bluuebunny/crossref_metadata_2025_split huggingface dataset:
TheLactobacillus casei ptsHI47T Mutation Causes Overexpression of a LevR-Regulated but RpoN-Independent Operon Encoding a Mannose Class Phosphotransferase System
Authors: Mazé Alain , Boël Grégory , Poncet Sandrine , Mijakovic Ivan , Le Breton Yoann , Benachour Abdellah , Monedero Vicente , Deutscher Josef , Hartke Axel 
Published: July 2004

ABSTRACTA proteome analysis ofLactobacillus caseimutants that are affected in carbon catabolite repression revealed that a 15-kDa protein was strongly overproduced in aptsHI47T 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)-dependentBacillus subtilistranscription activator LevR. The following pentacistronic operon is oriented in the opposite direction and encodes four proteins with strong similarity to the proteins of theB. subtilisLev-PTS and one protein of unknown function. The genes present on the 7.1-kb DNA fragment were therefore calledlevRandlevABCDX. ThelevABCDXoperon was induced by fructose and mannose. No “−12, −24” promoter typical of RpoN-dependent genes precedes theL. casei levoperon, and its expression was therefore RpoN independent but required LevR. Phosphorylation of LevR by P∼His-HPr stimulates its activity, while phosphorylation by P∼EIIBLevinhibits it. Disruption of the EIIBLev-encodinglevBgene therefore led to strong constitutive expression of thelevoperon, which was weaker in a strain carrying aptsImutation preventing phosphorylation by both P∼EIIBLevand P∼His-HPr. Expression of theL. casei levoperon 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 EIIALevare assumed to be responsible for the elevated expression of thelevoperon in theptsHI47T mutant.
Transposition of the Heat-Stable ToxinastAGene into a Gifsy-2-Related Prophage ofSalmonella entericaSerovar Abortusovis
Authors: Bacciu Donatella , Falchi Giovanni , Spazziani Alessandra , Bossi Lionello , Marogna Gavino , Leori Guido Sisinnio , Rubino Salvatore , Uzzau Sergio 
Published: July 2004

ABSTRACTThe horizontal transfer and acquisition of virulence genes via mobile genetic elements have been a major driving force in the evolution ofSalmonellapathogenicity. Serovars ofSalmonella entericacarry variable assortments of phage-encoded virulence genes, suggesting that temperate phages play a pivotal role in this process. Epidemic isolates ofS. entericaserovar Typhimurium are consistently lysogenic for two lambdoid phages, Gifsy-1 and Gifsy-2, carrying known virulence genes. Other serovars ofS. enterica, including serovars Dublin, Gallinarum, Enteritidis, and Hadar, carry distinct prophages with similarity to the Gifsy phages. In this study, we analyzed Gifsy-related loci fromS. entericaserovar Abortusovis, a pathogen associated exclusively with ovine infection. A cryptic prophage, closely related to serovar Typhimurium phage Gifsy-2, was identified. This element, named Gifsy-2AO, was shown to contribute to serovar Abortusovis systemic infection in lambs. Sequence analysis of the prophage b region showed a large deletion which covers genes encoding phage tail fiber proteins and putative virulence factors, including type III secreted effector protein SseI (GtgB, SrfH). This deletion was identified in most of the serovar Abortusovis isolates tested and might be dependent on the replicative transposition of an adjacent insertion sequence, IS1414, previously identified in pathogenicEscherichia colistrains. IS1414encodes heat-stable toxin EAST1 (astA) and showed multiple genomic copies in isolates of serovar Abortusovis. To our knowledge, this is the first evidence of intergeneric transfer of virulence genes via insertion sequence elements inSalmonella. The acquisition of IS1414(EAST1) and its frequent transposition within the chromosome might improve the fitness of serovar Abortusovis within its narrow ecological niche.
First Characterization of an Archaeal GTP-Dependent Phosphoenolpyruvate Carboxykinase from the Hyperthermophilic Archaeon
Thermococcus kodakaraensis
KOD1
Authors: Fukuda Wakao , Fukui Toshiaki , Atomi Haruyuki , Imanaka Tadayuki 
Published: July 2004

ABSTRACT

Phosphoenolpyruvate carboxykinase (PCK), which catalyzes the nucleotide-dependent, reversible decarboxylation of oxaloacetate to yield phosphoenolpyruvate and CO
2
, is one of the important enzymes in the interconversion between C
3
and C
4
metabolites. This study focused on the first characterization of the enzymatic properties and expression profile of an archaeal PCK from the hyperthermophilic archaeon
Thermococcus kodakaraensis
(Pck

