bluuebunny/crossref_metadata_2025_split huggingface dataset:
Control of 26S proteasome expression by transcription factors regulating multidrug resistance in Saccharomyces cerevisiae
Authors: Owsianik Grzegorz , Balzi Lisabetta , Ghislain Michel 
Published: March 2003

SummaryIn eukaryotic cells, intracellular proteolysis occurs mainly via the ubiquitin–proteasome system. Expression of the yeast proteasome is under the control of the transcription factor, Rpn4p (also known as Son1p/Ufd5p). We show here that the RPN4 gene
promoter contains regulatory sequences that bind Pdr1p and Pdr3p, two homologous
zinc finger‐containing transcription factors, which mediate multiple drug resistance
through the expression of membrane transporter proteins. Mutations in the RPN4
Pdr1p/Pdr3p binding sites lead to decreased expression of the proteasome RPT6
gene and to defective ubiquitin‐mediated proteolysis. Pdr3p, but not Pdr1p, is required
for normal levels of intracellular proteolysis, indicating that the two transcription
factors have distinct functions in the control of RPN4 expression. The RPN4
promoter contains an additional sequence that binds Yap1p, a bZIP‐type transcription
factor that plays an important role in the oxidative stress response and multidrug
resistance. We also show that the Yap1p response element is important in the transactivation
of RPN4 by Yap1p. In yeast cells lacking Pdr1p, ubiquitin‐Pro‐β‐galactosidase,
a short‐lived protein used to assay proteasome activity, is stabilized by the loss
of Yap1p. These data demonstrate that the ubiquitin–proteasome system is controlled
by transcriptional regulators of multidrug resistance via RPN4 expression.
The proper folding of a long C‐terminal segment of the yeast Lys14p regulator is required for activation of LYS genes in response to the metabolic effector
Authors: Alami Mohamed El , Feller André , Piérard André , Dubois Evelyne 
Published: March 2003

SummaryTranscription of lysine genes in Saccharomyces cerevisiae is dependent on

Lys14p and on α‐aminoadipate semialdehyde (αAASA), an intermediate

of the pathway. The two‐thirds C‐terminal end of Lys14p is sufficient to ensure the

activation function of the protein and its modulation by αAASA. Here,

we show that no single discrete domain of Lys14p is able to activate transcription

and that most of the deleted LexA–Lys14p proteins are inactive even in the

presence of a high αAASA concentration. The point mutations abolishing

the activation capacity of Lys14p are distributed all over the entire C‐terminal

segment. Although the deletion of 20 residues rich in leucine and located downstream

of the DNA‐binding domain converts Lys14p to a constitutive transcriptional activator,

our analysis provides evidence that the modulation process of Lys14p activity does

not involve an effector‐dependent masking/unmasking mechanism. Furthermore, we show

that the protein chaperone Hsp82p is required for full activation of LYS genes

by the αAASA‐activated Lys14p as well as by the constitutive Lys14p. Our

results suggest that the proper folding of the two‐thirds C‐terminal portion of Lys14p

is essential not only to activate transcription but also to modulate it according

to αAASA concentration.
C‐terminal domains of Listeria monocytogenes bacteriophage murein hydrolases determine specific recognition and high‐affinity binding to bacterial cell wall carbohydrates
Authors: Loessner Martin J. , Kramer Karl , Ebel Frank , Scherer Siegfried 
Published: March 2003

Summary Listeria monocytogenes phage endolysins Ply118 and Ply500 share a unique enzymatic activity and specifically hydrolyse Listeria cells at the completion of virus multiplication in order to release progeny phage. With the aim of determining the molecular basis for the lytic specificity of these enzymes, we have elucidated their domain structure and examined the function of their unrelated and unique C‐terminal cell wall binding domains (CBDs). Analysis of deletion mutants showed that both domains are needed for lytic activity. Fusions of CBDs with green fluorescent protein (GFP) demonstrated that the C‐terminal 140 amino acids of Ply500 and the C‐terminal 182 residues of Ply118 are necessary and sufficient to direct the murein hydrolases to the bacterial cell wall. CBD500 was able to target GFP to the surface of Listeria cells belonging to serovar groups 4, 5 and 6, resulting in an even staining of the entire cell surface. In contrast, the CBD118 hybrid bound to a ligand predominantly present at septal regions and cell poles, but only on cells of serovars 1/2, 3 and 7. Non‐covalent binding to surface carbohydrate ligands occurred in a rapid, saturation‐dependent manner. We measured 4 × 104 and 8 × 104 binding sites for CBD118 and CBD500 respectively. Surface plasmon resonance analysis revealed unexpected high molecular affinity constants for the CBD–ligand interactions, corresponding to nanomolar affinities. In conclusion, we show that the CBDs are responsible for targeting the phage endolysins to their substrates and function to confer recognition specificity on the proteins. As the CBD sequences contain no repeats and lack all known sequence motifs for anchoring of proteins to the bacterial cell, we conclude that they use unique structural motifs for specific association with the surface of Gram‐positive bacteria.
Generation of Campylobacter jejuni genetic diversity in vivo
Authors: Boer Paulo de , Wagenaar Jaap A. , Achterberg René P. , Putten Jos P. M. van , Schouls Leo M. , Duim Birgitta 
Published: March 2003

