bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification of an extracellular matrix protein adhesin, EmaA, which mediates the adhesion of Actinobacillus actinomycetemcomitans to collagen
Authors: Mintz Keith P.
Published: August 2004

Actinobacillus actinomycetemcomitansis an aetiologic agent in the development of periodontal and some systemic diseases in humans. This pathogen localizes to the underlying connective tissue of the oral cavity in individuals with periodontal disease. The adhesion ofA. actinomycetemcomitansto extracellular matrix components of the connective tissue prompted this study to identify gene products mediating the interaction ofA. actinomycetemcomitansto these molecules. A transposon mutagenesis system was optimized for use inA. actinomycetemcomitansand used to generate an insertional mutant library. A total of 2300 individual insertion transposon mutants were screened for changes in the adhesion to collagen and fibronectin. Mutants were identified which exhibited the following phenotypes: a decrease in collagen binding; a decrease in fibronectin binding; a decrease in binding to both proteins; and an increase in binding to both collagen and fibronectin. The identification of mutants defective in adhesion to the individual proteins indicates that distinct adhesins are expressed by this organism. Molecular analysis of these mutants implicated 11 independent loci in protein adhesion. One gene,emaA, is likely to encode a direct mediator of collagen adhesion, based on predicted protein features homologous to the collagen-binding protein YadA ofYersinia enterocolitica. EmaA was localized to the outer membrane, as expected for an adhesin. Reduction in fibronectin adhesion appeared to be influenced by abrogation of proteins involved in molybdenum-cofactor biosynthesis. Several other loci identified as reducing or increasing adhesion to both collagen and fibronectin are suggested to be involved in regulatory cascades that promote or repress expression of collagen and fibronectin adhesins. Collectively, the results support the hypothesis thatA. actinomycetemcomitanshost colonization involves afimbrial adhesins for extracellular matrix proteins, and that the expression of adhesion is modulated by global regulatory mechanisms.
Post-transcriptional regulation of the Bacillus subtilis pst operon encoding a phosphate-specific ABC transporter
Authors: Allenby Nicholas E. E. , O'Connor Nicola , Prágai Zoltán , Carter Noel M. , Miethke Marcus , Engelmann Susanne , Hecker Michael , Wipat Anil , Ward Alan C. , Harwood Colin R.
Published: August 2004

During phosphate starvation,Bacillus subtilisregulates genes in the PhoP regulon to reduce the cell's requirement for this essential substrate and to facilitate the recovery of inorganic phosphate from organic sources such as teichoic and nucleic acids. Among the proteins that are highly induced under these conditions is PstS, the phosphate-binding lipoprotein component of a high-affinity ABC-type phosphate transporter. PstS is encoded by the first gene in thepstoperon, the other four members of which encode the integral membrane and cytoplasmic components of the transporter. The transcription of thepstoperon was analysed using a combination of methods, including transcriptional reporter gene technology, Northern blotting and DNA arrays. It is shown that the primary transcript of thepstoperon is processed differentially to maintain higher concentrations of PstS relative to other components of the transporter. The comparative studies have revealed limitations in the use of reporter gene technology for analysing the transcription of operons in which the messenger RNA transcript is differentially processed.
Partial purification and characterization of a non-cyanobacterial cyanophycin synthetase from Acinetobacter calcoaceticus strain ADP1 with regard to substrate specificity, substrate affinity and binding to cyanophycin
Authors: Krehenbrink Martin , Steinbüchel Alexander
Published: August 2004

This study reports, for the first time, purification and biochemical characterization of a cyanophycin synthetase from a non-cyanobacterial strain. Cyanophycin synthetase of Acinetobacter calcoaceticus strain ADP1 was purified 69-fold from recombinant Escherichia coli by two chromatographic steps and one novel affinity step utilizing the Mg2+-dependent binding of the enzyme to cyanophycin. Unlike cyanobacterial cyanophycin synthetases characterized so far, the purified enzyme from A. calcoaceticus strain ADP1 did not accept lysine as an alternative substrate to arginine. The apparent K
m-values for arginine (47 μM) and aspartic acid (240 μM) were similar to those of known cyanophycin synthetases from cyanobacteria, but this enzyme had a slightly higher affinity for aspartic acid. In addition, the two different ATP-binding sites of the enzyme were characterized independently of each other with respect to K
m values for ATP. The ATP-binding site responsible for the addition of arginine was found to have a much higher affinity for ATP (38 μM) than that responsible for the addition of aspartate (210 mM). Furthermore, the binding of the enzyme to the two possible forms of cyanophycin granule polypeptide (CGP), CGP-Asp and CGP-Arg, was studied. While both forms bound around 30–40 % of the enzyme activity present under the assay conditions, binding was Mg2+-dependent in the case of CGP-Asp. Two-dimensional gel electrophoresis revealed that both forms of cyanophycin were equally abundant in cyanophycin-accumulating cells of A. calcoaceticus ADP1.
Kluyveromyces phaffii killer toxin active against wine spoilage yeasts: purification and characterization
Authors: Comitini Francesca , Pietro Natalia Di , Zacchi Laura , Mannazzu Ilaria , Ciani Maurizio
Published: August 2004

