bluuebunny/crossref_metadata_2025 huggingface dataset:
Inhibition of Dendritic Cell Maturation by Malaria Is Dose Dependent and Does Not RequirePlasmodium falciparumErythrocyte Membrane Protein 1
Authors: Elliott Salenna R. , Spurck Timothy P. , Dodin Joelle M. , Maier Alexander G. , Voss Till S. , Yosaatmadja Francisca , Payne Paul D. , McFadden Geoffrey I. , Cowman Alan F. , Rogerson Stephen J. , Schofield Louis , Brown Graham V.
Published: May 2007

Red blood cells infected withPlasmodium falciparum(iRBCs) have been shown to modulate maturation of human monocyte-derived dendritic cells (DCs), interfering with their ability to activate T cells. Interaction betweenPlasmodium falciparumerythrocyte membrane protein 1 (PfEMP1) and CD36 expressed by DCs is the proposed mechanism, but we show here that DC modulation does not require CD36 binding, PfEMP1, or contact between DCs and infected RBCs and depends on the iRBC dose. iRBCs expressing a PfEMP1 variant that binds chondroitin sulfate A (CSA) but not CD36 were phagocytosed, inhibited lipopolysaccharide (LPS)-induced phenotypic maturation and cytokine secretion, and abrogated the ability of DCs to stimulate allogeneic T-cell proliferation. CD36- and CSA-binding iRBCs showed comparable inhibition.P. falciparumlines rendered deficient in PfEMP1 expression by targeted gene knockout or knockdown also inhibited LPS-induced phenotypic maturation, and separation of DCs and iRBCs in transwells showed that inhibition was not contact dependent. Inhibition was observed at an iRBC:DC ratio of 100:1 but not at a ratio of 10:1. High doses of iRBCs were associated with apoptosis of DCs, which was not activation induced. Lower doses of iRBCs stimulated DC maturation sufficient to activate autologous T-cell proliferation. In conclusion, modulation of DC maturation byP. falciparumis dose dependent and does not require interaction between PfEMP1 and CD36. Inhibition and apoptosis of DCs by high-dose iRBCs may or may not be physiological. However, our observation that low-dose iRBCs initiate functional DC maturation warrants reevaluation and further investigation of DC interactions with blood-stageP. falciparum.
Marginal Zone B-Cell Depletion Impairs Murine Host Defense againstBorrelia burgdorferiInfection
Authors: Belperron Alexia A. , Dailey Catherine M. , Booth Carmen J. , Bockenstedt Linda K.
Published: May 2007

Marginal zone B (MZB) cells are a B-cell subset that produces T-cell-independent antibodies to blood-borne antigens. In this study, we examined the effects of MZB cell depletion on the immune response to the Lyme disease spirocheteBorrelia burgdorferi, an extracellular pathogen for which T-cell-independent antibody is an important host defense. MZB cell depletion of C3H/HeJ mice using monoclonal antibody to LFA-1 and α4β1integrins reducedB. burgdorferi-specific immunoglobulin M (IgM) titers, enhanced pathogen burden, and led to more severe arthritis assessed within the first 2 weeks of infection. In addition, MZB cell-depleted mice had reduced levels ofB. burgdorferi-specific IgG, which correlated with diminished splenic CD4+T-cell-activation, proliferation, and cytokine production. Passive transfer of immune mouse serum from infected control mice into infected MZB cell-depleted mice reduced pathogen burden but did not alter the expression of T-cell activation markers on splenic CD4+T cells. These findings demonstrate that MZB cells not only are a source of pathogen-specific IgM important for limiting spirochete burden and pathology but also play a prominent role in the priming of splenic T-cell responses to a blood-borne pathogen.
Characterization of aPseudomonas putidaRough Variant Evolved in a Mixed-Species Biofilm withAcinetobactersp. Strain C6
Authors: Hansen Susse Kirkelund , Haagensen Janus A. J. , Gjermansen Morten , Jørgensen Thomas Martini , Tolker-Nielsen Tim , Molin Søren
Published: April 2007

Genetic differentiation by natural selection is readily observed among microbial populations, but a more comprehensive understanding of evolutionary forces, genetic causes, and resulting phenotypic advantages is not often sought. Recently, a surface population ofPseudomonas putidabacteria was shown to evolve rapidly by natural selection of better-adapted variants in a mixed-species biofilm consortium (S. K. Hansen, P. B. Rainey, J. A. Haagensen, and S. Molin, Nature 445:533-536, 2007). Adaptation was caused by mutations in awapHhomolog (PP4943) involved in core lipopolysaccharide biosynthesis. Here we investigate further the biofilm physiology and the phenotypic characteristics of the selectedP. putidarough colony variants. The coexistence of theP. putidapopulation in a mixed-species biofilm withAcinetobactersp. strain C6 is dependent on the benzoate excreted fromAcinetobacterduring the catabolism of benzyl alcohol, the sole carbon source. Examination of biofilm development and the dynamics of the wild-type consortium revealed that the biofilm environment became oxygen limited, possibly with low oxygen concentrations aroundAcinetobactermicrocolonies. In contrast toP. putidawild-type cells, which readily dispersed from the mixed-species biofilm in response to oxygen starvation, the rough variant cells displayed a nondispersal phenotype. However, in monospecies biofilms proliferating on benzoate, the rough variant (like the wild-type population) dispersed in response to oxygen starvation. A key factor explaining this conditional, nondispersal phenotype is likely to be the acquired ability of the rough variant to coaggregate specifically withAcinetobactercells. We further show that theP. putidarough variant displayed enhanced production of a cellulose-like polymer as a consequence of the mutation inwapH. The resulting phenotypic characteristics of theP. putidarough variant explain its enhanced fitness and ability to form tight structural associations withAcinetobactermicrocolonies.
Roles of
rapH
and
rapG
in Positive Regulation of Rapamycin Biosynthesis in
Streptomyces hygroscopicus
Authors: Kuščer Enej , Coates Nigel , Challis Iain , Gregory Matt , Wilkinson Barrie , Sheridan Rose , Petković Hrvoje
Published: April 2007