Tk

). Pck

Tk

showed 30 to 35% identities to GTP-dependent PCKs from mammals and bacteria but was located in a branch distinct from that of the classical enzymes in the phylogenetic tree, together with other archaeal homologs from
Pyrococcus
and
Sulfolobus
spp. Several catalytically important regions and residues, found in all known PCKs irrespective of their nucleotide specificities, were conserved in Pck

Tk

. However, the predicted GTP-binding region was unique compared to those in other GTP-dependent PCKs. The recombinant Pck

Tk

actually exhibited GTP-dependent activity and was suggested to possess dual cation-binding sites specific for Mn
2+
and Mg
2+
. The enzyme preferred phosphoenolpyruvate formation from oxaloacetate, since the

K
m

value for oxaloacetate was much lower than that for phosphoenolpyruvate. The transcription and activity levels in
T. kodakaraensis
were higher under gluconeogenic conditions than under glycolytic conditions. These results agreed with the role of Pck

Tk

in providing phosphoenolpyruvate from oxaloacetate as the first step of gluconeogenesis in this hyperthermophilic archaeon. Additionally, under gluconeogenic conditions, we observed higher expression levels of Pck

Tk

on pyruvate than on amino acids, implying that it plays an additional role in the recycling of excess phosphoenolpyruvate produced from pyruvate, replacing the function of the anaplerotic phosphoenolpyruvate carboxylase that is missing from this archaeon.
Delineation of Upstream Signaling Events in theSalmonellaPathogenicity Island 2 Transcriptional Activation Pathway
Authors: Kim Charles C. , Falkow Stanley 
Published: July 2004

ABSTRACTSurvival and replication in the intracellular environment are critical components of the ability ofSalmonella entericaserovar Typhimurium to establish systemic infection in the murine host. Intracellular survival is mediated by a number of genetic loci, includingSalmonellapathogenicity island 2 (SPI2). SPI2 is a 40-kb locus encoding a type III secretion system that secretes effector molecules, which permits bacterial survival and replication in the intracellular environment of host cells. A two-component regulatory system,ssrAB, is also encoded in SPI2 and controls expression of the secretion system and effectors. While the environmental signals to which SPI2 responds in vivo are not known, activation of expression is dependent on OmpR and can be stimulated in vitro by chelation of cations or by a shift from rich to acidic minimal medium. In this work, we demonstrated that SPI2 activation is associated with OmpR in the phosphorylated form (OmpR-P). Mutations inenvZandackA-pta, which disrupted two distinct sources of OmpR phosphorylation, indicated that SPI2 activation by chelators or a shift from rich to acidic minimal medium is largely dependent on functional EnvZ. In contrast, the PhoPQ pathway is not required for SPI2 activation in the presence of OmpR-P. As in the case of in vitro stimulation, SPI2 expression in macrophages correlates with the presence of OmpR-P. Additionally, EnvZ, but not acetyl phosphate, is required for maximal expression of SPI2 in the intracellular environment, suggesting that the in vitro SPI2 activation pathway is the same as that used in vivo.
Iron Acquisition and Regulation in
Campylobacter jejuni
Authors: Palyada Kiran , Threadgill Deborah , Stintzi Alain 
Published: July 2004