The killer toxin secreted byKluyveromyces phaffii(KpKt) is active against spoilage yeast under winemaking conditions and thus has potential applications in the biocontrol of undesired micro-organisms in the wine industry. Biochemical characterization and N-terminal sequencing of the purified toxin show that KpKt is a glycosylated protein with a molecular mass of 33 kDa. Moreover, it shows 93 % and 80 % identity to aβ-1,3-glucanase ofSaccharomyces cerevisiaeand aβ-1,3-glucan transferase ofCandida albicans, respectively, and it is active on laminarin and glucan, thus showing aβ-glucanase activity. Competitive inhibition of killer activity by cell-wall polysaccharides suggests that glucan (β-1,3 andβ-1,6 branched glucans) represents the first receptor site of the toxin on the envelope of the sensitive target. Flow cytometry analysis of the sensitive target after treatment with KpKt and K1 toxin ofS. cerevisiae, known to cause loss of cell viability via formation of pores in the cell membrane, suggests a different mode of action for KpKt.
Attenuated virulence and protective efficacy of a Burkholderia pseudomallei bsa type III secretion mutant in murine models of melioidosis
Authors: Stevens Mark P. , Haque Ashraful , Atkins Timothy , Hill Jim , Wood Michael W. , Easton Anna , Nelson Michelle , Underwood-Fowler Cindy , Titball Richard W. , Bancroft Gregory J. , Galyov Edouard E.
Published: August 2004

Melioidosis is a severe infectious disease of animals and humans caused by the Gram-negative intracellular pathogen Burkholderia pseudomallei. An Inv/Mxi-Spa-like type III protein secretion apparatus, encoded by the B. pseudomallei bsa locus, facilitates bacterial invasion of epithelial cells, escape from endocytic vesicles and intracellular survival. This study investigated the role of the Bsa type III secretion system in the pathogenesis of melioidosis in murine models. B. pseudomallei bipD mutants, lacking a component of the translocation apparatus, were found to be significantly attenuated following intraperitoneal or intranasal challenge of BALB/c mice. Furthermore, a bipD mutant was attenuated in C57BL/6 IL-12 p40−/− mice, which are highly susceptible to B. pseudomallei infection. Mutation of bipD impaired bacterial replication in the liver and spleen of BALB/c mice in the early stages of infection. B. pseudomallei mutants lacking either the type III secreted guanine nucleotide exchange factor BopE or the putative effectors BopA or BopB exhibited varying degrees of attenuation, with mutations in bopA and bopB causing a significant delay in median time to death. This indicates that bsa-encoded type III secreted proteins may act in concert to determine the outcome of B. pseudomallei infection in mice. Mice inoculated with the B. pseudomallei bipD mutant were partially protected against subsequent challenge with wild-type B. pseudomallei. However, immunization of mice with purified BipD protein was not protective.
NblA is essential for phycobilisome degradation in Anabaena sp. strain PCC 7120 but not for development of functional heterocysts
Authors: Baier Kerstin , Lehmann Heike , Stephan Dirk Paul , Lockau Wolfgang
Published: August 2004

Phycobilisomes (PBS) are the major light-harvesting complexes of cyanobacteria. These usually blue-coloured multiprotein assemblies are rapidly degraded when the organisms are starved for combined nitrogen. This proteolytic process causes a colour change of the cyanobacterial cells from blue-green to yellow-green (‘bleaching’). As is well documented for the unicellular, non-diazotrophic cyanobacteriaSynechococcus elongatusPCC 7942 andSynechocystissp. PCC 6803, a gene termednblAplays a key role in PBS degradation. Filamentous, diazotrophic cyanobacteria likeAnabaenaadapt to nitrogen deprivation by differentiation of N2-fixing heterocysts. However, during the first hours after nitrogen deprivation all cells degrade their PBS. When heterocysts mature and nitrogenase becomes active, vegetative cells resynthesize their light-harvesting complexes while in heterocysts the phycobiliprotein content remains very low. Expression and function ofnblAinAnabaenasp. PCC 7120 was investigated. This strain has twonblAhomologous genes, one on the chromosome (nblA) and one on plasmid delta (nblA-p). Northern blot analysis indicated that only the chromosomalnblAgene is up-regulated upon nitrogen starvation. Mutants with interruptednblAandnblA-pgenes, respectively, grew on N2and developed functional heterocysts. Mutant ΔnblA-pbehaved like the wild-type. However, mutant ΔnblAwas unable to degrade its PBS, which was most obvious in non-bleaching heterocysts. The results show that NblA, encoded by the chromosomalnblAgene, is required for PBS degradation inAnabaenabut is not essential for heterocyst differentiation.
Characterization of the Bacillus subtilis YxdJ response regulator as the inducer of expression for the cognate ABC transporter YxdLM
Authors: Joseph Pascale , Guiseppi Annick , Sorokin Alexei , Denizot François
Published: August 2004

The genome ofBacillus subtilis, like those of some other AT-rich Gram-positive bacteria, has the uncommon feature of containing several copies of arrangements in which the genes encoding two-component and cognate ABC transporter systems are adjacent. As the function of one of these systems, the product of theyxdlocus, is still unknown, it was analysed further in order to get some clues on the physiological role of the gene products it encodes. TheyxdJgene was shown to encode a DNA-binding protein that directly controls transcription of the neighbouring operon encoding the ABC transporter YxdLM. Primer extension and DNase protection experiments allowed precise definition of theyxdLMtranscription start and controlling region. Two putative direct repeats were identified that are proposed to be the YxdJ response regulator binding sites. Whole-cell transcriptome analyses revealed that the YxdJ regulon is extremely restricted. In addition to theyxdJKLMyxeAoperon, only a few genes involved in modifications of the bacterial cell wall were shown to be regulated by YxdJ.
Expression of bacteriophage ϕEa1h lysozyme in Escherichia coli and its activity in growth inhibition of Erwinia amylovora
Authors: Kim Won-Sik , Salm Heike , Geider Klaus
Published: August 2004