Rapamycin is an important macrocyclic polyketide produced by
Streptomyces hygroscopicus
and showing immunosuppressive, antifungal, and antitumor activities as well as displaying anti-inflammatory and neuroregenerative properties. The immense pharmacological potential of rapamycin has led to the production of an array of analogues, including through genetic engineering of the rapamycin biosynthetic gene cluster. This cluster contains several putative regulatory genes. Based on DNA sequence analysis, the products of genes
rapH
and
rapG
showed high similarities with two different families of transcriptional activators, LAL and AraC, respectively. Overexpression of either gene resulted in a substantial increase in rapamycin biosynthesis, confirming their positive regulatory role, while deletion of both from the chromosome of
S. hygroscopicus
resulted in a complete loss of antibiotic production. Complementation studies indicated an essential role of the RapG regulator for rapamycin biosynthesis and a supportive role of RapH. A direct effect of
rapH
and
rapG
gene products on the promoter of the rapamycin polyketide synthase operon,
rapA
-
rapB
, was observed using the chalcone synthase gene
rppA
as a reporter system.
A Novel Cell Surface-Anchored Cellulose-Binding Protein Encoded by the
sca
Gene Cluster of
Ruminococcus flavefaciens
Authors: Rincon Marco T. , Cepeljnik Tadej , Martin Jennifer C. , Barak Yoav , Lamed Raphael , Bayer Edward A. , Flint Harry J.
Published: April 2007

Ruminococcus flavefaciens
produces a cellulosomal enzyme complex, based on the structural proteins ScaA, -B, and -C, that was recently shown to attach to the bacterial cell surface via the wall-anchored protein ScaE. ScaA, -B, -C, and -E are all cohesin-bearing proteins encoded by linked genes in the
sca
cluster. The product of an unknown open reading frame within the
sca
cluster, herein designated CttA, is similar in sequence at its C terminus to the corresponding region of ScaB, which contains an X module together with a dockerin sequence. The ScaB-XDoc dyad was shown previously to interact tenaciously with the cohesin of ScaE. Likewise, avid binding was confirmed between purified recombinant fragments of the CttA-XDoc dyad and the ScaE cohesin. In addition, the N-terminal regions of CttA were shown to bind to cellulose, thus suggesting that CttA is a cell wall-anchored, cellulose-binding protein. Proteomic analysis showed that the native CttA protein (∼130 kDa) corresponds to one of the three most abundant polypeptides binding tightly to insoluble cellulose in cellulose-grown
R. flavefaciens
17 cultures. Interestingly, this protein was also detected among cellulose-bound proteins in the related strain
R. flavefaciens
007C but not in a mutant derivative, 007S, that was previously shown to have lost the ability to grow on dewaxed cotton fibers. In
R. flavefaciens
, the presence of CttA on the cell surface is likely to provide an important mechanism for substrate binding, perhaps compensating for the absence of an identified cellulose-binding module in the major cellulosomal scaffolding proteins of this species.
Bacillus subtilisPellicle Formation Proceeds through Genetically Defined Morphological Changes
Authors: Kobayashi Kazuo
Published: April 2007

Biofilms are structured multicellular communities of bacteria that form through a developmental process. In standing culture, undomesticated strains ofBacillus subtilisproduce a floating biofilm, called a pellicle, with a distinct macroscopic architecture. Here we report on a comprehensive analysis ofB. subtilispellicle formation, with a focus on transcriptional regulators and morphological changes. To date, 288 known or putative transcriptional regulators encoded by theB. subtilisgenome have been identified or assigned based on similarity to other known proteins. The genes encoding these regulators were systematically disrupted, and the effects of the mutations on pellicle formation were examined, resulting in the identification of 19 regulators involved in pellicle formation. In addition, morphological analysis revealed that pellicle formation begins with the formation of cell chains, which is followed by clustering and degradation of cell chains. Genetic and morphological evidence showed that each stage of morphological change can be defined genetically, based on mutants of transcriptional regulators, each of which blocks pellicle formation at a specific morphological stage. Formation and degradation of cell chains are controlled by down- and up-regulation of σD- and σH-dependent autolysins expressed at specific stages during pellicle formation. Transcriptional analysis revealed that the transcriptional activation ofsigHdepends on the formation of cell clusters, which in turn activates transcription of σH-dependent autolysin in cell clusters. Taken together, our results reveal relationships between transcriptional regulators and morphological development during pellicle formation byB. subtilis.
Identification of the Missing
trans
-Acting Enoyl Reductase Required for Phthiocerol Dimycocerosate and Phenolglycolipid Biosynthesis in
Mycobacterium tuberculosis
Authors: Siméone Roxane , Constant Patricia , Guilhot Christophe , Daffé Mamadou , Chalut Christian
Published: April 2007

