bluuebunny/crossref_metadata_2025_split huggingface dataset:
SciN Is an Outer Membrane Lipoprotein Required for Type VI Secretion in EnteroaggregativeEscherichia coli
Authors: Aschtgen Marie-Stéphanie , Bernard Christophe S. , De Bentzmann Sophie , Lloubès Roland , Cascales Eric 
Published: September 2008

ABSTRACTEnteroaggregativeEscherichia coli(EAEC) is a pathogen implicated in several infant diarrhea or diarrheal outbreaks in areas of endemicity. Although multiple genes involved in EAEC pathogenesis have been identified, the overall mechanism of virulence is not well understood. Recently, a novel secretion system, called type VI secretion (T6S) system (T6SS), has been identified in EAEC and most animal or plant gram-negative pathogens. T6SSs are multicomponent cell envelope machines responsible for the secretion of at least two putative substrates, Hcp and VgrG. In EAEC, two copies of T6S gene clusters, calledsci-1andsci-2, are present on thepheUpathogenicity island. In this study, we focused our work on thesci-1gene cluster. The Sci-1 apparatus is probably composed of all, or a subset of, the 21 gene products encoded on the cluster. Among these subunits, some are shared by all T6SSs identified to date, including a ClpV-type AAA+ATPase (SciG) and an IcmF (SciS) and an IcmH (SciP) homologue, as well as a putative lipoprotein (SciN). In this study, we demonstrate thatsciNis a critical gene necessary for T6S-dependent secretion of the Hcp-like SciD protein and for biofilm formation. We further show that SciN is a lipoprotein, as shown by the inhibition of its processing by globomycin and in vivo labeling with [3H]palmitic acid. SciN is tethered to the outer membrane and exposed in the periplasm. Sequestration of SciN at the inner membrane by targeting the +2 residue responsible for lipoprotein localization (Gly2Asp) fails to complement ansciNmutant for SciD secretion and biofilm formation. Together, these results support a model in which SciN is an outer membrane lipoprotein exposed in the periplasm and essential for the Sci-1 apparatus function.
Characterization of a
Streptococcus
sp.-
Veillonella
sp. Community Micromanipulated from Dental Plaque
Authors: Chalmers Natalia I. , Palmer Robert J. , Cisar John O. , Kolenbrander Paul E. 
Published: September 2008

ABSTRACT

Streptococci and veillonellae occur in mixed-species colonies during formation of early dental plaque. One factor hypothesized to be important in assembly of these initial communities is coaggregation (cell-cell recognition by genetically distinct bacteria). Intrageneric coaggregation of streptococci occurs when a lectin-like adhesin on one streptococcal species recognizes a receptor polysaccharide (RPS) on the partner species. Veillonellae also coaggregate with streptococci. These genera interact metabolically; lactic acid produced by streptococci is a carbon source for veillonellae. To transpose these interactions from undisturbed dental plaque to an experimentally tractable in vitro biofilm model, a community consisting of RPS-bearing streptococci juxtaposed with veillonellae was targeted by quantum dot-based immunofluorescence and then micromanipulated off the enamel surface and cultured. Besides the expected antibody-reactive cell types, a non-antibody-reactive streptococcus invisible during micromanipulation was obtained. The streptococci were identified as
Streptococcus oralis
(RPS bearing) and
Streptococcus gordonii
(adhesin bearing). The veillonellae could not be cultivated; however, a veillonella 16S rRNA gene sequence was amplified from the original isolation mixture, and this sequence was identical to the sequence of the previously studied organism
Veillonella
sp. strain PK1910, an oral isolate in our culture collection.
S. oralis
coaggregated with
S. gordonii
by an RPS-dependent mechanism, and both streptococci coaggregated with PK1910, which was used as a surrogate during in vitro community reconstruction. The streptococci and strain PK1910 formed interdigitated three-species clusters when grown as a biofilm using saliva as the nutritional source. PK1910 grew only when streptococci were present. This study confirms that RPS-mediated intrageneric coaggregation occurs in the earliest stages of plaque formation by bringing bacteria together to create a functional community.
Mutations in Conserved Helix 69 of 23S rRNA of
Thermus thermophilus
That Affect Capreomycin Resistance but Not Posttranscriptional Modifications
Authors: Monshupanee Tanakarn , Gregory Steven T. , Douthwaite Stephen , Chungjatupornchai Wipa , Dahlberg Albert E. 
Published: September 2008