A 3·3 kb fragment from Erwinia amylovora phage ϕEa1h in plasmid pJH94 was previously characterized and found to contain an exopolysaccharide depolymerase (dpo) gene and two additional ORFs encoding 178 and 119 amino acids. ORF178 (lyz) and ORF119 (hol) were found to overlap by 19 bp and they resembled genes encoding lysozymes and holins. In nucleotide sequence alignments, lyz had structurally conserved regions with residues important for lysozyme function. The lyz gene was cloned into an expression vector and expressed in Escherichia coli. Active lysozyme was detected only when E. coli cells with the lyz gene and a kanamycin-resistance cassette were grown in the presence of kanamycin. Growth of Erw. amylovora was inhibited after addition of enzyme exceeding a threshold for lysozyme to target cells. When immature pears were soaked in lysates of induced cells, symptoms such as ooze formation and necrosis were retarded or inhibited after inoculation with Erw. amylovora.
Primitive neural stem cells from the mammalian epiblast differentiate to definitive neural stem cells under the control of Notch signaling
Authors: Hitoshi Seiji , Seaberg Raewyn M. , Koscik Cheryl , Alexson Tania , Kusunoki Susumu , Kanazawa Ichiro , Tsuji Shoji , van der Kooy Derek
Published: August 2004

Basic fibroblast growth
factor (FGF2)-responsive definitive neural stem cells first appear in
embryonic day 8.5 (E8.5) mouse embryos, but not in earlier embryos,
although neural tissue exists at E7.5. Here, we demonstrate that
leukemia inhibitory factor-dependent (but not FGF2-dependent)
sphere-forming cells are present in the earlier (E5.5–E7.5) mouse
embryo. The resultant clonal sphere cells possess self-renewal capacity
and neural multipotentiality, cardinal features of the neural stem cell.
However, they also retain some nonneural properties, suggesting that
they are the in vivo cells' equivalent of the primitive neural stem
cells that form in vitro from embryonic stem cells. The generation of
the in vivo primitive neural stem cell was independent of Notch
signaling, but the activation of the Notch pathway was important for the
transition from the primitive to full definitive neural stem cell
properties and for the maintenance of the definitive neural stem cell
state.
DNA trajectory in the Gal repressosome
Authors: Semsey Szabolcs , Tolstorukov Michail Y. , Virnik Konstantin , Zhurkin Victor B. , Adhya Sankar
Published: August 2004

The Gal repressosome is a higher-order nucleoprotein complex that represses transcription of the gal operon in Escherichia coli. During the repressosome assembly, a DNA loop is formed by the interaction of two GalR dimers, bound to two spatially separated operators, OE and OI, flanking the gal promoters. Structure-based genetic analysis indicated that GalR homodimers interact directly and form a V-shaped stacked tetramer in repressosome, further stabilized by HU binding to an architecturally critical position on the DNA. In this scheme of GalR tetramerization, the alignment of the operators in the DNA loop could be in either parallel (PL) or antiparallel (AL) mode. As each mode can have two alternative geometries differing in the mutual stacking of the OE- and OI-bound GalR dimers, it is possible to have four different DNA trajectories in the repressosome. Feasibilities of these trajectories were tested by in vitro transcription repression assays, first by isolating GalR mutants with altered operator specificity and then by constructing four different potential loops with mutant GalR heterodimers bound to specifically designed hybrid operators in such a way as to give rise to only one of the four putative trajectories. Results show that OE and OI adopt a mutual antiparallel orientation in an under-twisted DNA loop, consistent with the energetically optimal structural model. In this structure the center of the HU-binding site is located at the apex of the DNA loop. The approach reported here can be used to distinguish between otherwise indistinguishable DNA trajectories in complex nucleoprotein machines.
Effect of a C/EBP gene replacement on mitochondrial biogenesis in fat cells
Authors: Chiu Chih-Hsien , Lin Wen-Der , Huang Shao-Yong , Lee Ying-Hue
Published: August 2004

CCAAT/enhancer-binding proteins, C/EBPα and C/EBPβ, are required for fat cell differentiation and maturation. Previous studies showed that replacement of C/EBPα with C/EBPβ, generating the β/β alleles in the mouse genome, prevents lipid accumulation in white adipose tissue (WAT). In this study, β/β mice lived longer and had higher energy expenditure than their control littermates due to increased WAT energy oxidation. The WAT of β/β mice was enriched with metabolically active, thermogenic mitochondria known for energy burning. The β/β allele exerted its effect through the elevated expression of the G protein α stimulatory subunit (Gαs) in WAT. Gαs, when overexpressed in fat-laden 3T3-L1 cells, stimulated mitochondrial biogenesis similar to that seen in the WAT of β/β mice, and effectively diminished the stored lipid pool.
Reprogramming of a melanoma genome by nuclear transplantation
Authors: Hochedlinger Konrad , Blelloch Robert , Brennan Cameron , Yamada Yasuhiro , Kim Minjung , Chin Lynda , Jaenisch Rudolf
Published: August 2004