Phthiocerol dimycocerosates (DIM) and phenolglycolipids (PGL) are functionally important surface-exposed lipids of
Mycobacterium tuberculosis
. Their biosynthesis involves the products of several genes clustered in a 70-kb region of the
M. tuberculosis
chromosome. Among these products is PpsD, one of the modular type I polyketide synthases responsible for the synthesis of the lipid core common to DIM and PGL. Bioinformatic analyses have suggested that this protein lacks a functional enoyl reductase activity domain required for the synthesis of these lipids. We have identified a gene, Rv2953, that putatively encodes an enoyl reductase. Mutation in Rv2953 prevents conventional DIM formation and leads to the accumulation of a novel DIM-like product. This product is unsaturated between C-4 and C-5 of phthiocerol. Consistently, complementation of the mutant with a functional
pks15/1
gene from
Mycobacterium bovis
BCG resulted in the accumulation of an unsaturated PGL-like substance. When an intact Rv2953 gene was reintroduced into the mutant strain, the phenotype reverted to the wild type. These findings indicate that Rv2953 encodes a
trans
-acting enoyl reductase that acts with PpsD in phthiocerol and phenolphthiocerol biosynthesis.
Intracellular Parasitism of Chlamydiae: Specific Infectivity of Chlamydiaphage Chp2 in
Chlamydophila abortus
Authors: Skilton R. J. , Cutcliffe L. T. , Pickett M. A. , Lambden P. R. , Fane B. A. , Clarke I. N.
Published: April 2007

The obligate intracellular nature of chlamydiae presents challenges to the characterization of its phages, which are potential tools for a genetic transfer system. An assay for phage infectivity is described, and the infectious properties of phage Chp2 were determined.
A Single, Specific Thymine Mutation in the ComK-Binding Site Severely Decreases Binding and Transcription Activation by the Competence Transcription Factor ComK of
Bacillus subtilis
Authors: Susanna Kim A. , Mironczuk Aleksandra M. , Smits Wiep Klaas , Hamoen Leendert W. , Kuipers Oscar P.
Published: April 2007

The competence transcription factor ComK plays a central role in competence development in
Bacillus subtilis
by activating the transcription of the K regulon. ComK-activated genes are characterized by the presence of a specific sequence to which ComK binds, a K-box, in their upstream DNA region. Each K-box consists of two AT-boxes with the consensus sequence AAAA-(N)
5
-TTTT, which are separated by a flexible spacer resulting in either two, three, or four helical turns between the starting nucleotides of the repeating AT-box units. In this study, the effects of potential determinants of ComK regulation in K-boxes were investigated by testing ComK's transcription activation and DNA-binding affinity on altered K-boxes with mutations either in the spacer between the AT-boxes or in the consensus sequence of the AT-boxes. The most striking result demonstrates the importance of the second thymine base in the AT-boxes. Mutation of this T into a guanine resulted in a threefold reduction in transcription activation and DNA binding by ComK. Transcription activation, as well as DNA binding, was almost completely abolished when both AT-boxes contained a T
2
-to-G mutation. This result was corroborated by in silico analyses demonstrating that a combination of mutations at the T
2
positions of both AT-boxes is not found among any ComK-activated K-boxes, indicating that at least one consensus T
2
position is required to maintain a functional K-box. The results suggest an important structural role for T
2
in ComK binding, probably by its specific position in the minor groove of the DNA.
Flavodoxin:Quinone Reductase (FqrB): a Redox Partner of Pyruvate:Ferredoxin Oxidoreductase That Reversibly Couples Pyruvate Oxidation to NADPH Production in
Helicobacter pylori
and
Campylobacter jejuni
Authors: St. Maurice Martin , Cremades Nunilo , Croxen Matthew A. , Sisson Gary , Sancho Javier , Hoffman Paul S.
Published: April 2007

Pyruvate-dependent reduction of NADP has been demonstrated in cell extracts of the human gastric pathogen
Helicobacter pylori.
However, NADP is not a substrate of purified pyruvate:ferredoxin oxidoreductase (PFOR), suggesting that other redox active enzymes mediate this reaction. Here we show that
fqrB
(HP1164), which is essential and highly conserved among the epsilonproteobacteria, exhibits NADPH oxidoreductase activity. FqrB was purified by nickel interaction chromatography following overexpression in
Escherichia coli
. The protein contained flavin adenine dinucleotide and exhibited NADPH quinone reductase activity with menadione or benzoquinone and weak activity with cytochrome
c
, molecular oxygen, and 5,5′-dithio-bis-2-nitrobenzoic acid (DTNB). FqrB exhibited a ping-pong catalytic mechanism, a
k
cat
of 122 s
−1
, and an apparent

K
m

of 14 μM for menadione and 26 μM for NADPH. FqrB also reduced flavodoxin (FldA), the electron carrier of PFOR. In coupled enzyme assays with purified PFOR and FldA, FqrB reduced NADP in a pyruvate- and reduced coenzyme A (CoA)-dependent manner. Moreover, in the presence of NADPH, CO
2
, and acetyl-CoA, the PFOR:FldA:FqrB complex generated pyruvate via CO
2
fixation. PFOR was the rate-limiting enzyme in the complex, and nitazoxanide, a specific inhibitor of PFOR of
H. pylori
and
Campylobacter jejuni
, also inhibited NADP reduction in cell-free lysates. These capnophilic (CO
2
-requiring) organisms contain gaps in pathways of central metabolism that would benefit substantially from pyruvate formation via CO
2
fixation. Thus, FqrB provides a novel function in pyruvate metabolism and, together with production of superoxide anions via quinone reduction under high oxygen tensions, contributes to the unique microaerobic lifestyle that defines the epsilonproteobacterial group.
In Vivo-Selected Mutations in Methyl-Directed Mismatch Repair Suppress the Virulence Attenuation ofSalmonella damMutant Strains following Intraperitoneal, but Not Oral, Infection of Naïve Mice
Authors: Heithoff Douglas M. , Badie Golnaz , Julio Steven M. , Enioutina Elena Y. , Daynes Raymond A. , Sinsheimer Robert L. , Mahan Michael J.
Published: April 2007