ABSTRACT

Translocation during the elongation phase of protein synthesis involves the relative movement of the 30S and 50S ribosomal subunits. This movement is the target of tuberactinomycin antibiotics. Here, we describe the isolation and characterization of mutants of
Thermus thermophilus
selected for resistance to the tuberactinomycin antibiotic capreomycin. Two base substitutions, A1913U and mU1915G, and a single base deletion, ΔmU1915, were identified in helix 69 of 23S rRNA, a structural element that forms part of an interribosomal subunit bridge with the decoding center of 16S rRNA, the site of previously reported capreomycin resistance base substitutions. Capreomycin resistance in other bacteria has been shown to result from inactivation of the TlyA methyltransferase which 2′-O methylates C1920 of 23S rRNA. Inactivation of the
tlyA
gene in
T. thermophilus
does not affect its sensitivity to capreomycin. Finally, none of the mutations in helix 69 interferes with methylation at C1920 or with pseudouridylation at positions 1911 and 1917. We conclude that the resistance phenotype is a consequence of structural changes introduced by the mutations.
Characterization of Conserved Bases in 4.5S RNA ofEscherichia coliby Construction of New F′ Factors
Authors: Peterson James M. , Phillips Gregory J. 
Published: September 2008

ABSTRACTTo more clearly understand the function of conserved bases of 4.5S RNA, the product of the essentialffsgene ofEscherichia coli, and to address conflicting results reported in other studies, we have developed a new genetic system to characterizeffsmutants. Multipleffsalleles were generated by altering positions that correspond to the region of the RNA molecule that interacts directly with Ffh in assembly of the signal recognition particle. To facilitate characterization of theffsmutations with minimal manipulation, recombineering was used to construct new F′ factors to easily move each allele into different genetic backgrounds for expression in single copy. In combination with plasmids that expressedffsin multiple copy numbers, the F′ factors provided an accurate assessment of the ability of the different 4.5S RNA mutants to function in vivo. Consistent with structural analysis of the signal recognition particle (SRP), highly conserved bases in 4.5S RNA are important for binding Ffh. Despite the high degree of conservation, however, only a single base (C62) was indispensable for RNA function under all conditions tested. To quantify the interaction between 4.5S RNA and Ffh, an assay was developed to measure the ability of mutant 4.5S RNA molecules to copurify with Ffh. Defects in Ffh binding correlated with loss of SRP-dependent protein localization. Real-time quantitative PCR was also used to measure the levels of wild-type and mutant 4.5S RNA expressed in vivo. These results clarify inconsistencies from prior studies and yielded a convenient method to study the function of multiple alleles.
Synergistic Contribution of theLegionella pneumophila lqsGenes to Pathogen-Host Interactions
Authors: Tiaden André , Spirig Thomas , Carranza Paula , Brüggemann Holger , Riedel Kathrin , Eberl Leo , Buchrieser Carmen , Hilbi Hubert 
Published: September 2008

ABSTRACTThe causative agent of Legionnaires' disease,Legionella pneumophila, is a natural parasite of environmental protozoa and employs a biphasic life style to switch between a replicative and a transmissive (virulent) phase.L. pneumophilaharbors thelqs(Legionella quorumsensing) cluster, which includes genes encoding the autoinducer synthase LqsA, the sensor kinase LqsS, the response regulator LqsR, and a homologue of HdeD, which is involved in acid resistance inEscherichia coli. LqsR promotes host-cell interactions as an element of the stationary-phase virulence regulatory network. Here, we characterizeL. pneumophilamutant strains lacking all four genes of thelqscluster or only thehdeDgene. While anhdeDmutant strain did not have overt physiological or virulence phenotypes, anlqsmutant showed an aberrant morphology in stationary growth phase and was defective for intracellular growth, efficient phagocytosis, and cytotoxicity against host cells. Cytotoxicity was restored upon reintroduction of thelqsgenes into the chromosome of anlqsmutant strain. The deletion of thelqscluster caused more-severe phenotypes than deletion of onlylqsR, suggesting a synergistic effect of the otherlqsgenes. A transcriptome analysis indicated that in the stationary phase more than 380 genes were differentially regulated in thelqsmutant and wild-typeL. pneumophila. Genes involved in protein production, metabolism, and bioenergetics were upregulated in thelqsmutant, whereas genes encoding virulence factors, such as effectors secreted by the Icm/Dot type IV secretion system, were downregulated. A proteome analysis revealed that a set of Icm/Dot substrates is not produced in the absence of thelqsgene cluster, which confirms the findings from DNA microarray assays and mirrors the virulence phenotype of thelqsmutant strain.
Alr0397 Is an Outer Membrane Transporter for the Siderophore Schizokinen inAnabaenasp. Strain PCC 7120
Authors: Nicolaisen Kerstin , Moslavac Suncana , Samborski Anastazia , Valdebenito Marianne , Hantke Klaus , Maldener Iris , Muro-Pastor Alicia M. , Flores Enrique , Schleiff Enrico 
Published: September 2008