We have used nuclear transplantation to test whether the reprogramming activity of oocytes can reestablish developmental pluripotency of malignant cancer cells. We show here that the nuclei of leukemia, lymphoma, and breast cancer cells could support normal preimplantation development to the blastocyst stage but failed to produce embryonic stem (ES) cells. However, a blastocyst cloned from a RAS-inducible melanoma nucleus gave rise to ES cells with the potential to differentiate into multiple cell types in vivo including melanocytes, lymphocytes, and fibroblasts. Chimeras produced from these ES cells developed cancer with higher penetrance, shorter latency, and an expanded tumor spectrum when compared with the donor mouse model. These results demonstrate that the secondary changes of a melanoma nucleus are compatible with a broad developmental potential but predispose mice to melanomas and other malignant tumors on reactivation of RAS. Our findings serve as a paradigm for studying the tumorigenic effect of a given cancer genome in the context of a whole animal.
Mutually exclusive mutations of the Pten and ras pathways in skin tumor progression
Authors: Mao Jian-Hua , To Minh D. , Perez-Losada Jesus , Wu Di , Del Rosario Reyno , Balmain Allan
Published: August 2004

Pten heterozygous (Pten+/-) mice develop increased papilloma numbers and show decreased carcinoma latency time in comparison with controls after skin treatment with dimethyl benzanthracene (DMBA) and tetradecanoyl-phorbol acetate (TPA). H-ras mutation is normally a hallmark of DMBA-TPA-induced skin tumors, but 70% of carcinomas from Pten+/- mice do not exhibit this mutation, and in all cases have lost the wild-type Pten allele. Tumors that retain the Pten wild-type allele also have H-ras mutations, indicating that activation of H-ras and complete loss of Pten are mutually exclusive events in skin carcinomas. Mitogen-activated protein kinase (MAPK) is consistently activated in the tumors with H-ras mutations, but is strongly down-regulated in Pten-/- tumors, suggesting that this pathway is dispensable for skin carcinoma formation. These data have important implications in designing individual therapeutic strategies for the treatment of cancer.
Functional links between telomeres and proteins of the DNA-damage response
Authors: d'Adda di Fagagna Fabrizio , Teo Soo-Hwang , Jackson Stephen P.
Published: August 2004

In response to DNA
damage, cells engage a complex set of events that together comprise the
DNA-damage response (DDR). These events bring about the repair of the
damage and also slow down or halt cell cycle progression until the
damage has been removed. In stark contrast, the ends of linear
chromosomes, telomeres, are generally not perceived as DNA damage by the
cell even though they terminate the DNA double-helix. Nevertheless, it
has become clear over the past few years that many proteins involved in
the DDR, particularly those involved in responding to DNA double-strand
breaks, also play key roles in telomere maintenance. In this review, we
discuss the current knowledge of both the telomere and the DDR, and then
propose an integrated model for the events associated with the
metabolism of DNA ends in these two distinct physiological contexts.
Distinct roles of c-Abl and Atm in oxidative stress response are mediated by protein kinase C δ
Authors: Li Baojie , Wang Xueying , Rasheed Naslin , Hu Yuanyu , Boast Sharon , Ishii Tetsuro , Nakayama Keiko , Nakayama Keiichi I. , Goff Stephen P.
Published: August 2004

c-Abl and Atm have been implicated in cell responses to DNA damage and oxidative stress. However, the molecular mechanisms by which they regulate oxidative stress response remain unclear. In this report, we show that deficiency of c-Abl and deficiency of ATM differentially altered cell responses to oxidative stress by induction of antioxidant protein peroxiredoxin I (Prx I) via Nrf2 and cell death, both of which required protein kinase C (PKC) δ activation and were mediated by reactive oxygen species.c-abl-/-osteoblasts displayed enhanced Prx I induction, elevated Nrf2 levels, and hypersusceptibility to arsenate, which were reinstated by reconstitution of c-Abl;Atm-/-osteoblasts showed the opposite. These phenotypes correlated with increased PKC δ expression inc-abl-/-osteoblasts and decreased PKC δ expression inAtm-/-cells, respectively. The enhanced responses ofc-abl-/-osteoblasts could be mimicked by overexpression of PKC δ in normal cells and impeded by inhibition of PKC δ, and diminished responses ofAtm-/-cells could be rescued by PKC δ overexpression, indicating that PKC δ mediated the effects of c-Abl and ATM in oxidative stress response. Hence, our results unveiled a previously unrecognized mechanism by which c-Abl and Atm participate in oxidative stress response.
A role for centromere pairing in meiotic chromosome segregation
Authors: Kemp Benedict , Boumil Rebecca Maxfield , Stewart Mara N. , Dawson Dean S.
Published: August 2004

