bluuebunny/crossref_metadata_2025_split huggingface dataset:
ActP controls copper homeostasis in Rhizobium leguminosarum bv. viciae and Sinorhizobium meliloti preventing low pH‐induced copper toxicity
Authors: Reeve Wayne G. , Tiwari Ravi P. , Kale Neetin B. , Dilworth Michael J. , Glenn Andrew R. 
Published: March 2003

SummaryTwo ‘calcium‐irreparable’ acid‐sensitive mutants were identified after mutagenizing Rhizobium leguminosarum bv. viciae and Sinorhizobium
meliloti with Tn5. Each mutant contains a single copy of the transposon
which, inserted within the actP gene, prevents expression of a P‐type ATPase
that belongs to the CPx heavy metal‐transporting subfamily. Here, we show that both
actP‐knockout mutants show sensitivity to copper; omission of this heavy metal
from low pH‐buffered media restores acid tolerance to these strains. Furthermore,
complementation of the mutant phenotype requires only the actP gene. An actP–gusA
fusion in R. leguminosarum was transcriptionally regulated by copper in a
pH‐dependent manner. Downstream to actP in both organisms is the hmrR
gene that encodes a heavy metal‐responsive regulator
(HmrR) that belongs to the merR class of regulatory genes. Insertional inactivation
of hmrR abolished transcriptional activation of actP by copper ions
and increased the basal level of its expression in their absence. These observations
suggest that HmrR can regulate actP transcription positively and negatively. We show that copper homeostasis is an essential mechanism for the acid tolerance of these root nodule bacteria since it prevents this heavy metal from becoming overtly toxic in acidic conditions.
Spa15 of Shigella flexneri, a third type of chaperone in the type III secretion pathway
Authors: Page Anne‐Laure , Sansonetti Philippe , Parsot Claude 
Published: March 2003

SummaryThe type III secretion (TTS) pathway is used by numerous Gram‐negative pathogens to inject virulence factors into eukaryotic cells. In addition to a functional TTS apparatus, secretion of effector proteins depends upon specific chaperones. Using a two‐hybrid screen in yeast and a co‐purification assay in Shigella flexneri, we demonstrated that Spa15, which is encoded by an operon for components of the TTS apparatus, is associated in the cytoplasm with three proteins that are secreted by the TTS pathway, IpaA, IpgB1 and OspC3. Spa15 was found to be necessary for stability of IpgB1 but not IpaA, and for secretion of IpaA molecules that were stored in the cytoplasm but not those that were synthesized while the secretion apparatus was active. The ability of Spa15 to associate with several non‐homologous secreted proteins, the presence of Spa15 homologues in other TTS systems and the location of the corresponding genes within operons for components of the TTS apparatus suggest that Spa15 belongs to a new class of TTS chaperones.
Enhanced bacterial virulence through exploitation of host glycosaminoglycans
Authors: Menozzi Franco D. , Pethe Kevin , Bifani Pablo , Soncin Fabrice , Brennan Michael J. , Locht Camille 
Published: March 2003

SummaryPresent in the extracellular matrix and membranes of virtually all animal cells, proteoglycans (PGs) are among the first host macromolecules encountered by infectious agents. Because of their wide distribution and direct accessibility, it is not surprising that pathogenic bacteria have evolved mechanisms to exploit PGs for their own purposes, including mediating attachment to target cells. This is achieved through the expression of adhesins that recognize glycosa‐minoglycans (GAGs) linked to the core protein of PGs. Some pathogens, such as Bordetella pertussis and Chlamydia trachomatis,
may express more than one GAG‐binding adhesin. Bacterial interactions with PGs may
also facilitate cell invasion or systemic dissemination, as observed for Neisseria
gonorrhoeae and Mycobacterium tuberculosis respectively. Moreover, pathogenic bacteria can use PGs to enhance their virulence via a shedding of PGs that leads to the release of effectors that weaken the host defences. The exploitation of PGs by pathogenic bacteria is thus a multifaceted mechanistic process directly related to the potential virulence of a number of microorganisms.
Oxidative protein folding in bacteria
Authors: Collet Jean‐Francois , Bardwell James C. A. 
Published: March 2003