ABSTRACT

Iron affects the physiology of bacteria in two different ways: as a micronutrient for bacterial growth and as a catalyst for the formation of hydroxyl radicals. In this study, we used DNA microarrays to identify the
C. jejuni
genes that have their transcript abundance affected by iron availability. The transcript levels of 647 genes were affected after the addition of iron to iron-limited
C. jejuni
cells. Several classes of affected genes were revealed within 15 min, including immediate-early response genes as well as those specific to iron acquisition and metabolism. In contrast, only 208 genes were differentially expressed during steady-state experiments comparing iron-rich and iron-limited growth conditions. As expected, genes annotated as being involved in either iron acquisition or oxidative stress defense were downregulated during both time course and steady-state experiments, while genes encoding proteins involved in energy metabolism were upregulated. Because the level of protein glycosylation increased with iron limitation, iron may modulate the level of
C. jejuni
virulence by affecting the degree of protein glycosylation. Since iron homeostasis has been shown to be Fur regulated in
C. jejuni
, an isogenic
fur
mutant was used to define the Fur regulon by transcriptome profiling. A total of 53 genes were Fur regulated, including many genes not previously associated with Fur regulation. A putative Fur binding consensus sequence was identified in the promoter region of most iron-repressed and Fur-regulated genes. Interestingly, a
fur
mutant was found to be significantly affected in its ability to colonize the gastrointestinal tract of chicks, highlighting the importance of iron homeostasis in vivo. Directed mutagenesis of other genes identified by the microarray analyses allowed the characterization of the ferric enterobactin receptor, previously named CfrA. Chick colonization assays indicated that mutants defective in enterobactin-mediated iron acquisition were unable to colonize the gastrointestinal tract. In addition, a mutation in a receptor (Cj0178) for an uncharacterized iron source also resulted in reduced colonization potential. Overall, this work documents the complex response of
C. jejuni
to iron availability, describes the genetic network between the Fur and iron regulons, and provides insight regarding the role of iron in
C. jejuni
colonization in vivo.
Two C—P Lyase Operons in
Pseudomonas stutzeri
and Their Roles in the Oxidation of Phosphonates, Phosphite, and Hypophosphite
Authors: White Andrea K. , Metcalf William W. 
Published: July 2004

ABSTRACT

DNA sequencing and analysis of two distinct C—P lyase operons in
Pseudomonas stutzeri
WM88 were completed. The
htxABCDEFGHIJKLMN
operon encodes a hypophosphite-2-oxoglutarate dioxygenase (HtxA), whereas the predicted amino acid sequences of HtxB to HtxN are each homologous to the components of the
Escherichia coli phn
operon, which encodes C—P lyase, although homologs of
E. coli phnF
and
phnO
are absent. The genes in the
htx
operon are cotranscribed based on gene organization, and the presence of the intergenic sequences is verified by reverse transcription-PCR with total RNA. Deletion of the
htx
locus does not affect the ability of
P. stutzeri
to grow on phosphonates, indicating the presence of an additional C—P lyase pathway in this organism. To identify the genes comprising this pathway, a Δ
htx
strain was mutagenized and one mutant lacking the ability to grow on methylphosphonate as the sole P source was isolated. A ca.-10.6-kbp region surrounding the transposon insertion site of this mutant was sequenced, revealing 13 open reading frames, designated
phnCDEFGHIJKLMNP,
which were homologous to the
E. coli phn
genes. Deletion of both the
htx
and
phn
operons of
P. stutzeri
abolishes all growth on methylphosphonate and aminoethylphosphonate. Both operons individually support growth on methylphosphonate; however, the
phn
operon supports growth on aminoethylphosphonate and phosphite, as well. The substrate ranges of both C—P lyases are limited, as growth on other phosphonate compounds, including glyphosate and phenylphosphonate, was not observed.
Construction and Validation of the
Rhodobacter sphaeroides
2.4.1 DNA Microarray: Transcriptome Flexibility at Diverse Growth Modes
Authors: Pappas Christopher T. , Sram Jakub , Moskvin Oleg V. , Ivanov Pavel S. , Mackenzie R. Christopher , Choudhary Madhusudan , Land Miriam L. , Larimer Frank W. , Kaplan Samuel , Gomelsky Mark 
Published: July 2004

ABSTRACT

A high-density oligonucleotide DNA microarray, a genechip, representing the 4.6-Mb genome of the facultative phototrophic proteobacterium,
Rhodobacter sphaeroides
2.4.1, was custom-designed and manufactured by Affymetrix, Santa Clara, Calif. The genechip contains probe sets for 4,292 open reading frames (ORFs), 47 rRNA and tRNA genes, and 394 intergenic regions. The probe set sequences were derived from the genome annotation generated by Oak Ridge National Laboratory after extensive revision, which was based primarily upon codon usage characteristic of this GC-rich bacterium. As a result of the revision, numerous missing ORFs were uncovered, nonexistent ORFs were deleted, and misidentified start codons were corrected. To evaluate
R. sphaeroides
transcriptome flexibility, expression profiles for three diverse growth modes—aerobic respiration, anaerobic respiration in the dark, and anaerobic photosynthesis—were generated. Expression levels of one-fifth to one-third of the
R. sphaeroides
ORFs were significantly different in cells under any two growth modes. Pathways involved in energy generation and redox balance maintenance under three growth modes were reconstructed. Expression patterns of genes involved in these pathways mirrored known functional changes, suggesting that massive changes in gene expression are the major means used by
R. sphaeroides
in adaptation to diverse conditions. Differential expression was observed for genes encoding putative new participants in these pathways (additional photosystem genes, duplicate NADH dehydrogenase, ATP synthases), whose functionality has yet to be investigated. The DNA microarray data correlated well with data derived from quantitative reverse transcription-PCR, as well as with data from the literature, thus validating the
R. sphaeroides
genechip as a powerful and reliable tool for studying unprecedented metabolic versatility of this bacterium.
Evidence that the
algI
/
algJ
Gene Cassette, Required for O Acetylation of
Pseudomonas aeruginosa
Alginate, Evolved by Lateral Gene Transfer
Authors: Franklin Michael J. , Douthit Stephanie A. , McClure Marcella A. 
Published: July 2004