SummaryMolecular epidemiology studies suggest that horizontal genetic exchange is a major cause of pathogen biodiversity. We tested this concept for the bacterial enteropathogen Campylobacter jejuni by seeking direct in vivo evidence for the exchange of genetic material among Campylobacter strains. For this purpose, two antibiotic resistance markers were inserted into the hipO or htrA gene of genetically distinct and naturally transformable C. jejuni strains. Genetic exchange of the resistance markers was analysed after co‐cultivation of homologous and heterologous strains in vitro and in vivo during experimental infection of chickens. Double‐resistant recombinants were obtained both in vitro and from the chicken intestine for all combinations of strains tested. Bidirectional genetic exchange of DNA between homologous and heterologous strains was confirmed by Southern blotting in combination with flaA polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP), amplified fragment length polymorphism (AFLP) and pulsed field gel electrophoresis (PFGE). Extensive PFGE analyses of isolated recombinants indicated the frequent occurrence of genetic rearrangements during the experimental infection, in addition to the homologous recombination of the antibiotic resistance genes. Together, the data indicate unequivocally that interstrain genetic exchange as well as intragenomic alterations do occur in vivo during C. jejuni infection. These events probably explain the genome plasticity observed for this pathogen.
Characterization of Burkholderia pseudomallei infection and identification of novel virulence factors using a Caenorhabditis elegans host system
Authors: Gan Yunn‐Hwen , Chua Kim Lee , Chua Hui Hoon , Liu Boping , Hii Chung Shii , Chong Hwee Ling , Tan Patrick 
Published: March 2003

SummaryThe environmental saphrophyte Burkholderia pseudomallei is the causative
agent of melioidosis, a systemic, potentially life‐threatening condition endemic
to many parts of south‐east Asia and northern Australia. We have used the soil nematode
Caenorhabditis elegans as a model host to characterize the mechanisms by which
this bacterium mounts a successful infection. We find that C. elegans is susceptible
to a broad range of Burkholderia species, and that the virulence mechanisms
used by this pathogen to kill nematodes may be similar to those used to infect mammals.
We also find that the specific dynamics of the C. elegans–B. pseudomallei
host–pathogen interaction can be highly influenced by environmental factors,
and that nematode killing results at least in part from the presence of a diffusible
toxin. Finally, by screening for bacterial mutants attenuated in their ability to
kill C. elegans, we genetically identify several new potential virulence factors
in B. pseudomallei. The use of C. elegans as a model host should greatly
facilitate future investigations into how B. pseudomallei can interact with host organisms.
The datA locus predominantly contributes to the initiator titration mechanism in the control of replication initiation in Escherichia coli
Authors: Ogawa Tohru , Yamada Yoshitaka , Kuroda Takao , Kishi Tetsuya , Moriya Shigeki 
Published: March 2003

