonstrated that FruA is associated with the Ω4400 promoter region late in development, even in the absence of C-signaling. Based on these results, we propose that FruA directly activates Ω4400 transcription to a moderate level prior to C-signaling and, in response to C-signaling, binds near bp −80 and activates transcription to a higher level. Also, the highly localized effects of mutations between bp −86 and −77 on DNA binding in vitro, together with recently published footprints, allow us to predict a consensus binding site of GTCG/CGA/G for the FruA DNA-binding domain.
RNase E Maintenance of Proper FtsZ/FtsA Ratio Required for Nonfilamentous Growth of
Escherichia coli
Cells but Not for Colony-Forming Ability
Authors: Tamura Masaru , Lee Kangseok , Miller Christine A. , Moore Christopher J. , Shirako Yukio , Kobayashi Masahiko , Cohen Stanley N. 
Published: June 2006

ABSTRACT

Inactivation or deletion of the RNase E-encoding
rne
gene of
Escherichia coli
results in the growth of bacterial cells as filamentous chains in liquid culture (K. Goldblum and D. Apirion, J. Bacteriol.
146:
128-132, 1981) and the loss of colony-forming ability (CFA) on solid media. RNase E dysfunction is also associated with abnormal processing of
ftsQAZ
transcripts (K. Cam, G. Rome, H. M. Krisch, and J.-P. Bouché, Nucleic Acids Res.
24:
3065-3070, 1996), which encode proteins having a central role in septum formation during cell division. We show here that RNase E regulates the relative abundances of FtsZ and FtsA proteins and that RNase E depletion results in decreased FtsZ, increased FtsA, and consequently an altered FtsZ/FtsA ratio. However, while restoration of the level of FtsZ to normal in
rne
null mutant bacteria reverses the filamentation phenotype, it does not restore CFA. Conversely, overexpression of a related RNase, RNase G, in
rne
-deleted bacteria restores CFA, as previously reported, without affecting FtsZ abundance. Our results demonstrate that RNase E activity is required to maintain a proper cellular ratio of the FtsZ and FtsA proteins in
E. coli
but that FtsZ deficiency does not account for the nonviability of cells lacking RNase E.
Mutations of the Membrane-Bound Disulfide Reductase DsbD That Block Electron Transfer Steps from Cytoplasm to Periplasm in
Escherichia coli
Authors: Cho Seung-Hyun , Beckwith Jon 
Published: June 2006

ABSTRACT

The cytoplasmic membrane protein DsbD keeps the periplasmic disulfide isomerase DsbC reduced, using the cytoplasmic reducing power of thioredoxin. DsbD contains three domains, each containing two reactive cysteines. One membrane-embedded domain, DsbDβ, transfers electrons from thioredoxin to the carboxy-terminal thioredoxin-like periplasmic domain DsbDγ. To evaluate the role of conserved amino acid residues in DsbDβ in the electron transfer process, we substituted alanines for each of 19 conserved amino acid residues and assessed the in vivo redox states of DsbC and DsbD. The mutant DsbDs of 11 mutants which caused defects in DsbC reduction showed relatively oxidized redox states. To analyze the redox state of each DsbD domain, we constructed a thrombin-cleavable DsbD (DsbD
TH
) from which we could generate all three domains as separate polypeptide chains by thrombin treatment in vitro. We divided the mutants with strong defects into two classes. The first mutant class consists of mutant DsbDβ proteins that cannot receive electrons from cytoplasmic thioredoxin, resulting in a DsbD that has all six of its cysteines disulfide bonded. The second mutant class represents proteins in which the transfer of electrons from DsbDβ to DsbDγ appears to be blocked. This class includes the mutant with the most clear-cut defect, P284A. We relate the properties of the mutants to the positions of the amino acids in the structure of DsbD and discuss mechanisms that would interfere with the electron transfer process.
Interplay of the Wzx Translocase and the Corresponding Polymerase and Chain LengthRegulator Proteins in the Translocation and Periplasmic Assembly of Lipopolysaccharide O Antigen
Authors: Marolda Cristina L. , Tatar Laura D. , Alaimo Cristina , Aebi Markus , Valvano Miguel A. 
Published: June 2006

