bluuebunny/crossref_metadata_2025_split huggingface dataset:
Assembly of the Type II Secretion System: Identification of ExeA Residues Critical for Peptidoglycan Binding and Secretin Multimerization
Authors: Li Gang , Miller Alicia , Bull Harold , Howard S. Peter 
Published: October 2010

ABSTRACTAeromonas hydrophilasecretes a number of protein toxins across the outer membrane via the type II secretion system (T2SS). Assembly of the secretion channel ExeD secretin into the outer membrane is dependent on the peptidoglycan binding domain of ExeA. In this study, the peptidoglycan binding domain PF01471 family members were divided into a prokaryotic group and a eukaryotic group. By comparison of their sequence conservation profiles and their representative crystal structures, we found the prokaryotic members to have a highly conserved pocket(s) that is not present in the eukaryotic members. Substitution mutations of nine amino acids of the pocket were constructed in ExeA. Five of the substitution derivatives showed greatly decreased lipase secretion, accompanied by defects in secretin assembly. In addition, usingin vivocross-linking andin vitrocosedimentation assays, we showed that these mutations decreased ExeA-peptidoglycan interactions. These results suggest that the highly conserved pocket in ExeA is the binding site for its peptidoglycan ligand and identify residues critical for this binding.
Insights into the Extracytoplasmic Stress Response ofXanthomonas campestrispv.campestris: Role and Regulation of σE-Dependent Activity
Authors: Bordes Patricia , Lavatine Laure , Phok Kounthéa , Barriot Roland , Boulanger Alice , Castanié-Cornet Marie-Pierre , Déjean Guillaume , Lauber Emmanuelle , Becker Anke , Arlat Matthieu , Gutierrez Claude 
Published: October 2010

ABSTRACTXanthomonas campestrispv.campestrisis an epiphytic bacterium that can become a vascular pathogen responsible for black rot disease of crucifers. To adapt gene expression in response to ever-changing habitats, phytopathogenic bacteria have evolved signal transduction regulatory pathways, such as extracytoplasmic function (ECF) σ factors. The alternative sigma factor σE, encoded byrpoE, is crucial for envelope stress response and plays a role in the pathogenicity of many bacterial species. Here, we combine different approaches to investigate the role and mechanism of σE-dependent activation inX. campestrispv.campestris. We show that therpoEgene is organized as a single transcription unit with the anti-σ generseAand the protease genemucDand thatrpoEtranscription is autoregulated.rseAandmucDtranscription is also controlled by a highly conserved σE-dependent promoter within the σEgene sequence. The σE-mediated stress response is required for stationary-phase survival, resistance to cadmium, and adaptation to membrane-perturbing stresses (elevated temperature and ethanol). Using microarray technology, we started to define the σEregulon ofX. campestrispv.campestris. These genes encode proteins belonging to different classes, including periplasmic or membrane proteins, biosynthetic enzymes, classical heat shock proteins, and the heat stress σ factor σH. The consensus sequence for the predicted σE-regulated promoter elements is GGAACTN15-17GTCNNA. Determination of therpoHtranscription start site revealed thatrpoHwas directly regulated by σEunder both normal and heat stress conditions. Finally, σEactivity is regulated by the putative regulated intramembrane proteolysis (RIP) proteases RseP and DegS, as previously described in many other bacteria. However, our data suggest that RseP and DegS are not only dedicated to RseA cleavage and that the proteolytic cascade of RseA could involve other proteases.
Structural and Functional Analysis of the Type III Secretion System fromPseudomonas fluorescensQ8r1-96
Authors: Mavrodi Dmitri V. , Joe Anna , Mavrodi Olga V. , Hassan Karl A. , Weller David M. , Paulsen Ian T. , Loper Joyce E. , Alfano James R. , Thomashow Linda S. 
Published: October 2010

ABSTRACTPseudomonas fluorescensQ8r1-96 represents a group of rhizosphere strains responsible for the suppressiveness of agricultural soils to take-all disease of wheat. It produces the antibiotic 2,4-diacetylphloroglucinol and aggressively colonizes the roots of cereal crops. In this study, we analyzed the genome of Q8r1-96 and identified a type III protein secretion system (T3SS) gene cluster that has overall organization similar to that of the T3SS gene cluster of the plant pathogenPseudomonas syringae. We also screened a collection of 30 closely relatedP. fluorescensstrains and detected the T3SS genes in all but one of them. The Q8r1-96 genome containedropAAandropMtype III effector genes, which are orthologs of theP. syringaeeffector geneshopAA1-1andhopM1, as well as a novel type III effector gene designatedropB. These type III effector genes encoded proteins that were secreted in culture and injected into plant cells by bothP. syringaeand Q8r1-96 T3SSs. The Q8r1-96 T3SS was expressed in the rhizosphere, but mutants lacking a functional T3SS were not altered in their rhizosphere competence. The Q8r1-96 type III effectors RopAA, RopB, and RopM were capable of suppressing the hypersensitive response and production of reactive oxygen species, two plant immune responses.
Nucleotide Parasitism by
Simkania negevensis
(
Chlamydiae
)
Authors: Knab Silvia , Mushak Tanja M. , Schmitz-Esser Stephan , Horn Matthias , Haferkamp Ilka 
Published: October 2010