ABSTRACT

Pseudomonas aeruginosa
strains, isolated from chronically infected patients with cystic fibrosis, produce the O-acetylated extracellular polysaccharide, alginate, giving these strains a mucoid phenotype. O acetylation of alginate plays an important role in the ability of mucoid
P. aeruginosa
to form biofilms and to resist complement-mediated phagocytosis. The O-acetylation process is complex, requiring a protein with seven transmembrane domains (AlgI), a type II membrane protein (AlgJ), and a periplasmic protein (AlgF). The cellular localization of these proteins suggests a model wherein alginate is modified at the polymer level after the transport of O-acetyl groups to the periplasm. Here, we demonstrate that this mechanism for polysaccharide esterification may be common among bacteria, since AlgI homologs linked to type II membrane proteins are found in a variety of gram-positive and gram-negative bacteria. In some cases, genes for these homologs have been incorporated into polysaccharide biosynthetic operons other than for alginate biosynthesis. The phylogenies of AlgI do not correlate with the phylogeny of the host bacteria, based on 16S rRNA analysis. The
algI
homologs and the gene for their adjacent type II membrane protein present a mosaic pattern of gene arrangement, suggesting that individual components of the multigene cassette, as well as the entire cassette, evolved by lateral gene transfer. AlgJ and the other type II membrane proteins, although more diverged than AlgI, contain conserved motifs, including a motif surrounding a highly conserved histidine residue, which is required for alginate O-acetylation activity by AlgJ. The AlgI homologs also contain an ordered series of motifs that included conserved amino acid residues in the cytoplasmic domain CD-4; the transmembrane domains TM-C, TM-D, and TM-E; and the periplasmic domain PD-3. Site-directed mutagenesis studies were used to identify amino acids important for alginate O-acetylation activity, including those likely required for (i) the interaction of AlgI with the O-acetyl precursor in the cytoplasm, (ii) the export of the O-acetyl group across the cytoplasmic membrane, and (iii) the transfer of the O-acetyl group to a periplasmic protein or to alginate. These results indicate that AlgI belongs to a family of membrane proteins required for modification of polysaccharides and that a mechanism requiring an AlgI homolog and a type II membrane protein has evolved by lateral gene transfer for the esterification of many bacterial extracellular polysaccharides.
Characterization of NopP, a Type III Secreted Effector of
Rhizobium
sp. Strain NGR234
Authors: Ausmees Nora , Kobayashi Hajime , Deakin William J. , Marie Corinne , Krishnan Hari B. , Broughton William J. , Perret Xavier 
Published: July 2004

ABSTRACT

The type three secretion system (TTSS) encoded by pNGR234
a
, the symbiotic plasmid of
Rhizobium
sp. strain NGR234, is responsible for the flavonoid- and NodD1-dependent secretion of

n

odulation

o

uter

p

roteins (Nops). Abolition of secretion of all or specific Nops significantly alters the nodulation ability of NGR234 on many of its hosts. In the closely related strain
Rhizobium fredii
USDA257, inactivation of the TTSS modifies the host range of the mutant so that it includes the improved
Glycine max
variety McCall. To assess the impact of individual TTSS-secreted proteins on symbioses with legumes, various attempts were made to identify
nop
genes. Amino-terminal sequencing of peptides purified from gels was used to characterize NopA, NopL, and NopX, but it failed to identify SR3, a TTSS-dependent product of USDA257. By using phage display and antibodies that recognize SR3, the corresponding protein of NGR234 was identified as NopP. NopP, like NopL, is an effector secreted by the TTSS of NGR234, and depending on the legume host, it may have a deleterious or beneficial effect on nodulation or it may have little effect.
Interaction between Protein Subunits of the Type IV Secretion System ofBartonella henselae
Authors: Shamaei-Tousi Alireza , Cahill Rachel , Frankel Gad 
Published: July 2004