ABSTRACTIron uptake in proteobacteria by TonB-dependent outer membrane transporters represents a well-explored subject. In contrast, the same process has been scarcely investigated in cyanobacteria. The heterocyst-forming cyanobacteriumAnabaenasp. strain PCC 7120 is known to secrete the siderophore schizokinen, but its transport system has remained unidentified. Inspection of the genome of strain PCC 7120 shows that only one gene encoding a putative TonB-dependent iron transporter, namely alr0397, is positioned close to genes encoding enzymes involved in the biosynthesis of a hydroxamate siderophore. The expression of alr0397, which encodes an outer membrane protein, was elevated under iron-limited conditions. Inactivation of this gene caused a moderate phenotype of iron starvation in the mutant cells. The characterization of the mutant strain showed that Alr0397 is a TonB-dependent schizokinen transporter (SchT) of the outer membrane and that alr0397 expression and schizokinen production are regulated by the iron homeostasis of the cell.
Posttranslational Modification Influences the Effects of MgrA on
norA
Expression in
Staphylococcus aureus
Authors: Truong-Bolduc Que Chi , Ding Yanpeng , Hooper David C. 
Published: September 2008

ABSTRACT

MgrA is a global regulator in
Staphylococcus aureus
. Differences in the effects of MgrA on
norA
expression have been reported for different strains, which varied in
rsbU
, a gene that affects the expression of
sigB
, which encodes an alternative σ factor involved in stress responses. We hypothesized that MgrA was modified by
sigB
-dependent factors that affected its ability to control the expression of the
norA
efflux pump. Heterologously expressed MgrA purified from
Escherichia coli
was incubated with crude extracts (CE) from strains RN6390 (
rsbU
) and SH1000 (
rsbU
+
) and tested for binding to the
norA
promoter. Purified MgrA exhibited greater binding to
norA
promoter DNA after being incubated with SH1000 CE than MgrA incubated with the RN6390 CE. Phosphorylation of MgrA occurring in cell extracts caused it to lose the ability to bind
norA
promoter DNA. Overexpression of
pknB
, encoding a candidate serine/threonine kinase, produced increased phospho-MgrA and led to a fivefold increase in the transcript level of
norA
for both RN6390 and SH1000, as well as a fourfold increase in the MICs of norfloxacin and ciprofloxacin for these two strains. The levels of expression of
pknB
in RN6390 and SH1000, however, indicated that additional factors related to
rsbU
or
sigB
contribute to the differential regulatory effects of MgrA on
norA
expression.
Modification of
Pseudomonas aeruginosa
Pa5196 Type IV Pilins at Multiple Sites with
d
-Ara
f
by a Novel GT-C Family Arabinosyltransferase, TfpW
Authors: Kus Julianne V. , Kelly John , Tessier Luc , Harvey Hanjeong , Cvitkovitch Dennis G. , Burrows Lori L. 
Published: September 2008