In meiosis I, exchanges provide a connection between homologous chromosome pairs that facilitates their proper attachment to the meiotic spindle. In many eukaryotes, homologous chromosomes that fail to become linked by exchanges exhibit elevated levels of meiotic errors, but they do not segregate randomly, demonstrating that mechanisms beyond exchange can promote proper meiosis I segregation. The experiments described here demonstrate the existence of a meiotic centromere pairing mechanism in budding yeast. This centromere pairing mediates the meiosis I bipolar spindle attachment of nonexchange chromosome pairs and likely plays the same role for all homologous chromosome pairs.
Stress-Induced DNA Duplex Destabilization (SIDD) in the E. coli Genome: SIDD Sites Are Closely Associated With Promoters
Authors: Wang Huiquan , Noordewier Michiel , Benham Craig J.
Published: August 2004

We present the first analysis of stress-induced DNA duplex destabilization (SIDD) in a complete chromosome, the Escherichia coli K12 genome. We used a newly developed method to calculate the locations and extents of stress-induced destabilization to single-base resolution at superhelix density σ = –0.06. We find that SIDD sites in this genome show a statistically highly significant tendency to avoid coding regions. And among intergenic regions, those that either contain documented promoters or occur between divergently transcribing coding regions, and hence may be inferred to contain promoters, are associated with strong SIDD sites in a statistically highly significant manner. Intergenic regions located between convergently transcribing genes, which are inferred not to contain promoters, are not significantly enriched for destabilized sites. Statistical analysis shows that a strongly destabilized intergenic region has an 80% chance of containing a promoter, whereas an intergenic region that does not contain a strong SIDD site has only a 24% chance. We describe how these observations may illuminate specific mechanisms of regulation, and assist in the computational identification of promoter locations in prokaryotes.
Genetic Divergence of the Rhesus Macaque Major Histocompatibility Complex
Authors: Daza-Vamenta Riza , Glusman Gustavo , Rowen Lee , Guthrie Brandon , Geraghty Daniel E.
Published: August 2004

The major histocompatibility complex (MHC) is comprised of the class I, class II, and class III regions, including the MHC class I and class II genes that play a primary role in the immune response and serve as an important model in studies of primate evolution. Although nonhuman primates contribute significantly to comparative human studies, relatively little is known about the genetic diversity and genomics underlying nonhuman primate immunity. To address this issue, we sequenced a complete rhesus macaque MHC spanning over 5.3 Mb, and obtained an additional 2.3 Mb from a second haplotype, including class II and portions of class I and class III. A major expansion of from six class I genes in humans to as many as 22 active MHC class I genes in rhesus and levels of sequence divergence some 10-fold higher than a similar human comparison were found, averaging from 2% to 6% throughout extended portions of class I and class II. These data pose new interpretations of the evolutionary constraints operating between MHC diversity and T-cell selection by contrasting with models predicting an optimal number of antigen presenting genes. For the clinical model, these data and derivative genetic tools can be implemented in ongoing genetic and disease studies that involve the rhesus macaque.
Evolution of Eukaryotic Transcription: Insights From the Genome of Giardia lamblia
Authors: Best Aaron A. , Morrison Hilary G. , McArthur Andrew G. , Sogin Mitchell L. , Olsen Gary J.
Published: August 2004

The Giardia lamblia genome sequencing project affords us a unique opportunity to conduct comparative analyses of core cellular systems between early and late-diverging eukaryotes on a genome-wide scale. We report a survey to identify canonical transcription components in Giardia, focusing on RNA polymerase (RNAP) subunits and transcription-initiation factors. Our survey revealed that Giardia contains homologs to 21 of the 28 polypeptides comprising eukaryal RNAPI, RNAPII, and RNAPIII; six of the seven RNAP subunits without giardial homologs are polymerase specific. Components of only four of the 12 general transcription initiation factors have giardial homologs. Surprisingly, giardial TATA-binding protein (TBP) is highly divergent with respect to archaeal and higher eukaryotic TBPs, and a giardial homolog of transcription factor IIB was not identified. We conclude that Giardia represents a transition during the evolution of eukaryal transcription systems, exhibiting a relatively complete set of RNAP subunits and a rudimentary basal initiation apparatus for each transcription system. Most class-specific RNAP subunits and basal initiation factors appear to have evolved after the divergence of Giardia from the main eukaryotic line of descent. Consequently, Giardia is predicted to be unique in many aspects of transcription initiation with respect to paradigms derived from studies in crown eukaryotes.
Large-Scale Validation of Single Nucleotide Polymorphisms in Gene Regions
Authors: Nelson Matthew R. , Marnellos George , Kammerer Stefan , Hoyal Carolyn R. , Shi Michael M. , Cantor Charles R. , Braun Andreas
Published: August 2004

Genome-wide association studies using large numbers of bi-allelic single nucleotide polymorphisms (SNPs) have been proposed as a potentially powerful method for identifying genes involved in common diseases. To assemble a SNP collection appropriate for large-scale association, we designed assays for 226,099 publicly available SNPs located primarily within known and predicted gene regions. Allele frequencies were estimated in a sample of 92 CEPH Caucasians using chip-based MALDI-TOF mass spectrometry with pooled DNA. Of the 204,200 designed assays that were functional, 125,799 SNPs were determined to be polymorphic (minor allele frequency >0.02), of which 101,729 map uniquely to the human genome. Many of the commonly available RefSNP annotations were predictive of polymorphic status and could be used to improve the selection of SNPs from the public domain for genetic research. The set of uniquely mapping, polymorphic SNPs is located within 10 kb of 66% of known and predicted genes annotated in LocusLink, which could prove useful for large-scale disease association studies.
Regional Patterns of Gene Expression in Human and Chimpanzee Brains
Authors: Khaitovich Philipp , Muetzel Bjoern , She Xinwei , Lachmann Michael , Hellmann Ines , Dietzsch Janko , Steigele Stephan , Do Hong-Hai , Weiss Gunter , Enard Wolfgang , Heissig Florian , Arendt Thomas , Nieselt-Struwe Kay , Eichler Evan E. , Pääbo Svante
Published: August 2004