SummaryReplication of the Escherichia coli chromosome is initiated synchronously
from all origins (oriC) present in a cell at a fixed time in the cell cycle
under given steady state culture conditions. A mechanism to ensure the cyclic initiation
events operates through the chromosomal site, datA, which titrates exceptionally
large amounts of the bacterial initiator protein, DnaA, to prevent overinitiation.
Deletion of the datA locus results in extra initiations and altered temporal
control of replication. There are many other sites on the E. coli chromosome
that can bind DnaA protein, but the contribution of these sites to the control of
replication initiation has not been investigated. In the present study, seven major
DnaA binding sites other than datA have been examined for their influence
on the timing of replication initiation. Disruption of these seven major binding
sites, either individually or together, had no effect on the timing of initiation
of replication. Thus, datA seems to be a unique site that adjusts the balance
between free and bound DnaA to ensure that there is only a single initiation event
in each bacterial cell cycle. Mutation either in the second or the third DnaA box
(a 9 basepair DnaA‐binding sequence) in datA was enough to induce asynchronous
and extra initiations of replication to a similar extent as that observed with the
datA‐deleted strain. These DnaA boxes may act as cores for the cooperative
binding of DnaA to the entire datA region.
LitR, a new transcriptional activator in Vibrio fischeri, regulates luminescence and symbiotic light organ colonization
Authors: Fidopiastis Pat M. , Miyamoto Carol M. , Jobling Michael G. , Meighen Edward A. , Ruby Edward G. 
Published: March 2003

Summary Vibrio fischeri
is the bacterial symbiont within the light‐emitting organ of the sepiolid squid
Euprymna scolopes
. Upon colonizing juvenile squids, bacterial symbionts grow on host‐supplied nutrients, while providing a bioluminescence that the host uses during its nocturnal activities. Mutant bacterial strains that are unable to emit light have been shown to be defective in normal colonization. A 606 bp open reading frame was cloned from
V. fischeri
that encoded a protein, which we named LitR, that had about 60% identity to four related regulator proteins:
Vibrio cholerae
HapR,
Vibrio harveyi
LuxR,
Vibrio parahaemolyticus
OpaR and
Vibrio vulnificus
SmcR. When grown in culture, cells of
V. fischeri
strain PMF8, in which
litR
was insertionally inactivated, were delayed in the onset of luminescence induction and emitted only about 20% as much light per cell as its parent. Protein‐binding studies suggested that LitR enhances quorum sensing by regulating the transcription of the
luxR
gene. Interestingly, when competed against its parent in mixed inocula, PMF8 became the predominant symbiont present in 83% of light organs. Thus, the
litR mutation appears to represent a novel class of mutations in which the loss of a regulatory gene function enhances the bacterium's competence in initiating a benign infection.
The role of heterologous receptors in McpB‐mediated signalling in Bacillus subtilis chemotaxis
Authors: Zimmer Michael A. , Szurmant Hendrik , Saulmon Michael M. , Collins Marissa A. , Bant Jason S. , Ordal George W. 
Published: March 2003

SummaryAsparagine chemotaxis in Bacillus subtilis appears to involve two partially redundant adaptation mechanisms: a receptor methylation‐independent process that operates at low attractant concentrations and a receptor methylation‐dependent process that is required for optimal responses to high concentrations. In order to elucidate these processes, chemotactic responses were assessed for strains expressing methylation‐defective mutations in the asparagine receptor, McpB, in which all 10 putative receptors (10del), five receptors (5del) or only the native copy of mcpB were deleted. This was done in both the presence and the absence of the methylesterase CheB. We found that: (i) only responses to high concentrations of asparagine were impaired; (ii) the presence of all heterologous receptors fully compensated for this defect, whereas responses progressively worsened as more receptors were taken away; (iii) methyl‐group turnover occurred on heterologous receptors after the addition of asparagine, and these methylation changes were required for the restoration of normal swimming behaviour; (iv) in the absence of the methyleste‐rase, the presence of heterologous receptors in some cases caused impaired chemotaxis; and (v) either a certain threshold number of receptors must be present to promote basal CheA activity, or one or more of the receptors missing in the 10del background (but present in the 5del background) is required for establishing basal CheA activity. Taken together, these findings suggest that many or all chemoreceptors work as an ensemble that constitutes a robust chemotaxis system. We propose that the ability of non‐McpB receptors to compensate for the methylation‐defective McpB mutations involves lateral transmission of the adapted conformational change across the ensemble.
Proteoglycans mediate malaria sporozoite targeting to the liver
Authors: Pradel Gabriele , Garapaty Shivani , Frevert Ute 
Published: March 2003