ABSTRACT

Intracellular bacteria live in an environment rich in most essential metabolites but need special mechanisms to access these substrates. Nucleotide transport proteins (NTTs) catalyze the import of ATP and other nucleotides from the eukaryotic host into the bacterial cell and render
de novo
synthesis of these compounds dispensable. The draft genome sequence of
Simkania negevensis
strain Z, a chlamydial organism considered a newly emerging pathogen, revealed four genes encoding putative nucleotide transport proteins (
Sn
NTT1 to
Sn
NTT4), all of which are transcribed during growth of
S. negevensis
in
Acanthamoeba
host cells, as confirmed by reverse transcription-PCR. Using heterologous expression in
Escherichia coli
, we could show that
Sn
NTT1 functions as an ATP/ADP antiporter,
Sn
NTT2 as a guanine nucleotide/ATP/H
+
symporter driven by the membrane potential, and
Sn
NTT3 as a nucleotide triphosphate antiporter. In addition,
Sn
NTT3 is able to transport dCTP, which has not been shown for a prokaryotic transport protein before. No substrate could be identified for
Sn
NTT4. Taking these data together,
S. negevensis
employs a set of nucleotide transport proteins to efficiently tap its host's energy and nucleotide pools. Although similar to other chlamydiae, these transporters show distinct and unique adaptations with respect to substrate specificities and mode of transport.
Mismatch Repair Modulation of MutY Activity Drives
Bacillus subtilis
Stationary-Phase Mutagenesis
Authors: Debora Bernardo N. , Vidales Luz E. , Ramírez Rosario , Ramírez Mariana , Robleto Eduardo A. , Yasbin Ronald E. , Pedraza-Reyes Mario 
Published: October 2010

ABSTRACT

Stress-promoted mutations that occur in nondividing cells (adaptive mutations) have been implicated strongly in causing genetic variability as well as in species survival and evolutionary processes. Oxidative stress-induced DNA damage has been associated with generation of adaptive His
+
and Met
+
but not Leu
+
revertants in strain
Bacillus subtilis
YB955 (
hisC952 metB5 leuC427
). Here we report that an interplay between MutY and MutSL (mismatch repair system [MMR]) plays a pivotal role in the production of adaptive Leu
+
revertants. Essentially, the genetic disruption of MutY dramatically reduced the reversion frequency to the
leu
allele in this model system. Moreover, the increased rate of adaptive Leu
+
revertants produced by a MutSL knockout strain was significantly diminished following
mutY
disruption. Interestingly, although the expression of
mutY
took place during growth and stationary phase and was not under the control of RecA, PerR, or σ
B
, a null mutation in the
mutSL
operon increased the expression of
mutY
several times. Thus, in starved cells, saturation of the MMR system may induce the expression of
mutY
, disturbing the balance between MutY and MMR proteins and aiding in the production of types of mutations detected by reversion to leucine prototrophy. In conclusion, our results support the idea that MMR regulation of the mutagenic/antimutagenic properties of MutY promotes stationary-phase mutagenesis in
B. subtilis
cells.
Mapping Interactions between Germinants and
Clostridium difficile
Spores
Authors: Howerton Amber , Ramirez Norma , Abel-Santos Ernesto 
Published: October 2010