Salmonella entericaserovar Typhimurium that lacks the DNA adenine methylase (Dam) ectopically expresses multiple genes that are preferentially expressed during infection, is attenuated for virulence, and confers heightened immunity in vaccinated hosts. The safety ofdammutantSalmonellavaccines was evaluated by screening within infected mice for isolates that have an increased capacity to cause disease relative to the attenuated parental strain. Sincedammutant strains are sensitive to the DNA base analog 2-aminopurine (2-AP), we screened for 2-AP-resistant (2-APr) isolates in systemic tissues of mice infected withdammutantSalmonella. Such 2-APrderivatives were isolated following intraperitoneal but not oral administration and were shown to be competent for infectivity via intraperitoneal but not oral infection of naïve mice. These 2-APrderivatives were deficient in methyl-directed mismatch repair and were resistant to nitric oxide, yet they retained the bile-sensitive phenotype of the parentaldammutant strain. Additionally, introduction of amutHnull mutation intodammutant cells suppressed the inherent defects in intraperitoneal infectivity and nitric oxide resistance, as well as overexpression of SpvB, an actin cytotoxin required forSalmonellasystemic survival. These data suggest that restoration of intraperitoneal virulence ofdammutant strains is associated with deficiencies in methyl-directed mismatch repair that correlate with the production of systemically related virulence functions.
Diffusible Signal Factor-Dependent Cell-Cell Signaling and Virulence in the Nosocomial PathogenStenotrophomonas maltophilia
Authors: Fouhy Yvonne , Scanlon Karl , Schouest Katherine , Spillane Charles , Crossman Lisa , Avison Matthew B. , Ryan Robert P. , Dow J. Maxwell
Published: April 2007

The genome ofStenotrophomonas maltophiliaencodes a cell-cell signaling system that is highly related to the diffusible signal factor (DSF)-dependent system of the phytopathogenXanthomonas campestris. Here we show that inS. maltophilia, DSF signaling controls factors contributing to the virulence and antibiotic resistance of this important nosocomial pathogen.
Genomic SELEX Search for Target Promoters under the Control of the PhoQP-RstBA Signal Relay Cascade
Authors: Ogasawara Hiroshi , Hasegawa Akiko , Kanda Emi , Miki Takenori , Yamamoto Kaneyoshi , Ishihama Akira
Published: April 2007

RstBA, a two-component regulatory system ofEscherichia coliwith an unidentified regulatory function, is under the control of a Mg2+-sensing PhoQP two-component system. In order to identify the network of transcription regulation downstream of RstBA, we isolated a set of RstA-binding sequences from theE. coligenome by using the genomic SELEX system. A gel mobility shift assay indicated the binding of RstA to two SELEX DNA fragments, one including the promoter region ofasr(acid shock RNA) and another including the promoter forcsgD(a regulator of the curli operon). Using a DNase I footprinting assay, we determined the RstA-binding sites (RstA boxes) with the consensus sequence TACATNTNGTTACA. Transcription of theasrgene was induced 10- to 60-fold either in low-pH (pH 4.5) LB medium or in low-phosphate minimal medium as detected by promoter assay. The acid-induced in vivo transcription ofasrwas reduced after the deletion ofrstA. In vivo transcription of theasrpromoter was observed only in the presence of RstA. In agreement with the PhoQP-RstBA network, the addition of Mg2+led to a severe reduction of theasrpromoter activity, and the disruption ofphoPalso reduced theasrpromoter activity, albeit to a lesser extent. These observations altogether indicate that RstA is an activator ofasrtranscription. In contrast, transcription ofcsgDwas repressed by overexpression of RstA, indicating that RstA is a repressor forcsgD. With these data taken together, we conclude that the expression of bothasrandcsgDis under the direct control of the PhoQP-RstBA signal relay cascade.
Clocking Out: Modeling Phage-Induced Lysis of
Escherichia coli
Authors: Ryan Gillian L. , Rutenberg Andrew D.
Published: April 2007

Phage λ lyses the host
Escherichia coli
at a precisely scheduled time after induction. Lysis timing is determined by the action of phage holins, which are small proteins that induce hole formation in the bacterium's cytoplasmic membrane. We present a two-stage nucleation model of lysis timing, with the nucleation of condensed holin rafts on the inner membrane followed by the nucleation of a hole within those rafts. The nucleation of holin rafts accounts for most of the delay of lysis after induction. Our simulations of this model recover the accurate lysis timing seen experimentally and show that the timing accuracy is optimal. An enhanced holin-holin interaction is needed in our model to recover experimental lysis delays after the application of membrane poison, and such early triggering of lysis is possible only after the inner membrane is supersaturated with holin. Antiholin reduces the delay between membrane depolarization and lysis and leads to an earlier time after which triggered lysis is possible.
Specific Bonds between an Iron Oxide Surface and Outer Membrane Cytochromes MtrC and OmcA from
Shewanella oneidensis
MR-1
Authors: Lower Brian H. , Shi Liang , Yongsunthon Ruchirej , Droubay Timothy C. , McCready David E. , Lower Steven K.
Published: April 2007