We have analyzed gene expression in various brain regions of humans and chimpanzees. Within both human and chimpanzee individuals, the transcriptomes of the cerebral cortex are very similar to each other and differ more between individuals than among regions within an individual. In contrast, the transcriptomes of the cerebral cortex, the caudate nucleus, and the cerebellum differ substantially from each other. Between humans and chimpanzees, 10% of genes differ in their expression in at least one region of the brain. The majority of these expression differences are shared among all brain regions. Whereas genes encoding proteins involved in signal transduction and cell differentiation differ significantly between brain regions within individuals, no such pattern is seen between the species. However, a subset of genes that show expression differences between humans and chimpanzees are distributed nonrandomly across the genome. Furthermore, genes that show an elevated expression level in humans are statistically significantly enriched in regions that are recently duplicated in humans.
A Non-EST-Based Method for Exon-Skipping Prediction
Authors: Sorek Rotem , Shemesh Ronen , Cohen Yuval , Basechess Ortal , Ast Gil , Shamir Ron
Published: August 2004

It is estimated that between 35% and 74% of all human genes can undergo alternative splicing. Currently, the most efficient methods for large-scale detection of alternative splicing use expressed sequence tags (ESTs) or microarray analysis. As these methods merely sample the transcriptome, splice variants that do not appear in deeply sampled tissues have a low probability of being detected. We present a new method by which we can predict that an internal exon is skipped (namely whether it is a cassette-exon) merely based on its naked genomic sequence and on the sequence of its mouse ortholog. No other data, such as ESTs, are required for the prediction. Using our method, which was experimentally validated, we detected hundreds of novel splice variants that were not detectable using ESTs. We show that a substantial fraction of the splice variants in the human genome could not be identified through current human EST or cDNA data.
Segmental Phylogenetic Relationships of Inbred Mouse Strains Revealed by Fine-Scale Analysis of Sequence Variation Across 4.6 Mb of Mouse Genome
Authors: Frazer Kelly A. , Wade Claire M. , Hinds David A. , Patil Nila , Cox David R. , Daly Mark J.
Published: August 2004

High-density SNP screening of panels of inbred mouse strains has been proposed as a method to accelerate the identification of genes associated with complex biomedical phenotypes. To evaluate the potential of these studies, a more detailed understanding of the fine structure of sequence variation across inbred mouse strains is needed. Here, we use high-density oligonucleotide arrays to discover an extremely dense set of SNPs in 13 classical and two wild-derived inbred strains in five genomic intervals totaling 4.6 Mb of DNA sequence, and then analyze the segmental haplotype structure defined by these high-density SNPs. This analysis reveals segments ranging from 12 to 608 kb in length within which the inbred strains have a simple and distinct phylogenetic relationship with typically two or three clades accounting for the 13 classical strains examined. The phylogenetic relationships among strains change abruptly and unpredictably from segment to segment, and are distinct in each of the five genomic regions examined. The data suggest that at least 12 strains would need to be resequenced for exhaustive SNP discovery in every region of the mouse genome, that ∼97% of the variation among inbred strains is ancestral (between clades) and ∼3% private (within clades), and provides critical insights into the proposed use of panels of inbred strains to identify genes underlying quantitative trait loci.
cpnDB: A Chaperonin Sequence Database
Authors: Hill Janet E. , Penny Susanne L. , Crowell Kenneth G. , Goh Swee Han , Hemmingsen Sean M.
Published: August 2004

Type I chaperonins are molecular chaperones present in virtually all bacteria, some archaea and the plastids and mitochondria of eukaryotes. Sequences of cpn60 genes, encoding 60-kDa chaperonin protein subunits (CPN60, also known as GroEL or HSP60), are useful for phylogenetic studies and as targets for detection and identification of organisms. Conveniently, a 549–567-bp segment of the cpn60 coding region can be amplified with universal PCR primers. Here, we introduce cpnDB, a curated collection of cpn60 sequence data collected from public databases or generated by a network of collaborators exploiting the cpn60 target in clinical, phylogenetic, and microbial ecology studies. The growing database currently contains ∼2000 records covering over 240 genera of bacteria, eukaryotes, and archaea. The database also contains over 60 sequences for the archaeal Type II chaperonin (thermosome, a homolog of eukaryotic cytoplasmic chaperonin) from 19 archaeal genera. As the largest curated collection of sequences available for a protein-encoding gene, cpnDB provides a resource for researchers interested in exploiting the power of cpn60 as a diagnostic or as a target for phylogenetic or microbial ecology studies, as well as those interested in broader subjects such as lateral gene transfer and codon usage. We built cpnDB from open source tools and it is available at http://cpndb.cbr.nrc.ca.
The Complete Genome and Proteome of Mycoplasma mobile
Authors: Jaffe Jacob D. , Stange-Thomann Nicole , Smith Cherylyn , DeCaprio David , Fisher Sheila , Butler Jonathan , Calvo Sarah , Elkins Tim , FitzGerald Michael G. , Hafez Nabil , Kodira Chinnappa D. , Major John , Wang Shunguang , Wilkinson Jane , Nicol Robert , Nusbaum Chad , Birren Bruce , Berg Howard C. , Church George M.
Published: August 2004