ABSTRACT

Germination of
Clostridium difficile
spores is the first required step in establishing
C. difficile
-associated disease (CDAD). Taurocholate (a bile salt) and glycine (an amino acid) have been shown to be important germinants of
C. difficile
spores. In the present study, we tested a series of glycine and taurocholate analogs for the ability to induce or inhibit
C. difficile
spore germination. Testing of glycine analogs revealed that both the carboxy and amino groups are important epitopes for recognition and that the glycine binding site can accommodate compounds with more widely separated termini. The
C. difficile
germination machinery also recognizes other hydrophobic amino acids. In general, linear alkyl side chains are better activators of spore germination than their branched analogs. However,
l
-phenylalanine and
l
-arginine are also good germinants and are probably recognized by distinct binding sites. Testing of taurocholate analogs revealed that the 12-hydroxyl group of taurocholate is necessary, but not sufficient, to activate spore germination. In contrast, the 6- and 7-hydroxyl groups are required for inhibition of
C. difficile
spore germination. Similarly,
C. difficile
spores are able to detect taurocholate analogs with shorter, but not longer, alkyl amino sulfonic acid side chains. Furthermore, the sulfonic acid group can be partially substituted with other acidic groups. Finally, a taurocholate analog with an
m
-aminobenzenesulfonic acid side chain is a strong inhibitor of
C. difficile
spore germination. In conclusion,
C. difficile
spores recognize both amino acids and taurocholate through multiple interactions that are required to bind the germinants and/or activate the germination machinery.
Regulation and Function of
Escherichia coli
Sugar Efflux Transporter A (SetA) during Glucose-Phosphate Stress
Authors: Sun Yan , Vanderpool Carin K. 
Published: October 2010

ABSTRACT

Accumulation of certain nonmetabolizable sugar-phosphates (including α-methyl glucoside-6-phosphate) in
Escherichia coli
is growth inhibitory and elicits the glucose-phosphate stress response. The transcription factor SgrR activates transcription of the small RNA SgrS under stress conditions. SgrS represses translation of mRNAs encoding sugar transporters. The
sgrR
and
sgrS
genes are located directly upstream of
setA
, and this gene organization is conserved in numerous enteric species, prompting the hypothesis that SetA contributes to the glucose-phosphate stress response. SetA is a proton motive force-driven efflux pump capable of transporting various sugars and sugar analogs
in vitro
. This study demonstrates that
setA
expression is induced in response to glucose-phosphate stress, and this requires SgrR. Under stress conditions,
setA
is cotranscribed with
sgrS
from the
sgrS
promoter. A
setA
mutant exhibits a growth defect under stress conditions that can be complemented by
setA
in
trans
, suggesting that SetA contributes to the optimal cellular recovery from stress. Despite previous
in vitro
evidence that SetA can promote efflux of the stress-causing glucose analog α-methyl glucoside,
in vivo
data in this study indicate that SetA is not the major efflux pump responsible for removal of α-methyl glucoside under stress conditions.
The Nodulation of Alfalfa by the Acid-TolerantRhizobiumsp. Strain LPU83 Does Not Require Sulfated Forms of Lipochitooligosaccharide Nodulation Signals
Authors: Torres Tejerizo Gonzalo , Del Papa María Florencia , Soria-Diaz M. Eugenia , Draghi Walter , Lozano Mauricio , Giusti María de los Ángeles , Manyani Hamid , Megías Manuel , Gil Serrano Antonio , Pühler Alfred , Niehaus Karsten , Lagares Antonio , Pistorio Mariano 
Published: October 2010

ABSTRACTThe induction of root nodules by the majority of rhizobia has a strict requirement for the secretion of symbiosis-specific lipochitooligosaccharides (nodulation factors [NFs]). The nature of the chemical substitution on the NFs depends on the particular rhizobium and contributes to the host specificity imparted by the NFs. We present here a description of the genetic organization of thenodgene cluster and the characterization of the chemical structure of the NFs associated with the broad-host-rangeRhizobiumsp. strain LPU83, a bacterium capable of nodulating at least alfalfa, bean, andLeucena leucocephala. Thenodgene cluster was located on the plasmid pLPU83b. The organization of the cluster showed synteny with those of the alfalfa-nodulating rhizobia,Sinorhizobium melilotiandSinorhizobium medicae. Interestingly, the strongest sequence similarity observed was between the partialnodsequences ofRhizobium mongolenseUSDA 1844 and the corresponding LPU83nodgenes sequences. The phylogenetic analysis of the intergenic regionnodEGpositions strain LPU83 and the type strainR. mongolense1844 in the same branch, which indicates thatRhizobiumsp. strain LPU83 might represent an early alfalfa-nodulating genotype. The NF chemical structures obtained for the wild-type strain consist of a trimeric, tetrameric, and pentameric chitin backbone that shares some substitutions with both alfalfa- and bean-nodulating rhizobia. Remarkably, while in strain LPU83 most of the NFs were sulfated in their reducing terminal residue, none of the NFs isolated from thenodHmutant LPU83-H were sulfated. The evidence obtained supports the notion that the sulfate decoration of NFs in LPU83 is not necessary for alfalfa nodulation.
The Biocide Chlorine Dioxide Stimulates Biofilm Formation inBacillus subtilisby Activation of the Histidine Kinase KinC
Authors: Shemesh Moshe , Kolter Roberto , Losick Richard 
Published: October 2010