Shewanella oneidensis
MR-1 is purported to express outer membrane cytochromes (e.g., MtrC and OmcA) that transfer electrons directly to Fe(III) in a mineral during anaerobic respiration. A prerequisite for this type of reaction would be the formation of a stable bond between a cytochrome and an iron oxide surface. Atomic force microscopy (AFM) was used to detect whether a specific bond forms between a hematite (Fe
2
O
3
) thin film, created with oxygen plasma-assisted molecular beam epitaxy, and recombinant MtrC or OmcA molecules coupled to gold substrates. Force spectra displayed a unique force signature indicative of a specific bond between each cytochrome and the hematite surface. The strength of the OmcA-hematite bond was approximately twice that of the MtrC-hematite bond, but direct binding to hematite was twice as favorable for MtrC. Reversible folding/unfolding reactions were observed for mechanically denatured MtrC molecules bound to hematite. The force measurements for the hematite-cytochrome pairs were compared to spectra collected for an iron oxide and
S. oneidensis
under anaerobic conditions. There is a strong correlation between the whole-cell and pure-protein force spectra, suggesting that the unique binding attributes of each cytochrome complement one another and allow both MtrC and OmcA to play a prominent role in the transfer of electrons to Fe(III) in minerals. Finally, by comparing the magnitudes of binding force for the whole-cell versus pure-protein data, we were able to estimate that a single bacterium of
S. oneidensis
(2 by 0.5 μm) expresses ∼10
4
cytochromes on its outer surface.
YbxF, a Protein Associated with Exponential-Phase Ribosomes inBacillus subtilis
Authors: Sojka Luděk , Fučík Vladimír , Krásný Libor , Barvík Ivan , Jonák Jiří
Published: April 2007

TheybxFgene is a member of the streptomycin operon in a wide range of gram-positive bacteria. InBacillus subtilis, it codes for a small basic protein (82 amino acids, pI 9.51) of unknown function. We demonstrate that, inB. subtilis, YbxF localizes to the ribosome, primarily to the 50S subunit, with dependence on growth phase. Based on three-dimensional structures of YbxF generated by homology modeling, we identified helix 2 as important for the interaction with the ribosome. Subsequent mutational analysis of helix 2 revealed Lys24 as crucial for the interaction. Neither theB. subtilis ybxFgene nor its paralogue, theymxCgene, is essential, as shown by probing ΔybxF, ΔymxC, or ΔybxFΔymxCdouble deletion strains in several functional assays.
l
-Sorbose Reductase and Its Transcriptional Regulator Involved in
l
-Sorbose Utilization of
Gluconobacter frateurii
Authors: Soemphol Wichai , Toyama Hirohide , Moonmangmee Duangtip , Adachi Osao , Matsushita Kazunobu
Published: April 2007

Upstream of the gene for flavin adenine dinucleotide (FAD)-dependent
d
-sorbitol dehydrogenase (SLDH),
sldSLC
, a putative transcriptional regulator was found in
Gluconobacter frateurii
THD32 (NBRC 101656). In this study, the whole
sboR
gene and the adjacent gene,
sboA
, were cloned and analyzed.
sboR
mutation did not affect FAD-SLDH activity in the membrane fractions. The SboA enzyme expressed and purified from an
Escherichia coli
transformant showed NADPH-dependent
l
-sorbose reductase (NADPH-SR) activity, and the enzyme was different from the NADPH-SR previously reported for
Gluconobacter suboxydans
IFO 3291 in molecular size and amino acid sequence. A mutant defective in
sboA
showed significantly reduced growth on
l
-sorbose, indicating that the SboA enzyme is required for efficient growth on
l
-sorbose. The
sboR
mutant grew on
l
-sorbose even better than the wild-type strain did, and higher NADPH-SR activity was detected in cytoplasm fractions. Reverse transcription-PCR experiments indicated that
sboRA
comprises an operon. These data suggest that
sboR
is involved in the repression of
sboA
, but not in the induction of
sldSLC
, on
d
-sorbitol and that another activator is required for the induction of these genes by
d
-sorbitol or
l
-sorbose.
Genetic Diversity of Toxigenic and Nontoxigenic
Vibrio cholerae
Serogroups O1 and O139 Revealed by Array-Based Comparative Genomic Hybridization
Authors: Pang Bo , Yan Meiying , Cui Zhigang , Ye Xiaofen , Diao Baowei , Ren Yonghong , Gao Shouyi , Zhang Liang , Kan Biao
Published: April 2007

Toxigenic serogroups O1 and O139 of
Vibrio cholerae
may cause cholera epidemics or pandemics. Nontoxigenic strains within these serogroups also exist in the environment, and also some may cause sporadic cases of disease. Herein, we investigate the genomic diversity among toxigenic and nontoxigenic O1 and O139 strains by comparative genomic microarray hybridization with the genome of El Tor strain N16961 as a base. Conservation of the toxigenic O1 El Tor and O139 strains is found as previously reported, whereas accumulation of genome changes was documented in toxigenic El Tor strains isolated within the 40 years of the seventh pandemic. High phylogenetic diversity in nontoxigenic O1 and O139 strains is observed, and most of the genes absent from nontoxigenic strains are clustered together in the N16961 genome. By comparing these toxigenic and nontoxigenic strains, we observed that the small chromosome of
V. cholerae
is quite conservative and stable, outside of the superintegron region. In contrast to the general stability of the genome, the superintegron demonstrates pronounced divergence among toxigenic and nontoxigenic strains. Additionally, sequence variation in virulence-related genes is found in nontoxigenic El Tor strains, and we speculate that these intermediate strains may have pathogenic potential should they acquire CTX prophage alleles and other gene clusters. This genome-wide comparison of toxigenic and nontoxigenic
V. cholerae
strains may promote understanding of clonal differentiation of
V. cholerae
and contribute to an understanding of the origins and clonal selection of epidemic strains.
Clinical relevance of molecular genetics to paediatric sarcomas
Authors: Slater Olga , Shipley Janet
Published: April 2007