Although often considered “minimal” organisms, mycoplasmas show a wide range of diversity with respect to host environment, phenotypic traits, and pathogenicity. Here we report the complete genomic sequence and proteogenomic map for the piscine mycoplasma Mycoplasma mobile, noted for its robust gliding motility. For the first time, proteomic data are used in the primary annotation of a new genome, providing validation of expression for many of the predicted proteins. Several novel features were discovered including a long repeating unit of DNA of ∼2435 bp present in five complete copies that are shown to code for nearly identical yet uniquely expressed proteins. M. mobile has among the lowest DNA GC contents (24.9%) and most reduced set of tRNAs of any organism yet reported (28). Numerous instances of tandem duplication as well as lateral gene transfer are evident in the genome. The multiple available complete genome sequences for other motile and immotile mycoplasmas enabled us to use comparative genomic and phylogenetic methods to suggest several candidate genes that might be involved in motility. The results of these analyses leave open the possibility that gliding motility might have arisen independently more than once in the mycoplasma lineage.
Increased Glutathione Biosynthesis Plays a Role in Nickel Tolerance in Thlaspi Nickel Hyperaccumulators[W]
Authors: Freeman John L. , Persans Michael W. , Nieman Ken , Albrecht Carrie , Peer Wendy , Pickering Ingrid J. , Salt David E.
Published: August 2004

Worldwide more than 400 plant species are now known that hyperaccumulate various trace metals (Cd, Co, Cu, Mn, Ni, and Zn), metalloids (As) and nonmetals (Se) in their shoots. Of these, almost one-quarter are Brassicaceae family members, including numerous Thlaspi species that hyperaccumulate Ni up to 3% of there shoot dry weight. We observed that concentrations of glutathione, Cys, and O-acetyl-l-serine (OAS), in shoot tissue, are strongly correlated with the ability to hyperaccumulate Ni in various Thlaspi hyperaccumulators collected from serpentine soils, including Thlaspi goesingense, T. oxyceras, and T. rosulare, and nonaccumulator relatives, including T. perfoliatum, T. arvense, and Arabidopsis thaliana. Further analysis of the Austrian Ni hyperaccumulator T. goesingense revealed that the high concentrations of OAS, Cys, and GSH observed in this hyperaccumulator coincide with constitutively high activity of both serine acetyltransferase (SAT) and glutathione reductase. SAT catalyzes the acetylation of l-Ser to produce OAS, which acts as both a key positive regulator of sulfur assimilation and forms the carbon skeleton for Cys biosynthesis. These changes in Cys and GSH metabolism also coincide with the ability of T. goesingense to both hyperaccumulate Ni and resist its damaging oxidative effects. Overproduction of T. goesingense SAT in the nonaccumulator Brassicaceae family member Arabidopsis was found to cause accumulation of OAS, Cys, and glutathione, mimicking the biochemical changes observed in the Ni hyperaccumulators. In these transgenic Arabidopsis, glutathione concentrations strongly correlate with increased resistance to both the growth inhibitory and oxidative stress induced effects of Ni. Taken together, such evidence supports our conclusion that elevated GSH concentrations, driven by constitutively elevated SAT activity, are involved in conferring tolerance to Ni-induced oxidative stress in Thlaspi Ni hyperaccumulators.
The story of a schizoid organization
Authors: Richardson Susan , Cullen John , Richardson Bill
Published: July 2004

Reports on the uncovering of a small‐firm reality where the self‐gratifying behaviour of the owner/manager threatens the survival of the organization. Takes a “middle‐range thinking” approach to mould ethnographic data, and demonstrates and develops, via a case study, models of change within organizations. Identifies the importance of power in such change‐resistant contexts, and draws conclusions from the work which suggest that in such contexts “second order” change is unlikely to be achieved and only “pseudo‐colonization” can be sustained in the long term. It is the authors’ view that this context is a common one and is worthy of further research since there may be far‐reaching implications for a whole range of stakeholders associated with such an organization.
Auditor time budget pressure: consequences and antecedents
Authors: Otley David T. , Pierce Bernard J.
Published: July 2004

Research suggests that dysfunctional behaviour by auditors may be related to the perceived tightness of time budgets. Using data collected from practising auditors, examines the nature of such a relationship. Found that the frequency of dysfunctional behaviour increased sharply as budgets were seen to approach unattainable levels of performance. Recognizing the importance of auditors’ perceptions regarding the attainability of budgets, examines antecedent variables affecting budget attainability. Found that the influence of client fee expectations, the level of audit senior participation and the influence of the audit programme were significant influences. Discusses implications for practice and possibilities for future research.
Lobbying on accounting issues
Authors: Weetman P. , Davie E.S. , Collins W.
Published: July 2004