SummaryTen years ago it was thought that disulphide bond formation in prokaryotes occurred spontaneously. Now two pathways involved in disulphide bond formation have been well characterized, the oxidative pathway, which is responsible for the formation of disulphides, and the isomerization pathway, which shuffles incorrectly formed disulphides. Disulphide bonds are donated directly to unfolded polypeptides by the DsbA protein; DsbA is reoxidized by DsbB. DsbB generates disulphides de novo from oxidized quinones.
These quinones are reoxidized by the electron transport chain, showing that disulphide
bond formation is actually driven by electron transport. Disulphide isomerization
requires that incorrect disulphides be attacked using a reduced catalyst, followed
by the redonation of the disulphide, allowing alternative disulphide pairing. Two
isomerases exist in Escherichia coli, DsbC and DsbG. The membrane protein DsbD maintains these disulphide isomerases in their reduced and thereby active form. DsbD is kept reduced by cytosolic thioredoxin in an NADPH‐dependent reaction.
Two domains of a dual‐function NarK protein are required for nitrate uptake, the first step of denitrification in Paracoccus pantotrophus
Authors: Wood Nicholas J. , Alizadeh Tooba , Richardson David J. , Ferguson Stuart J. , Moir James W. B. 
Published: March 2003

SummaryUptake of nitrate into the cytoplasm is the first but least well understood step of denitrification; no gene has previously been identified to be necessary for this process. Upstream from the structural genes of the membrane‐bound nitrate reductase (narGHJI) in Paracoccus pantotrophus there is a fusion of two genes,

each homologous to members of the narK family. The single open reading frame

is predicted to encode 24 transmembrane helices, comprising two domains, NarK1 and

NarK2. Analysis of both the accumulation of intracellular nitrite and electron transport

through the nitrate reductase enzyme in narK mutants reveals that NarK1 and

NarK2 are both involved in nitrate uptake. Maximal rate of nitrate transport via

NarK2 was dependent upon nitrite, indicating that NarK2 encodes a nitrate/nitrite

antiporter. The uncouplers S13 and dinitrophenol showed that NarK2 was not dependent

on the proton motive force for maximal nitrate transport activity. Nitrate transport

via NarK1 was dependent on proton motive force, indicating that it is likely to be

a nitrate/proton symporter. Low expression of membrane‐bound nitrate reductase in

narK mutants was counteracted by azide, which induced nitrate reductase expression only if the transcriptional activator NarR was present.
Export of the siderophore enterobactin in Escherichia coli: involvement of a 43 kDa membrane exporter
Authors: Furrer Jason L. , Sanders Douglas N. , Hook‐Barnard India G. , McIntosh Mark A. 
Published: March 2003

SummaryThe enterobactin system for iron transport in Escherichia coli is well characterized
with the exception of the mechanism of enterobactin secretion to the extracellular
environment. Escherichia coli membrane protein P43, encoded by ybdA
in the chromosomal region of genes involved in enterobactin synthesis, shows strong
homology to the 12‐transmembrane segment major facilitator superfamily of export
pumps. A P43‐null mutation was created and siderophore nutrition assays, performed
with a panel of E. coli strains expressing one or more outer membrane receptors
for enterobactin‐related compounds, demonstrated that the P43 mutant was unable to
secrete enterobactin efficiently. Products released from the mutant strain were identified
with thin‐layer chromatography (TLC) and high‐performance liquid chromatography (HPLC),
revealing that the P43 mutant secretes little, if any, enterobactin, but elevated
levels of enterobactin breakdown products 2,3‐ dihydroxybenzoylserine (DHBS) monomer,
dimer, and trimer. These data establish that P43 is a critical component of the E.
coli enterobactin secretion machinery and provides a rationale for the designation
of the previous genetic locus ybdA as entS to reflect its relevant biological function.
A hypermutator phenotype attenuates the virulence of Listeria monocytogenes in a mouse model
Authors: Mérino Delphine , Réglier‐Poupet Hélène , Berche Patrick ,   The European Listeria Genome Consortium, Charbit Alain 
Published: March 2003