The application of cytogenetic and molecular genetic analyses to paediatric sarcomas has identified a number of characteristic changes associated with types and subtypes of sarcomas. This has led to increased understanding of the underlying molecular biology of some sarcomas and provided an important adjunct to standard morphological and immunohistochemical diagnoses. Characteristic genetic abnormalities, particularly specific chromosome translocations and associated fusion genes, have diagnostic and in some cases prognostic value. There is also the potential to detect micrometastastic disease. Fusion genes are most readily detected by fluorescence in situ hybridisation and reverse transcription-PCR technologies. The expression profiles of tumours with specific fusion genes are characteristically similar and the molecular signatures of sarcomas are also proving to be of diagnostic and prognostic value. Furthermore, fusion genes and other emerging molecular events associated with sarcomas represent potential targets for novel therapeutic approaches which are desperately required to improve the outcome of children with certain categories of sarcoma, including rhabdomyosarcomas and the Ewing’s family of tumours. Increased understanding of the molecular biology of sarcomas is leading towards more effective treatments which may complement or be less toxic than conventional radiotherapy and cytotoxic chemotherapy. Here we review paediatric sarcomas that have associated molecular genetic changes which can increase diagnostic and prognostic accuracy and impact on clinical management.
Expansion of a mutated clone: from stem cell to tumour
Authors: Leedham S J , Wright N A
Published: April 2007

Intestinal stem cells are adult, tissue-based stem cells located at the base of the intestinal crypt and are capable of regenerating all intestinal cell types. The progeny of mutated stem cells can expand to fill an entire crypt as a consequence of genetic drift, selective advantage or hitchhiking—eventually forming a clonal crypt population by a process called “niche succession”. Cancer is believed to be a disease of stem cells. The digestive tract has a very high cancer prevalence partly due to rapid epithelial cell turnover and exposure to dietary toxins. Work on the hereditary cancer syndromes, including familial adenomatous polyposis (FAP), has led to significant advances, including the adenoma–carcinoma sequence. The initial mutation involved in this stepwise progression is in the “gatekeeper” tumour suppressor geneadenomatous polyposis coli (APC). In FAP somatic, second hits in this gene are non-random events, selected for by the position of the germline mutation. The early growth of adenomas is contentious, with two main theories, the “top-down” and “bottom-up” hypotheses, attempting to explain the spread of dysplastic tissue in the bowel. Initial X chromosome inactivation studies suggested that colorectal tumours were monoclonal; however, work on a rare XO/XY human patient with FAP and chimeric Min mice showed that approximately 76% of adenomas were polyclonal. A reduction in tumour multiplicity in the chimeric mouse model has been achieved by the introduction of a homozygous tumour resistance allele. This model has been used to suggest that short-range interaction between adjacent initiated crypts, not random polyp collision, is responsible for tumour polyclonality.
Expression of ZNF652, a novel zinc finger protein, in vulvar carcinomas and its relation to prognosis
Authors: Holm R , Knopp S , Kumar R , Lee J , Nesland J M , Tropè C , Callen D F
Published: April 2007

Aims:

To determine the levels of expression of ZNF652 and its relevance to prognosis in vulvar squamous cell carcinomas.

Methods:

22 cases of vulvar intraepithelial neoplasia (VIN) and tumours from 217 patients with vulvar squamous cell carcinomas were investigated for expression of ZNF652 using immunostaining methods. The effect of ZNF652 ectopic expression was determined in the vulvar carcinoma cell line SW954 by western and cell-based assays.

Results:

High levels of ZNF652 nuclear expression were observed in 5 (100%) of VIN I, 6 (75%) of VIN II and 109 (50.2%) of the vulvar carcinomas, whereas low levels were seen in 2 (25%) VIN II, 9 (100%) of VIN III and 108 (49.8%) of the vulvar carcinomas. High levels of ZNF652 expression in the vulvar carcinomas were significantly correlated to high expression of EphA2. However, when correcting for multiple testing this correlation was lost. No association was identified between ZNF652 expression and p16, p21, p27, p53, cyclin A, D1, D3, E, EphrinA-1 and human papillomavirus. Variations in levels of ZNF652 were not related to prognosis. Low levels of ZNF652 protein were identified in the vulvar carcinoma cell line SW954. Furthermore, SW954 cells ectopically expressing ZNF652 showed reduced cell proliferation and the ability to form colonies on plastic.

Conclusions:

ZNF652 protein expression is reduced in 25% of VIN II, 100% of VIN III and approximately 50% of the cases of vulvar squamous cell carcinoma, and may be an early event in the pathogenesis of vulvar squamous cell carcinoma. Variations in the levels of ZNF652 were not related to patient’s prognosis.
Effects of fixation on RNA integrity in a liquid-based cervical cytology setting
Authors: Horvath C , Boulet G , Sahebali S , Bogers J , Depuydt C , Vereecken A , Vermeulen T , Vanden Broeck D
Published: April 2007

Aims:

Despite many improvements, cervical cancer screening is still subject to shortcomings. Diagnostic accuracy may improve by using molecular biological techniques, requiring RNA of superior quality. This study determined the effect of SurePath fixation on RNA integrity to assess the suitability of clinical samples collected in this medium for RNA-based molecular assays.

Methods:

RNA isolation was performed on fresh and fixed HeLa cells and exfoliated cervical cells fixed in SurePath. The RNA integrity was evaluated by analysis of ribosomal RNA as an indicator of quality. The effect of SurePath preservation on PCR amplification was evaluated by real-time reverse transcriptase (RT)-PCR.

Results:

In contrast to unfixed cells, SurePath-fixed cells yielded less and severely degraded RNA, as shown by the absence of ribosomal RNA bands. RNA derived from SurePath-fixed cells showed poor real-time RT-PCR amplification characteristics, as evidenced by the absent correlation between threshold values and log cDNA concentration.