ABSTRACT

Genetic evidence suggests that a family of bacterial and eukaryotic integral
membrane proteins (referred to as Wzx and Rft1, respectively) mediates
the transbilayer movement of isoprenoid lipid-linked glycans. Recent
work in our laboratory has shown that Wzx proteins involved in
O-antigen lipopolysaccharide (LPS) assembly have relaxed specificity
for the carbohydrate structure of the O-antigen subunit. Furthermore,
the proximal sugar bound to the isoprenoid lipid carrier,
undecaprenyl-phosphate (Und-P), is the minimal structure required for
translocation. In
Escherichia coli
K-12,
N
-acetylglucosamine (GlcNAc) is the proximal sugar of
the O16 and enterobacterial common antigen (ECA) subunits. Both O16 and
ECA systems have their respective translocases, Wzx
O16
and
Wzx
E
, and also corresponding polymerases (Wzy
O16
and Wzy
E
) and O-antigen chain-length regulators
(Wzz
O16
and Wzz
E
), respectively. In
this study, we show that the
E. coli wzx
E
gene can
fully complement a
wzx
O16
translocase deletion
mutant only if the majority of the ECA gene cluster is deleted. In
addition, we demonstrate that introduction of plasmids expressing
either the Wzy
E
polymerase or the Wzz
E
chain-length regulator proteins drastically reduces the O16
LPS-complementing activity of Wzx
E
. We also show that this
property is not unique to Wzx
E
, since Wzx
O16
and
Wzx
O7
can cross-complement translocase defects in the O16
and O7 antigen clusters only in the absence of their corresponding Wzz
and Wzy proteins. These genetic data are consistent with the notion
that the translocation of O-antigen and ECA subunits across the plasma
membrane and the subsequent assembly of periplasmic O-antigen and ECA
Und-PP-linked polymers depend on interactions among Wzx, Wzz, and Wzy,
which presumably form a multiprotein
complex.
Global Transcriptome Analysis ofTropheryma whippleiin Response to Temperature Stresses
Authors: Crapoulet Nicolas , Barbry Pascal , Raoult Didier , Renesto Patricia 
Published: June 2006

ABSTRACTTropheryma whipplei, the agent responsible for Whipple disease, is a poorly known pathogen suspected to have an environmental origin. The availability of the sequence of the 0.92-Mb genome of this organism made a global gene expression analysis in response to thermal stresses feasible, which resulted in unique transcription profiles. A few genes were differentially transcribed after 15 min of exposure at 43°C. The effects observed included up-regulation of thednaKregulon, which is composed of six genes and is likely to be under control of two HspR-associated inverted repeats (HAIR motifs) found in the 5′ region. Putative virulence factors, like the RibC and IspDF proteins, were also overexpressed. While it was not affected much by heat shock, theT. whippleitranscriptome was strongly modified following cold shock at 4°C. For the 149 genes that were differentially transcribed, eight regulons were identified, and one of them was composed of five genes exhibiting similarity with genes encoding ABC transporters. Up-regulation of these genes suggested that there was an increase in nutrient uptake when the bacterium was exposed to cold stress. As observed for other bacterial species, the major classes of differentially transcribed genes encode membrane proteins and enzymes involved in fatty acid biosynthesis, indicating that membrane modifications are critical. Paradoxically, the heat shock proteins GroEL2 and ClpP1 were up-regulated. Altogether, the data show that despite the lack of classical regulation pathways,T. whippleiexhibits an adaptive response to thermal stresses which is consistent with its specific environmental origin and could allow survival under cold conditions.
Role of DNA Replication and Repair in Thymineless Death in
Escherichia coli
Authors: Morganroth Pamela A. , Hanawalt Philip C. 
Published: June 2006

ABSTRACT

Inhibition of DNA replication with hydroxyurea during thymine starvation of
Escherichia coli
shows that active DNA synthesis is not required for thymineless death (TLD). Hydroxyurea experiments and thymine starvation of
lexA3
and
uvrA
DNA repair mutants rule out unbalanced growth, the SOS response, and nucleotide excision repair as explanations for TLD.
H-NS RepressesinvTranscription inYersinia enterocoliticathrough Competition with RovA and Interaction with YmoA
Authors: Ellison Damon W. , Miller Virginia L. 
Published: June 2006

ABSTRACTYersinia enterocoliticais able to efficiently invade Peyer's patches with the aid of invasin, an outer member protein involved in the attachment and invasion of M cells. Invasin is encoded byinv, which is positively regulated by RovA in bothY. enterocoliticaandYersinia pseudotuberculosiswhile negatively regulated by YmoA inY. enterocoliticaand H-NS inY. pseudotuberculosis. In this study we present data indicating H-NS and RovA bind directly and specifically to theinvpromoter ofY. enterocolitica. We also show that RovA and H-NS fromY. enterocoliticabind to a similar region of theinvpromoter and suggest they compete for binding sites. This is similar to recently published data fromY. pseudotuberculosis, revealing a potentially conserved mechanism ofinvregulation betweenY. enterocoliticaandY. pseudotuberculosis. Furthermore, we present data suggesting H-NS and YmoA form a repression complex on theinvpromoter, with H-NS providing the binding specificity and YmoA interacting with H-NS to form a repression complex. We also demonstrate that deletion of the predicted H-NS binding region relieves the requirement for RovA-dependent transcription of theinvpromoter, consistent with RovA acting as a derepressor of H-NS-mediated repression. Levels of H-NS and YmoA are similar between 26°C and 37°C, suggesting that the H-NS/YmoA repression complex is present at both temperatures, while the levels ofrovAtranscript are low at 37°C and high at 26°C, leading to expression ofinvat 26°C. Expression of RovA at 37°C results in transcription ofinvand production of invasin. Data presented here support a model ofinvregulation where the level of RovA within the cell governsinvexpression. As RovA levels increase, RovA can successfully compete for binding to theinvpromoter with the H-NS/YmoA complex, resulting in derepression ofinvtranscription.