ABSTRACT

Pseudomonas aeruginosa
Pa5196 produces type IV pilins modified with unusual α1,5-linked
d
-arabinofuranose (α1,5-
d
-Ara
f
) glycans, identical to those in the lipoarabinomannan and arabinogalactan cell wall polymers from
Mycobacterium
spp. In this work, we identify a second strain of
P. aeruginosa
, PA7, capable of expressing arabinosylated pilins and use a combination of site-directed mutagenesis, electrospray ionization mass spectrometry (MS), and electron transfer dissociation MS to identify the exact sites and extent of pilin modification in strain Pa5196. Unlike previously characterized type IV pilins that are glycosylated at a single position, those from strain Pa5196 were modified at multiple sites, with modifications of αβ-loop residues Thr64 and Thr66 being important for normal pilus assembly. Trisaccharides of α1,5-
d
-Ara
f
were the principal modifications at Thr64 and Thr66, with additional mono- and disaccharides identified on Ser residues within the antiparallel beta sheet region of the pilin. TfpW was hypothesized to encode the pilin glycosyltransferase based on its genetic linkage to the pilin, weak similarity to membrane-bound GT-C family glycosyltransferases (which include the
Mycobacterium
arabinosyltransferases EmbA/B/C), and the presence of characteristic motifs. Loss of TfpW or mutation of key residues within the signature GT-C glycosyltransferase motif completely abrogated pilin glycosylation, confirming its involvement in this process. A Pa5196
pilA
mutant complemented with other
Pseudomonas
pilins containing potential sites of modification expressed nonglycosylated pilins, showing that TfpW's pilin substrate specificity is restricted. TfpW is the prototype of a new type IV pilin posttranslational modification system and the first reported gram-negative member of the GT-C glycosyltransferase family.
Regulation of the
Salmonella enterica std
Fimbrial Operon by DNA Adenine Methylation, SeqA, and HdfR
Authors: Jakomin Marcello , Chessa Daniela , Bäumler Andreas J. , Casadesús Josep 
Published: September 2008

ABSTRACT

DNA adenine methylase (
dam
) mutants of
Salmonella enterica
serovar Typhimurium grown under laboratory conditions express the
std
fimbrial operon, which is tightly repressed in the wild type. Here, we show that uncontrolled production of Std fimbriae in
S. enterica
serovar Typhimurium
dam
mutants contributes to attenuation in mice, as indicated by the observation that an
stdA dam
strain is more competitive than a
dam
strain upon oral infection. Dam methylation appears to regulate
std
transcription, rather than
std
mRNA stability or turnover. A genetic screen for
std
regulators showed that the GATC-binding protein SeqA directly or indirectly represses
std
expression, while the poorly characterized
yifA
gene product serves as an
std
activator. YifA encodes a putative LysR-like protein and has been renamed HdfR, like its
Escherichia coli
homolog. Activation of
std
expression by HdfR is observed only in
dam
and
seqA
backgrounds. These data suggest that HdfR directly or indirectly activates
std
transcription. Since SeqA is unable to bind nonmethylated DNA, it is possible that
std
operon derepression in
dam
and
seqA
mutants may result from unconstrained HdfR-mediated activation of
std
transcription. Derepression of
std
in
dam
and
seqA
mutants of
S. enterica
occurs in only a fraction of the bacterial population, suggesting the occurrence of either bistable expression or phase variation.
Transcriptional Regulation of the Citrate Gene Cluster of
Enterococcus faecalis
Involves the GntR Family Transcriptional Activator CitO
Authors: Blancato Víctor S. , Repizo Guillermo D. , Suárez Cristian A. , Magni Christian 
Published: September 2008

ABSTRACT

The genome of the gram-positive bacterium
Enterococcus faecalis
contains the genes that encode the citrate lyase complex. This complex splits citrate into oxaloacetate and acetate and is involved in all the known anaerobic bacterial citrate fermentation pathways. Although citrate fermentation in
E. faecalis
has been investigated before, the regulation and transcriptional pattern of the
cit
locus has still not been fully explored. To fill this gap, in this paper we demonstrate that the GntR transcriptional regulator CitO is a novel positive regulator involved in the expression of the
cit
operons. The transcriptional analysis of the
cit
clusters revealed two divergent operons:
citHO
, which codes for the transporter (
citH
) and the regulatory protein (
citO
), and upstream from it and in the opposite direction the
oadHDB-citCDEFX-oadA-citMG
operon, which includes the citrate lyase subunits (
citD
,
citE
, and
citF
), the soluble oxaloacetate decarboxylase (
citM
), and also the genes encoding a putative oxaloacetate decarboxylase complex (
oadB
,
oadA
,
oadD
and
oadH
). This analysis also showed that both operons are specifically activated by the addition of citrate to the medium. In order to study the functional role of CitO, a mutant strain with an interrupted
citO
gene was constructed, causing a total loss of the ability to degrade citrate. Reintroduction of a functional copy of
citO
to the
citO
-deficient strain restored the response to citrate and the Cit
+
phenotype. Furthermore, we present evidence that CitO binds to the
cis
-acting sequences O
1
and O
2
, located in the
cit
intergenic region, increasing its affinity for these binding sites when citrate is present and allowing the induction of both
cit
promoters.
Mortality and cancer incidence of Australian participants in the British nuclear tests in Australia
Authors: Gun R T , Parsons J , Crouch P , Ryan P , Hiller J E 
Published: September 2008