Conclusions:

Implementation of molecular biology in a clinical context is on the rise and may alleviate shortcomings in current screening and diagnostics. This study shows that SurePath fixation gives rise to highly fragmented RNA with insufficient quality for further reliable analysis by standard real-time RT-PCR applications. The increasing prominence of molecular screening stresses the importance of this finding, which must be considered in relation to choice of an appropriate liquid-based cytology system.
Axillary sentinel lymph node micrometastases with extracapsular extension: a distinct pattern of breast cancer metastasis?
Authors: Cserni G
Published: April 2007

Aims:

To examine the frequency of extracapsular extension (ECE) of sentinel lymph node (SLN) metastases in breast cancer according to metastasis size, and to characterise ECE in micometastases.

Methods:

If initially negative, SLNs were examined by step-sectioning and immunohistochemistry. Non-SLNs were not subjected to enhanced pathology. Positive axillary SLNs were analysed for metastasis size and the presence of ECE.

Results:

Of 885 successful SLN biopsy cases, 343 (39%) exhibited SLN involvement, and 115 (34%) displayed ECE. Of the latter, 107 underwent axillary dissection, and 63 (59%) of these demonstrated non-SLN metastases. The presence of ECE correlated with metastasis size (coefficient 0.92). Eight (10%) of the 84 micrometastatic SLN cases had ECE, and two of these were associated with non-SLN metastases. Only ECE and only the intraparenchymal nodal part of the micrometastasis were seen in some sections of five cases each. The primary tumours of the micrometastatic cases with ECE were non-high-grade and often of tubular type.

Conclusions:

The frequency of ECE increases with increasing nodal metastasis size. Minimal nodal metastases with ECE may represent a distinct pattern of nodal involvement with a predominant capsular and extracapsular, but only minimal or no nodal parenchymal component, predominantly seen in non-poorly differentiated and/or tubular carcinomas. This presentation of nodal metastasis can sometimes pose differential diagnostic problems, and should be distinguished from massive metastases presenting with ECE because it does not seem to be so commonly associated with non-SLN metastases or a massive metastatic load to the axilla as ECE of SLN metastases in general.
Parafibromin expression in breast cancer: a novel marker for prognostication?
Authors: Selvarajan S , Sii L-H , Lee A , Yip G , Bay B-H , Tan M-H , Teh B-T , Tan P-H
Published: April 2007

Background:

Parafibromin is a novel protein product ofHRPT2, a recently identified tumour suppressor gene. Mutations of theHRPT2gene are common in parathyroid carcinomas, and these exhibit reduced protein expression. Parafibromin expression in breast cancer has not been previously studied.

Aims:

To determine the distribution of parafibromin in breast cancer tissues, and correlate its expression with conventional pathological parameters.

Methods:

Tissue microarrays were constructed from archival paraffin embedded breast cancer samples. Sections cut from tissue microarray blocks were subjected to immunohistochemistry. Immunopositivity for parafibromin and intensity-percentage scores were derived by blinded evaluation. Findings were correlated with clinicopathological parameters.

Results:

163 breast cancers were assessed. Larger tumours were less likely to express parafibromin than smaller ones, with the association approaching statistical significance (p = 0.05). Staining intensity correlated inversely with tumour size (p = 0.016) and pathological stage (p = 0.008); as did parafibromin intensity-percentage score with pathological stage (p = 0.03), lymphovascular invasion (p = 0.03) and cerbB2 intensity-percentage score (p = 0.04).

Conclusion:

Parafibromin in breast cancer, as in parathyroid tumours, appears to have tumour suppressor functions, with loss of protein expression associated with adverse pathological parameters. These findings may indicate a potential role of parafibromin as a prognostic marker in breast cancer.
A Bayesian hierarchical meta-analysis of growth for the genusSebastesin the eastern Pacific Ocean
Authors: Helser Thomas E , Stewart Ian J , Lai Han-Lin
Published: May 2007

We conducted a meta-analysis of growth for 46 species of the genus Sebastes in the eastern Pacific Ocean using a Bayesian hierarchical model to estimate parameters, to investigate growth variability, and to elucidate meaningful biological covariates. Growth in terms of maximum attainable size (L∞) ranged from 12 to 80 cm, and instantaneous growth rates varied by over an order of magnitude (K; 0.03–0.34·year–1). Results from this method also confirm the theoretical, but often untested, view that growth parameters L∞and K are negatively correlated among populations or species of fish; Bayesian credibility intervals for correlation ranged from –0.2 to –0.7, with the posterior median of –0.4. The Bayesian hierarchical growth model showed less variability in growth parameters and lower correlations among parameters than those from standard techniques used in population ecology, suggesting that the absolute value of the correlation between L∞and K may be lower than the general perception in the ecological literature. Exploration of several covariates revealed that asymptotic size varied positively as a function of the size at 50% maturity. Finally, posterior probability distributions of the hyperparameters from this analysis provide plausible informative priors of growth for stock assessments of data-poor species.
Centres de bandes et constantes d'anharmonicité de la molécule du trifluorure de phosphore PF3
Authors: Najib Hamid , Jabar Abderrahim
Published: May 2007

In this paper, we establish interesting relationships linking rovibrational band-centers and anharmonic constants of symmetric top molecule. Using these relationships, nine accurate values of the xij and gij anharmonic constants have been deduced from the band-centers of fundamental, overtone, combination, and hot bands of the PF3 gas molecule. Taking into account the Fermi resonance between the ν3 and ν2 + ν4 bands, with a W234 = 2.86 cm–1 coupling term, the band-centers and the anharmonic constants of the molecular potential have been corrected.
Long-term variations of the oxygen red 630 nm line nightglow intensity
Authors: Gudadze N B , Didebulidze G G , Javakhishvili G Sh. , Shepherd M G , Vardosanidze M V
Published: May 2007