SummaryThe integrity of the genetic material of bacteria is guaranteed by a set of distinct repair mechanisms. The participation of these repair systems in bacterial pathogenicity has been addressed only recently. Here, we study for the first time the participation in virulence of the MutSL mismatch repair system of Listeria monocytogenes. The mutS and mutL genes, which are contiguous in the L. monocytogenes chromosome, were identified after in silico analysis. The deduced MutS shares 62% identity with MutS of Bacillus subtilis and 50% identity with HexA, its homologue in Streptococcus pneumoniae; MutL shares 59% identity with MutL of B. subtilis and 47% identity with HexB of S. pneumoniae. Functional analysis of the mutSL locus was studied by constructing a double knock‐out mutant. We showed that the deletion ΔmutSL induces: (i) a 100‐ to 1000‐fold increase in the spontaneous mutation rate; and (ii) a 10‐ to 15‐fold increase in the frequency of transduction, thus demonstrating the role of mutSL of L. monocytogenes in both mismatch repair and homologous recombination. We found that the deletion ΔmutSL moderately affected bacterial virulence, with a 1‐log increase in the lethal dose 50% (LD50) in the mouse. Strikingly, repeated passages of the mutant strain in mice reduced virulence further. Competition assays between wild‐type and mutant strains showed that the deletion ΔmutSL reduced the capacity of L. monocytogenes to survive and multiply in mice. These results thus demonstrate that, for the intracellular pathogen L. monocytogenes, a hypermutator phenotype is more deleterious than profitable to its virulence.
MntR modulates expression of the PerR regulon and superoxide resistance in Staphylococcus aureus through control of manganese uptake
Authors: Horsburgh Malcolm J. , Wharton Stephen J. , Cox Alan G. , Ingham Eileen , Peacock Sharon , Foster Simon J. 
Published: March 2003

SummaryThe Staphylococcus aureus DtxR‐like protein, MntR, controls expression of
the mntABC and mntH genes, which encode putative manganese transporters.
Mutation of mntABC produced a growth defect in metal‐depleted medium and increased
sensitivity to intracellularly generated superoxide radicals. These phenotypes resulted
from diminished uptake of manganese and were rescued by the addition of excess Mn(II).
Resistance to superoxide was incompletely rescued by Mn(II) for STE035 (mntA mntH),
and the strain had reduced virulence in a murine abscess model of infection. Expression
of mntABC was repressed by Mn(II) in an MntR‐dependent manner, which contrasts
with the expression of mntH that was not repressed in elevated Mn(II) and
was decreased in an mntR mutant. This demonstrates that MntR acts as a negative
and positive regulator of these loci re‐spectively. PerR, the peroxide resistance
regulon repressor, acts with MntR to control the expression of mntABC and
manganese uptake. The expression of the PerR‐regulated genes, katA (catalase),
ftn (ferritin) and fur (ferric uptake regulator), was diminished in
STE031 (mntR) when grown in excess Mn(II). Therefore, the control of Mn(II)‐regulated
members of the PerR regulon and the Fur protein is modulated by MntR through its
control of Mn(II) uptake. The co‐ordinated regulation of metal ion homeostasis and
oxidative stress resistance via the regulators MntR, PerR and Fur of S. aureus is discussed.
Replication fork reversal in DNA polymerase III mutants of Escherichia coli: a role for the β clamp
Authors: Grompone Gianfranco , Seigneur Marie , Ehrlich S. Dusko , Michel Bénédicte 
Published: March 2003

SummaryCertain replication mutations lead in Escherichia coli to a specific reaction
named replication fork reversal: at blocked forks, annealing of the nascent strands
and pairing of the template strands form a four‐way junction. RuvABC‐catalysed resolution
of this Holliday junction causes chromosome double‐strand breaks (DSBs) in a recBC
context and therefore creates a requirement for the recombination proteins RecBC
for viability. In the present work, two mutants were tested for replication fork
reversal: a dnaEts mutant and a dnaNts mutant, affected in the alpha
(polymerase) and beta (processivity clamp) subunits of DNA polymerase III holoenzyme
respectively. In the dnaEts recB strain, RuvABC‐dependent DSBs caused by the
dnaEts mutation occurred at 37°C or 42°C, indicating
the occurrence of replication fork reversal upon partial or complete inactivation
of the DNA polymerase alpha subunit. DSB formation was independent of RecA, RecQ
and the helicase function of PriA. In the dnaNts recB mutant, RuvABC‐dependent
DSB caused by the dnaNts mutation occurred only at semi‐permissive temperature,
37°C, indicating the occurrence of replication fork reversal in conditions
in which the remaining activity of the beta clamp is sufficient for viability. In
contrast, the dnaNts mutation did not cause chromosome breakage at 42°C,
a temperature at which DnaN is totally inactive and the dnaNts mutant is inviable.
We propose that a residual activity of the DNA polymerase III beta clamp is required
for replication fork reversal in the dnaNts mutant.
Mutual dependence of the expression of the cell differentiation regulatory protein HetR and the global nitrogen regulator NtcA during heterocyst development
Authors: Muro‐Pastor Alicia M. , Valladares Ana , Flores Enrique , Herrero Antonia 
Published: March 2003