SummaryMalaria sporozoites are rapidly targeted to the liver where they pass through Kupffer cells and infect hepatocytes, their initial site of replication in the mammalian host. We show that sporozoites, as well as their major surface proteins, the CS protein and TRAP, recognize distinct cell type‐specific surface proteoglycans from primary Kupffer cells, hepatocytes and stellate cells, but not from sinusoidal endothelia. RecombinantPlasmodium falciparumCS protein and TRAP bind to heparan sulphate on hepatocytes and both heparan and chondroitin sulphate proteoglycans on stellate cells. On Kupffer cells, CS protein predominantly recognizes chondroitin sulphate, whereas TRAP binding is glycosaminoglycan independent.Plasmodium bergheisporozoites attach to heparan sulphate on hepatocytes and stellate cells, whereas Kupffer cell recognition involves both chondroitin sulphate and heparan sulphate proteoglycans. CS protein also interacts with secreted proteoglycans from stellate cells, the major producers of extracellular matrix in the liver.In situbinding studies using frozen liver sections indicate that the majority of the CS protein binding sites are associated with these matrix proteoglycans. Our data sug‐gest that sporozoites are first arrested in the sinusoid by binding to extracellular matrix proteoglycans and then recognize proteoglycans on the surface of Kupffer cells, which they use to traverse the sinusoidal cell barrier.
The cellular level of the recognition factor RssB is rate‐limiting for σS proteolysis: implications for RssB regulation and signal transduction in σS turnover in Escherichia coli
Authors: Pruteanu Mihaela , Hengge‐Aronis Regine 
Published: March 2003

SummaryDegradation of the general stress sigma factor σS of Escherichia coli is a prime example of regulated proteolysis in prokaryotes. Whereas exponentially growing cells rapidly degrade σS, various stress conditions result in stabilization and, therefore, rapid accumulation of σS. Proteolysis of σS requires the response regulator RssB, a direct recognition factor with phosphorylation‐dependent affinity for σS, which targets σS to the ClpXP protease. Here, we demonstrate that a sudden increase in σS synthesis results in σS stabilization, indicating titration of an essential proteolytic component. Evidence is provided that RssB is the overall rate‐limiting factor for σS proteolysis. As a consequence, the cell has to continuously adjust the expression of RssB to σS in order to maintain σS proteolysis in growing cells, despite variations in the rate of σS synthesis. Such homeostatic feedback‐coupling is provided by rssB transcription being dependent on the σS‐controlled rssAB operon promoter. However, strong and rapid increases in σS synthesis, in re‐sponse to acute stress, exceed the compensatory potential of this feedback loop with the result that σS is stabilized because of RssB titration. We propose that RssB control of σS proteolysis functions as a genetic switch, in which (i) the ‘off’ state (low σS levels caused by proteolysis) is stabilized by a homeostatic negative feedback, and (ii) the threshold for switching to the ‘on’ state (high levels of stable σS) is dependent on the cellular level of active, i.e. phosphorylated RssB.
Thinking and decision making, bacterial style: Bacterial Neural Networks, Obernai, France, 7th–12th June 2002
Authors: Armitage Judith P. , Dorman Charles J. , Hellingwerf Klaas , Schmitt Rudiger , Summers David , Holland Barry 
Published: March 2003

SummaryBacteria exhibit a bewildering range of behavioural responses and permutations of metabolic pathways for maximum exploitation of their environment. These are based on sensory perception of external and internal signals through batteries of surface and cytoplasmic receptors, evaluation of complex information flows and rapid decision making. Appreciation of the diversity of bacterial behaviour and adaptation capacities requires the study of a broad range of organisms and at this meeting we sampled more than 30 species with new findings which included the nature of gaseous receptors, advances in chemotaxis, subversion of host defences by pathogens, adaptation to high salt, community life and its obvious benefits, cell to cell communications and even the nature of bacterial circadian rhythms. With around 80 bacterial genomes now completed, and many more almost there, it was appropriate to complete the meeting with an introduction to Systems Biology and prospects for simulating the virtual cell. The versatility and seemingly ‘intelligent’ behaviour of bacteria will continue to fascinate, and this meeting on Bacterial Neural Networks fully reflected the excitement of this field.
A screen in Saccharomyces cerevisiae identified CaMCM1, an essential gene in Candida albicans crucial for morphogenesis
Authors: Rottmann M. , Dieter S. , Brunner H. , Rupp S. 
Published: March 2003