The long-term data set of total nightglow intensity of the oxygen red 630.0~nm line observed at Abastumani (41.8°N, 42.8°E) between 1957–1993 is investigated. The long-term trend and characteristic variations in solar radiation during an 11 year cycle of the red-line intensity are different after astronomical twilight (premidnight) and at midnight. The amplitude of deviation of the red-line intensity from its mean value at solar maximum and (or) minimum phase is greatest after astronomical twilight and decreases toward midnight. The long-term trend of these variations changes from its value about 0.74 R/year premidnight to its minimum negative value of about –1.92 R/year at and after midnight. This behavior of the long-term trend is considered as a possible result of an increase in electron density below the peak height (hmF2) of the ionospheric F2 layer and lowering of the height hmF2 after midnight predicted by the TIME-GCM model on the assumption of an increase in density of greenhouse gases in the lower atmosphere. The third-order regression equation (with different solar activity indices) is considered to be convenient for describing long-term variations in the mean annual red-line intensity.PACS Nos.: 94.10.Rk, 94.20.Ji, 92.60.Vb
Thermal expansion: a fresh look at an old problem
Authors: Mohazzabi Pirooz , Behroozi Feredoon
Published: May 2007

The phenomenon of thermal expansion is discussed from a basic perspective. Using a new approach, the well-known equation that relates the interatomic distance to the mean total energy per atom in a solid is obtained. Unlike the standard derivation, the new treatment is free from the computational inconsistencies and yields the correct expression for thermal expansion at any temperature. The standard derivation of the equation is outlined and its shortcomings are discussed.

PACS Nos.: 65.40.–b, 65.40.De
Transfer of ultracold 87Rb from a QUIC magnetic trap into a far off resonance optical trap
Authors: Ming H , van Wijngaarden W A
Published: May 2007

Ultracold 87Rb atoms were transferred from a QUIC (quadrupole and Ioffe configuration) magnetic trap into a far off resonance optical trap (FORT). FORTs were created by focusing a 150 mW laser beam having a wavelength of 852 nm to a spot having a radius of 20 and 30 µm. A probe laser then passed through the ultracold atom cloud after the magnetic trap was turned off to study the temporal evolution of the optically trapped atoms. Nearly 106 atoms could be transferred into the FORT at temperatures as low as 1 µK with an efficiency as high as 50%.

PACS No.: 32.80.Pj
Collective bands in 80,82Kr
Authors: Mishra S , Tripathy K C , Sahu R
Published: May 2007

Deformed shell-model calculations are performed to study the structure of the collective bands in 80,82Kr. In our microscopic model, the single particle orbits 1p3/2, 0f5/2, 1p1/2, and 0g9/2 constitute the configuration space with 56Ni as the inert core. A modified Kuo interaction for this basis space is used in our calculation. The different levels are classified into collective bands on the basis of the B(E2) values among them. The calculated ground bands and quasi-gamma bands for both the nuclei agree reasonably well with experiment. The negative parity bands are also well-reproduced in our calculation. The calculated B(E2) values are compared with available experimental data. The nature of angular momentum alignment in the ground band is also discussed.

PACS Nos.: 21.10.–k, 21.60.Jz, 27.50.+e
Stress-dependent regulation of Pbh1, a BIR domain-containing protein, in the fission yeast
Authors: Cho Nam-Chul , Kang Hyun-Jung , Lim Hye-Won , Kim Byung-Chul , Park Eun-Hee , Lim Chang-Jin
Published: May 2007

To elicit the physiological roles of Pbh1, a baculoviral IAP repeat (BIR) domain-containing protein, in Schizosaccharomyces pombe, we investigated if Pbh1 expression is regulated by stress. The upstream region (1221 bp) of the pbh1 gene was fused into the promoterless β-galactosidase gene of the shuttle vector YEp367R, and the resultant fusion plasmid was named pPbh04. The synthesis of β-galactosidase from the pbh1-lacZ fusion gene was markedly enhanced by sodium nitroprusside (SNP) generating nitric oxide. The basal expression of the pbh1 gene required the presence of Pap1. Pap1 also mediated the induction of the pbh1 gene by SNP and nitrogen starvation. Pap1-dependent induction of the pbh1 gene by SNP was confirmed by the enhanced level of the pbh1 mRNA in Pap1-positive cells but not in Pap1-negative cells. Taken together, it was demonstrated that the pbh1 genes are positively regulated by nitrosative and nitrogen starvation stresses in Pap1-dependent manner.Key words: fission yeast, nitrosative stress, nutritional stress, nitrogen starvation, Pap1, Pbh1, regulation, Schizosaccharomyces pombe.
Fish protein hydrolysis by a psychrotrophic marine bacterium isolated from the gut of hake (Merluccius hubbsi)
Authors: Belchior Silvia G. Estevao , Vacca Gabriela
Published: May 2007

The aims of this study were to identify a psychrotrophic bacterium, strain CR41, producing a cold adapted protease during growth at low temperatures and to evaluate the ability of the cells to hydrolyze hake fish protein. The strain was isolated from the intestinal tract of hake collected from the San Jorge Gulf (Patagonia, Argentina) and it was identified as Pseudoalteromonas. Growth and fish protein hydrolysis were determined using an aerated simple mineral medium plus 10% fish protein concentrate. Proteolytic activity was measured at 7 and 22 °C during culture in the concentrate. Protease production started in the exponential growth phase and reached a maximum during stationary phase. Protease activity at 7 °C was lower than at 22 °C. After 8 h of incubation, the percentage of hydrolyzed protein was 84% at 7 °C and 95% at 22 °C. Electrophoresis detection showed that degradation of muscle hake proteins was complete at both temperatures, and in gelatin zymograms extracellular activity showed two proteolytic bands with apparent molecular masses of approximately 31.6 and 62 kDa.Key words: hake, protein hydrolysis, psychrotrophic bacteria, Pseudoalteromonas sp.