ABSTRACTIn this study we used the yeast two-hybrid system to identify interactions between protein subunits of thevirBtype IV secretion system ofBartonella henselae. We report interactions between inner membrane and periplasmic proteins, the pilus polypeptide, and the core complex and a novel interaction between VirB3 and VirB5.
Role of
Escherichia coli
DNA Polymerase IV in In Vivo Replication Fidelity
Authors: Kuban Wojciech , Jonczyk Piotr , Gawel Damian , Malanowska Karolina , Schaaper Roel M. , Fijalkowska Iwona J. 
Published: July 2004

ABSTRACT

We have investigated whether DNA polymerase IV (Pol IV; the
dinB
gene product) contributes to the error rate of chromosomal DNA replication in
Escherichia coli
. We compared mutation frequencies in mismatch repair-defective strains that were either
dinB
positive or
dinB
deficient, using a series of mutational markers, including
lac
targets in both orientations on the chromosome. Virtually no contribution of Pol IV to the chromosomal mutation rate was observed. On the other hand, a significant effect of
dinB
was observed for reversion of a
lac
allele when the
lac
gene resided on an F′(
pro-lac
) episome.
Loop Deletions Indicate Regions Important for FhuA Transport and Receptor Functions in
Escherichia coli
Authors: Endriß Franziska , Braun Volkmar 
Published: July 2004

ABSTRACT

Precise deletions of cell surface-exposed loops of FhuA resulted in mutants of
Escherichia coli
with distinct phenotypes. Deletion of loop 3 or 11 inactivated ferrichrome transport activity. Deletion of loop 8 inactivated receptor activity for colicin M and the phages T1, T5, and φ80. The loop 7 deletion mutant was colicin M resistant but fully phage sensitive. The loop 4 deletion mutant was resistant to the TonB-dependent phages T1 and φ80 but fully sensitive to the TonB-independent phage T5. The phenotypes of the deletion mutants revealed important sites for the multiple FhuA transport and receptor activities. The ligand binding sites are nonidentical and are distributed among the entire exposed surface. Presumably, FhuA evolved as a ferrichrome transporter and was subsequently used as a receptor by the phages and colicin M, which selected the same as well as distinct loops as receptor sites.
Manipulating dendritic cell biology for the active immunotherapy of cancer
Authors: O'Neill David W. , Adams Sylvia , Bhardwaj Nina 
Published: July 2004

Abstract
Dendritic cells (DCs) are specialized antigen-presenting cells (APCs) that have an unequaled capacity to initiate primary immune responses, including tolerogenic responses. Because of the importance of DCs in the induction and control of immunity, an understanding of their biology is central to the development of potent immunotherapies for cancer, chronic infections, autoimmune disease, and induction of transplantation tolerance. This review discusses recent advances in DC research and the application of this knowledge toward new strategies for the clinical manipulation of DCs for cancer immunotherapy.
Increase in laminin expression in allergic airway remodelling and decrease by dexamethasone
Authors: Christie P.E. , Jonas M. , Tsai C-H. , Chi E.Y. , Henderson W.R. 
Published: July 2004

Lung expression of the extracellular matrix protein, laminin, and its receptor, laminin‐1 receptor, were examined in a mouse model of asthma with airway remodelling.Ovalbumin (OVA) was administered to BALB/c mice, intraperitoneally on days 0 and 14, and intranasally periodically between days 14 and 75.The mice developed airway eosinophil and mononuclear inflammatory cell infiltration and fibrosis. On day 76, a marked increase in total laminin was seen in the airways of OVA‐treated mice compared to controls by Western blot analysis. The increased laminin expression was detected immunocytochemically in the thickened subepithelial basement membrane and around airways and blood vessels. The OVA‐treated mice showed increased expression of the α1, β1, and γ1 chains of the laminin‐1 isoform in monocytes, macrophages and eosinophils infiltrating the airways. Laminin‐1 receptor expression was increased in inflammatory and endothelial cells in the lungs of OVA‐treated mice compared to controls. Treatment of OVA‐sensitised/challenged mice with dexamethasone reduced airway expression of laminin and laminin‐1 receptor in OVA‐treated mice but not airway hyperresponsiveness to methacholine.Laminin deposition may be an important component of the airway remodelling observed in chronic allergic lung inflammation and is a process modulated by corticosteroids.