ABSTRACTBacillus subtilisforms biofilms in response to signals that remain poorly defined. We report that biofilm formation is stimulated by sublethal doses of chlorine dioxide (ClO2), an extremely effective and fast-acting biocide. ClO2accelerated biofilm formation inB. subtilisas well as in other bacteria, suggesting that biofilm formation is a widely conserved response to sublethal doses of the agent. Biofilm formation depends on the synthesis of an extracellular matrix that holds the constituent cells together. We show that the transcription of the major operons responsible for the matrix production inB. subtilis,epsA-epsOandyqxM-sipW-tasA, was enhanced by ClO2, in a manner that depended on the membrane-bound kinase KinC. Activation of KinC appeared to be due to the ability of ClO2to collapse the membrane potential. Importantly, strains unable to make a matrix were hypersensitive to ClO2, indicating that biofilm formation is a defensive response that helps protect cells from the toxic effects of the biocide.
Effects of Deficiency and Overdose of Group 2 Sigma Factors in Triple Inactivation Strains of
Synechocystis
sp. Strain PCC 6803
Authors: Pollari Maija , Rantamäki Susanne , Huokko Tuomas , Kårlund-Marttila Anna , Virjamo Virpi , Tyystjärvi Esa , Tyystjärvi Taina 
Published: October 2010

ABSTRACT

Acclimation of cyanobacteria to environmental changes includes major changes in the gene expression patterns partly orchestrated by the replacement of a particular σ subunit with another in the RNA polymerase holoenzyme. The cyanobacterium
Synechocystis
sp. strain PCC 6803 encodes nine σ factors, all belonging to the σ
70
family. Cyanobacteria typically encode many group 2 σ factors that closely resemble the principal σ factor. We inactivated three out of the four group 2 σ factors of
Synechocystis
simultaneously in all possible combinations and found that all triple inactivation strains grow well under standard conditions. Unlike the other strains, the Δ
sigBCD
strain, which contains SigE as the only functional group 2 σ factor, did not grow faster under mixotrophic than under autotrophic conditions. The SigB and SigD factors were important in low-temperature acclimation, especially under diurnal light rhythm. The Δ
sigBCD
, Δ
sigBCE
, and Δ
sigBDE
strains were sensitive to high-light-induced photoinhibition, indicating a central role of the SigB factor in high-light tolerance. Furthermore, the Δ
sigBCE
strain (SigD is the only functional group 2 σ factor) appeared to be locked in the high-fluorescence state (state 1) and grew slowly in blue but not in orange or white light. Our results suggest that features of the triple inactivation strains can be categorized as (i) direct consequences of the inactivation of a particular σ factor(s) and (ii) effects resulting from the higher probability that the remaining group 2 σ factors associate with the RNA polymerase core.
Glucose-Dependent Activation ofBacillus anthracisToxin Gene Expression and Virulence Requires the Carbon Catabolite Protein CcpA
Authors: Chiang Christina , Bongiorni Cristina , Perego Marta 
Published: October 2010

ABSTRACTSensing environmental conditions is an essential aspect of bacterial physiology and virulence. InBacillus anthracis, the causative agent of anthrax, transcription of the two major virulence factors, toxin and capsule, is triggered by bicarbonate, a major compound in the mammalian body. Here it is shown that glucose is an additional signaling molecule recognized byB. anthracisfor toxin synthesis. The presence of glucose increased the expression of the protective antigen toxin component-encoding gene (pagA) by stimulating induction of transcription of the AtxA virulence transcription factor. Induction ofatxAtranscription by glucose required the carbon catabolite protein CcpA via an indirect mechanism. CcpA did not bind specifically to any region of the extendedatxApromoter. The virulence of aB. anthracisstrain from which theccpAgene was deleted was significantly attenuated in a mouse model of infection. The data demonstrated that glucose is an important host environment-derived signaling molecule and that CcpA is a molecular link between environmental sensing andB. anthracispathogenesis.
Glypican-3 expression predicts poor clinical outcome of patients with early-stage clear cell carcinoma of the ovary
Authors: Umezu Tomokazu , Shibata Kiyosumi , Kajiyama Hiroaki , Yamamoto Eiko , Nawa Akihiro , Kikkawa Fumitaka 
Published: October 2010