Objectives:To investigate the mortality and cancer incidence of Australian nuclear test participants, and to identify any association with exposure to ionising radiation.Methods:A retrospective cohort study was carried out in which the mortality and cancer incidence rates of participants (n = 10 983) were compared with rates in the general male Australian population. Dose reconstructions were carried out by a panel of health physicists. Mortality and cancer rates were compared with the general population and between groups of subjects categorised by assessed radiation exposure.Results:All-cause mortality was not raised. Mortality and incidence were significantly raised for cancers of the head and neck, lung, colon and rectum, and prostate, and for all cancers combined. For oesophageal cancer, melanoma, all leukaemias and non-chronic lymphatic leukaemia (non-CLL leukaemia), incidence was significantly raised, but mortality was non-significantly raised. No association was found between radiation exposure and overall cancer incidence or mortality, or with any cancer or cancer deaths occurring in excess.Conclusions:There is no evidence that the excess cancers and cancer deaths were caused by radiation exposure at the test sites. Possible contributing factors are high smoking prevalence and demographic differences from the Australian population with whom rates were compared. Asbestos is a likely contributor to some cancers in naval personnel.
Occupation/industry and risk of non-Hodgkin’s lymphoma in the United States
Authors: Schenk M , Purdue M P , Colt J S , Hartge P , Blair A , Stewart P , Cerhan J R , De Roos A J , Cozen W , Severson R K 
Published: September 2008

Aims:To identify occupations and industries associated with non-Hodgkin’s lymphoma (NHL) in a large population-based, case-control study in the USA.Methods:Cases (n = 1189) of histologically confirmed malignant NHL ages 20–74 were prospectively identified in four geographic areas covered by the National Cancer Institute SEER Program. Controls (n = 982) were selected from the general population by random digit dialling (<65 years of age) and from residents listed in Medicare files (65–74 years of age). Odds ratios and 95% confidence intervals for occupations and industries were calculated by unconditional logistic regression analyses, adjusting for age, gender, ethnicity and study centre. Further analyses stratified for gender and histological subtype were also performed.Results:Risk of NHL was increased for a few occupations and industries. Several white collar occupations, with no obvious hazardous exposures, had elevated risks, including purchasing agents and buyers, religious workers, physical therapists and information clerks. Occupations with excesses that may have exposures of interest include launderers and ironers, service occupations, food/beverage preparation supervisors, hand packers and packagers, roofing and siding, leather and leather products, transportation by air, nursing and personal care facilities, and specialty outpatient clinics. Significantly decreased risks of NHL were found for a number of occupations and industries including post-secondary teachers and chemical and allied products.Conclusions:The results of this study suggest that several occupations and industries may alter the risk of NHL. Our results support previously reported increased risks among farmers, printers, medical professionals, electronic workers and leather workers. These findings should be evaluated further in larger studies that have the power to focus on specific exposures and histological subtypes of NHL.
Work pattern causes bias in self-reported activity duration: a randomised study of mechanisms and implications for exposure assessment and epidemiology
Authors: Barrero L H , Katz J N , Perry M J , Krishnan R , Ware J H , Dennerlein J T 
Published: September 2008

Background:Self-reported activity duration is used to estimate cumulative exposures in epidemiological research.Objective:The effects of work pattern, self-reported task dullness (a measure of cognitive task demand), and heart rate ratio and perceived physical exertion (measures of physical task demands) on error in task duration estimation were investigated.Methods:24 participants (23–54 years old, 12 males) were randomly assigned to execute three tasks in either a continuous (three periods of 40 continuous minutes, one for each task) or a discontinuous work pattern (40 min tasks each divided into four periods of 4, 8, 12 and 16 min). Heart rate was measured during tasks. After completing the 2 h work session, subjects reported the perceived duration, dullness and physical exertion for each of the three tasks. Multivariate models were fitted to analyse errors and their absolute value to assess the accuracy in task duration estimation and the mediating role of task demands on the observed results.Results:Participants overestimated the time spent shelving boxes (up to 38%) and filing journals (up to 9%), and underestimated the time typing articles (up to −22%). Over- and underestimates and absolute errors were greater in the discontinuous work pattern group. Only the self-reported task dullness mediated the differences in task duration estimation accuracy between work patterns.Conclusions:Task-related factors can affect self-reported activity duration. Exposure assessment strategies requiring workers to allocate work time to different tasks could result in biased measures of association depending on the demands of the tasks during which the exposure of interest occurs.