SummaryHeterocyst differentiation in the cyanobacterium Anabaena sp. strain PCC
7120 depends on both the global nitrogen regulator NtcA and the cell differentiation
regulatory protein HetR, and induction of hetR upon nitrogen step‐down depends
on NtcA. The use of two out of the four transcription start points (tsps) described
for the hetR gene (those located at positions –728 and –271)
was found to be dependent on NtcA, and the use of the tsp located at position –271
was also dependent on HetR. Thus, autoregulation of hetR could take place
via the activation of transcription from this tsp. Expression of ntcA in nitrogen‐fixing
cultures was higher than in cells growing in the presence of ammonium or nitrate,
and high expression of ntcA under nitrogen deficiency resulted from an increased
use of tsps located at positions –180 and –49. The induction of the
use of these tsps did not take place in ntcA or hetR mutant strains.
These results indicate a mutual dependency in the induction of the regulatory genes
hetR and ntcA that takes place in response to nitrogen step‐down in
Anabaena cells. Expression of the hetC gene, which is also involved in the early steps of heterocyst differentiation, from its NtcA‐dependent tsp was, however, not dependent on HetR.
A single flavoprotein, AppA, integrates both redox and light signals in Rhodobacter sphaeroides
Authors: Braatsch Stephan , Gomelsky Mark , Kuphal Silke , Klug Gabriele 
Published: March 2003

SummaryAnoxygenic photosynthetic proteobacteria exhibit various light responses, including changing levels of expression of photosynthesis genes. However, the underlying mechanisms are largely unknown. We show that expression of the puf and puc operons encoding structural proteins of the photosynthetic complexes is strongly repressed by blue light under semi‐aerobic growth in Rhodobacter sphaeroides but not in the related species Rhodobacter capsulatus. At very low oxygen tension, puf and puc expression is independent of blue light in both species. Photosynthetic electron transport does not mediate the blue light repression, implying the existence of specific photoreceptors. Here, we show that the flavoprotein AppA is likely to act as the photoreceptor for blue light‐dependent repression during continuous illumination. The FAD cofactor of AppA is essential for the blue light‐dependent sensory transduction of this response. AppA, which is present in R. sphaeroides but not in R. capsulatus, is known to participate in the redox‐dependent control of photosynthesis gene expression. Thus, AppA is the first example of a protein with dual sensing capabilities that integrates both redox and light signals.
Isolation and characterization of capsule structure mutant strains of Cryptococcus neoformans
Authors: Moyrand Frédérique , Klaproth Birgit , Himmelreich Uwe , Dromer Françoise , Janbon Guilhem 
Published: March 2003

SummaryThe capsule of Cryptococcus neoformans is the most obvious virulence factor of this pathogenic yeast. The main capsule constituents are glucuronoxylomannans (GXM). Although several studies have focused on GXM composition and structure, very little is known about their genetics. To elucidate the relationship between the capsule structure and the pathophysiology of the cryptococcosis, genetic screening for mutant strains producing a structurally modified capsule was set up. Using monoclonal antibodies specific for different capsule sugar epitopes, we isolated strains with different mutated capsule structures (Cas mutants). According to their reactivities with various monoclonal antibodies, the mutants were classified into six groups (Cas1 to Cas6). One Cas2 mutant was used to clone the corresponding gene by complementation. This gene (USX1) encodes the previously identified UDP‐xylose synthase. We demonstrated that it is necessary for both capsule xylosylation and C. neoformans virulence.
Natrialba magadii virus φCh1: first complete nucleotide sequence and functional organization of a virus infecting a haloalkaliphilic archaeon
Authors: Klein R. , Baranyi U. , Rössler N. , Greineder B. , Scholz H. , Witte A. 
Published: March 2003