BackgroundGlypican-3 (GPC3), a membrane-bound heparan sulphate proteoglycan, may play a role in promoting cancer cell growth and differentiation. Recent studies reported that GPC3 is overexpressed in clear cell carcinoma (CCC) of the ovary, and not other ovarian histotypes. However, in CCC patients, the relationship between the overexpression of GPC3 and prognosis has not yet been clarified.AimTo evaluate GPC3 expression by immunohistochemistry in CCC.Methods and ResultsIn 52 CCC patients, GPC3 expression was observed in 40.4%. In cases of CCC, no correlations were identified between GPC3 expression and clinicopathological factors, such as age, FIGO stage, CA125 values, peritoneal cytology, ascitic fluid volume and mortality rate, except for the residual tumour size. GPC3 expression was associated with poor progression-free survival in stage I CCC patients. The numbers of Ki-67-stained cells in GPC3-positive areas were lower than those in GPC3-negative areas. GPC3 expression may be associated with a low proliferation rate in CCC cells. In the early stage of CCC, GPC3-expressing patients tended to be resistant to taxane-based treatment.ConclusionsResults suggest that the overexpression of GPC3 may be related to the low-level proliferation of tumours; it may be associated with resistance to taxane-based chemotherapy and a poor prognosis in CCC of the ovary.
Management of in situ lobular neoplasia detected on needle core biopsy of breast
Authors: Purdie Colin A , McLean Denis , Stormonth Elizabeth , Macaskill E Jane , McCullough Jean B , Edwards Sharon L , Brown Douglas C , Jordan Lee B 
Published: October 2010

AimsTo evaluate the risk of having occult ductal carcinoma in situ or invasive carcinoma in the region of a focus of lobular (in situ) neoplasia (LN) diagnosed on needle core biopsy (NCB) of breast.MethodsAll cases of LN diagnosed on NCB of breast over 10 years (2000–2009 inclusive) were reviewed. The clinical presentation, radiological appearances and final pathological diagnosis on open diagnostic biopsy (ODB) were correlated.Results125 cases of LN on NCB were identified from diagnostic codes. Of these, 72 (58%) had a coexistent, higher-grade lesion that mandated surgery. Fifty of the remaining 53 (94%) underwent ODB. The majority of patients were asymptomatic, with 68% presenting through the breast screening programme, and in 89% of patients, the target abnormality was microcalcification. Of the 50 patients, 13 (26%) had a final diagnosis of in situ or invasive carcinoma requiring therapeutic surgery. When the cases of pleomorphic LN were excluded, 21% (10/47) were upgraded. Two of these 10 cases had discordant radiology which could have been diagnosed on repeat NCB leaving an upgrade rate of 18% (8/45). In four of the eight cases of invasive malignancy, the disease was multifocal.ConclusionsLN is frequently asymptomatic, being identified by mammographic microcalcification alone. In 21% of classical LN cases, it is associated with an undiagnosed, higher-grade lesion requiring oncological management. In our view, patients with LN discovered on NCB should undergo open diagnostic biopsy.
Zoledronic Acid Potentiates mTOR Inhibition and Abolishes the Resistance of Osteosarcoma Cells to RAD001 (Everolimus): Pivotal Role of the Prenylation Process
Authors: Moriceau Gatien , Ory Benjamin , Mitrofan Laura , Riganti Chiara , Blanchard Frédéric , Brion Régis , Charrier Céline , Battaglia Séverine , Pilet Paul , Denis Marc G. , Shultz Leonard D. , Mönkkönen Jukka , Rédini Françoise , Heymann Dominique 
Published: October 2010

Abstract
Despite recent improvements in therapeutic management of osteosarcoma, ongoing challenges in improving the response to chemotherapy warrants new strategies still needed to improve overall patient survival. In this study, we investigated in vivo the effects of RAD001 (Everolimus), a new orally available mTOR inhibitor, on the growth of human and mouse osteosarcoma cells either alone or in combination with zoledronate (ZOL), an anti-osteoporotic drug used to treat bone metastases. RAD001 inhibited osteosarcoma cell proliferation in a dose- and time-dependent manner with no modification of cell-cycle distribution. Combination with ZOL augmented this inhibition of cell proliferation, decreasing PI3K/mTOR signaling compared with single treatments. Notably, in contrast to RAD001, ZOL downregulated isoprenylated membrane-bound Ras concomitantly with an increase of nonisoprenylated cytosolic Ras in sensitive and resistant osteosarcoma cell lines to both drugs. Moreover, ZOL and RAD001 synergized to decrease Ras isoprenylation and GTP-bound Ras levels. Further, the drug combination reduced tumor development in two murine models of osteoblastic or osteolytic osteosarcoma. We found that ZOL could reverse RAD001 resistance in osteosarcoma, limiting osteosarcoma cell growth in combination with RAD001. Our findings rationalize further study of the applications of mTOR and mevalonate pathway inhibitors that can limit protein prenylation pathways. Cancer Res; 70(24); 10329–39. ©2010 AACR.