SummaryMorphogenesis in Saccharomyces cerevisiae and the pathogenic yeast Candida albicans is governed in part by the same molecular circuits. In S. cerevisiae, FLO11/MUC1 expression has been shown to be modulated by multiple signalling pathways required for pseudohyphal development. We have established a screen in S. cerevisiae to identify regulators of fungal development in C. albicans based on FLO11::lacZ expression as a reporter. This screen identified both known components of the mitogen‐activated protein kinase (MAPK) cascade and the cAMP cascade that are important for hyphal development in C. albicans, as well as genes not yet known to be involved in morphogenesis. The Candida homologue of MCM1 is one of the novel factors identified in this screen as being important for morphogenesis. CaMcm1p levels do not vary significantly in different cell types and respond to an autoregulatory feedback mechanism, arguing that CaMcm1p activity is regulated by post‐translational modifications. Both overexpression and repression of this essential gene led to the induction of hyphae. Moreover, we found that the expression of HWP1, a hyphae‐specific gene, was induced by repression of CaMCM1. The changes in morphology and HWP1 expression were not the result of a change in expression levels of NRG1 or TUP1, known repressors of hyphal development. Thus, CaMcm1p is a component of a hitherto unknown regulatory mechanism of hyphal growth.
Nucleotide sequence, functional characterization and evolution of pFKN, a virulence plasmid in Pseudomonas syringae pathovar maculicola
Authors: Rohmer Laurence , Kjemtrup Susanne , Marchesini Patrizia , Dangl Jeffery L. 
Published: March 2003

Summary Pseudomonas syringae pv. maculicola strain M6 (Psm M6) carries the avrRpm1 gene, encoding a type III effector, on a 40 kb plasmid, pFKN. We hypothesized that this plasmid might carry additional genes required for pathogenesis on plants. We report the sequence and features of pFKN. In addition to avrRpm1, pFKN carries an allele of another type III effector, termed avrPphE, and a gene of unknown function (ORF8), expression of which is induced in planta, suggesting a role in the plant–pathogen interaction. The region of pFKN carrying avrRpm1, avrPphE and ORF8 exhibits several features of pathogenicity islands (PAIs). Curing of pFKN (creating Psm M6C) caused a significant reduction in virulence on Arabidopsis leaves. However, complementation studies using Psm M6C demonstrated an obvious virulence function only for avrRpm1. pFKN can integrate and excise from the chromosome of Psm M6 at low frequency via homologous recombination between identical sequence segments located on the chromosome and on pFKN. These segments are part of two nearly identical transposons carrying avrPphE. The avrPphE transposon was also detected in other strains of P. s. pv. maculicola and in P. s. tomato strain DC3000. The avrPphE transposon was found inserted at different loci in different strains. The analysis of sequences surrounding the avrPphE transposon insertion site in the chromosome of Psm M6 indicates that pFKN integrates into a PAI that encodes type III effectors. The integration of pFKN into this chromosomal region may therefore be seen as an evolutionary process determining the formation of a new PAI in the chromosome of Psm M6.
Tocolytic therapy for preterm labour: assessing its potential for reducing preterm delivery
Authors: West Suzanne L. , Yawn Barbara P. , Thorp John M. , Korhonen Maarit J. H. , Savitz David A. , Guess Harry A. 
Published: March 2003

SummaryPreterm labour (PTL) is a major contributor to preterm delivery (PTD) but delivery is often not preventable by current therapies. We conducted this study to determine the proportion of women with PTL who were and who were not candidates for tocolytic therapy. The cohort comprised residents of Olmsted County, Minnesota who delivered at >20 weeks’ gestation in 1985–94 and who experienced PTL. Medical records were abstracted to identify episodes of PTL, its treatment and outcome. We developed an algorithm that accounted for gestation at delivery and pregnancy complications to determine the proportion of pregnancies complicated by PTL that were candidates for tocolytic therapy. Of 651 pregnancies complicated by PTL, a 50% probability sample, stratified by delivery year, were selected and abstracted. The cumulative incidence of PTL ranged from 3.6 to 6.4 per 100 deliveries of live or stillborn infants. Tocolysis was not contraindicated for 49.4% of all women with PTL and for a third of women with only one PTL episode. Delivery was delayed to >35 weeks in 53.8% of candidates for tocolysis. Only an additional 11.7% of women with one or more PTL episodes could have had their PTD delayed beyond 35 weeks if a perfect tocolytic therapy had been available. Many pregnancies complicated by PTL occurred at ≥35 weeks or involved maternal or obstetric factors that contraindicated tocolytic medications. The maximum incremental benefit that could be expected of a new safe and efficacious tocolytic therapy would be to reduce current PTD rates resulting from PTL by about 12%.