SummaryThe double‐stranded (ds)DNA virus φCh1 infects the haloalkaliphilic archaeon Natrialba magadii. The complete DNA sequence of 58 498 bp of the temperate virus was established, and the probable functions of 21 of 98 φCh1‐encoded open reading frames (ORFs) have been assigned. This knowledge has been used to propose functional modules each required for specific functions during virus development. The φCh1 DNA is terminally redundant and circularly permuted and therefore appears to be packaged by the so‐called headful mechanism. The presence of ORFs encoding homologues of proteins involved in plasmid replication as well as experimental evidence indicate a plasmid‐mediated replication strategy of the virus. Results from nanosequencing of virion components suggest covalent cross‐linking of monomers of at least one of the structural proteins during virus maturation. A comparison of the φCh1 genome with the partly sequenced genome of Halobacterium salinarum virus φH revealed a close relationship between the two viruses, although their host organisms live in distinct environments with respect to the different pH values required for growth.
SakA MAP kinase is involved in stress signal transduction, sexual development and spore viability in Aspergillus nidulans
Authors: Kawasaki Laura , Sánchez Olivia , Shiozaki Kazuhiro , Aguirre Jesús 
Published: March 2003

SummaryIn eukaryotic cells, environmental stress signals are transmitted by evolutionarily conserved MAPKs, such as Hog1 in the budding yeast Saccharomyces cerevisiae, Spc1 in the fission yeast Schizosaccharomyces pombe and p38/JNK in mammalian cells. Here, we report the identification of the Aspergillus nidulans sakA gene, which encodes a member of the stress MAPK family. The sakA gene is able to complement the S. pombe spc1− defects in both osmo‐regulation and cell cycle progression. Moreover, SakA MAPK is activated in response to osmotic and oxidative stress in both S. pombe and A. nidulans. However, in contrast to hog1 and spc1 mutants, the sakA null mutant is not sensitive to high osmolarity stress, indicating a different regulation of the osmostress response in this fungus. On the other hand, the ΔsakA mutant shows development and cell‐specific phenotypes. First, it displays premature steA‐dependent sexual development. Second, ΔsakA mutant produces asexual spores that are highly sensitive to oxidative and heat shock stress and lose viability upon storage. Indeed, SakA is transiently activated early after induction of conidiation. Our results indicate that SakA MAPK is involved in stress signal transduction and repression of sexual development, and is required for spore stress resistance and survival.
OhrR, a transcription repressor that senses and responds to changes in organic peroxide levels in Xanthomonas campestris pv. phaseoli
Authors: Panmanee Warunya , Vattanaviboon Paiboon , Eiamphungporn Warawan , Whangsuk Wirongrong , Sallabhan Ratiboot , Mongkolsuk Skorn 
Published: March 2003

SummaryWe report the physiological role of OhrR as an organic peroxide sensor and transcription repressor in Xanthomonas campestris pv. phaseoli. In vivo exposure of X. campestris pv. phaseoli to either tert‐butyl or cumene hydroperoxides efficiently neutralized OhrR repression of expression from the OhrR‐regulated P1 promoter. H2O2 was a weak and non‐physiological inducer of the system while other oxidants and metabolites of organic peroxide metabolism did not induce the expression from the P1. Northern blotting results indicated a correlation between concentrations of tert‐butyl hydroperoxide used in the treatment and the induction of ohr (an OhrR‐regulated gene) expression. In addition, the levels of ohr mRNA in cultures induced by various concentrations of tert‐butyl hydroperoxide were reduced in cells with high levels of an organic peroxide metabolising enzyme (AhpC‐AhpF) but not in cells with high catalase levels suggesting that organic peroxide interacts with OhrR. DNA band shift experiments using purified OhrR and the P1 promoter fragment showed that organic peroxide treatment prevented binding of the protein to the P1 promoter by oxidation of OhrR, as the inhibition of binding to the P1 promoter was reversed by addition of a reducing agent, DTT. The highly conserved cysteine residue C22 of OhrR is required for organic peroxide inducible gene expression. A mutant protein, OhrRC22S can repress the P1 promoter activity but is insensitive to organic peroxide treatment. Thus, OhrR is the first transcription repressor characterized that appeared to evolve to physiologically sense organic peroxides.