The lack of user response in written submissions on accounting standards has been noted in previous research. That lack of user response has limited empirical investigations with the result that lobbying by users has to be understood largely from theoretical literature in the political and social arena. Aims to add empirical data to the understanding of the user perspective, taking as a focus for the investigation the lobbying of the UK Accounting Standards Board in respect of theOperating and Financial Review.Four issues are chosen for analysis: usefulness to users, effectiveness of voluntary compliance, confidentiality and provision of forward‐looking information. Shows that the ASB’s responses on these four major issues are rational in terms of the relative intensity of lobbying. Using interview methods to obtain user responses shows that a different perception of lobbying intensity may be derived from a comparison of preparer and user responses, such that preparers’ views may have given too negative an impression to the ASB. The relative persuasive power of different arguments is shown to depend on how the comparison is undertaken. Uses the quantitative analysis of responses as a base from which to explore the potential for strategic lobbying to disguise self‐interested motivation and provides some insight on the lack of user response.
Some determinants of social and environmental disclosures in New Zealand companies
Authors: Hackston David , Milne Markus J.
Published: July 2004

Reports the results of a study on the social and environmental disclosure practices of New Zealand companies. In addition to providing an up‐to‐date description of such practices, and placing them in an international context, examines some potential determinants of these practices. Replicates and extends a recent US study (Patten, 1991). Makes improvements on Patten’s study by measuring the amount of disclosure as a continuous variable using both page amount and the number of sentences. The results indicate both measures are highly correlated. Consistent with Patten (1991) and other studies, reports that both company size and industry are significantly associated with social and environmental disclosures, and that profitability (both current and lagged) is not. In addition to these variables, provides some tentative evidence that New Zealand companies with dual (overseas) stock exchange listings are greater disclosers of social and environmental information.
How calcium inhibits the magnesium‐dependent enzyme human phosphoserine phosphatase
Authors: Peeraer Yves , Rabijns Anja , Collet Jean‐François , Van Schaftingen Emile , De Ranter Camiel
Published: August 2004

The structure of the Mg2+‐dependent enzyme human phosphoserine phosphatase (HPSP) was exploited to examine the structural and functional role of the divalent cation in the active site of phosphatases. Most interesting is the biochemical observation that a Ca2+ ion inhibits the activity of HPSP, even in the presence of added Mg2+. The sixfold coordinated Mg2+ ion present in the active site of HPSP under normal physiological conditions, was replaced by a Ca2+ ion by using a crystallization condition with high concentration of CaCl2 (0.7 m). The resulting HPSP structure now shows a sevenfold coordinated Ca2+ ion in the active site that might explain the inhibitory effect of Ca2+ on the enzyme. Indeed, the Ca2+ ion in the active site captures both side‐chain oxygen atoms of the catalytic Asp20 as a ligand, while a Mg2+ ion ligates only one oxygen atom of this Asp residue. The bidentate character of Asp20 towards Ca2+ hampers the nucleophilic attack of one of the Asp20 side chain oxygen atoms on the phosphorus atom of the substrate phosphoserine.
Ventrally emigrating neural tube (VENT) cells: a second neural tube‐derived cell population
Authors: Dickinson Douglas P. , Machnicki Michal , Ali Mohammed M. , Zhang Zhanying , Sohal Gurkirpal S.
Published: August 2004

Two embryological fates for cells of the neural tube are well established. Cells from the dorsal part of the developing neural tube emigrate and become neural crest cells, which in turn contribute to the development of the peripheral nervous system and a variety of non‐neural structures. Other neural tube cells form the neurons and glial cells of the central nervous system (CNS). This has led to the neural crest being treated as the sole neural tube‐derived emigrating cell population, with the remaining neural tube cells assumed to be restricted to forming the CNS. However, this restriction has not been tested fully. Our investigations of chick, quail and duck embryos utilizing a variety of different labelling techniques (DiI, LacZ, GFP and quail chimera) demonstrate the existence of a second neural tube‐derived emigrating cell population. These cells originate from the ventral part of the cranial neural tube, emigrate at the exit/entry site of the cranial nerves, migrate in association with the nerves and populate their target tissues. On the basis of its site of origin and route of migration we have named this cell population the ventrally emigrating neural tube (VENT) cells. VENT cells also differ from neural crest cells in that they emigrate considerably after the emigration of neural crest cells, and lack expression of the neural crest cell antigen HNK‐1. VENT cells are multipotent, differentiating into cell types belonging to all four basic tissues in the body: the nerve, muscle, connective and epithelium. Thus, the neural tube provides at least two cell populations – neural crest and VENT cells – that contribute to the development of the peripheral nervous system and various non‐neural structures. This review describes the origin of the idea of VENT cells, and discusses evidence for their existence and subsequent fates.
Brassinolide interacts with auxin and ethylene in the root gravitropic response of maize (Zea mays)
Authors: Chang Soo Chul , Kim Young‐Soo , Lee Jin Young , Kaufman Peter B. , Kirakosyan Ara , Yun Hye Sup , Kim Tae‐Wuk , Kim Soon Young , Cho Myeon Haeng , Lee June Seung , Kim Seong‐Ki
Published: August 2004

This study examines how brassinolide (BL) and ethylene interact in the gravitropic response mechanism of maize (Zea mays) primary roots. When applied exogenously, ethylene increases the rate of gravitropic curvature in a dose‐dependent manner. This effect of ethylene was confirmed by the fact that AVG, a specific action inhibitor of ACC synthase, reduces the gravitropic curvature in the presence and absence of BL. Since AVG did not inhibit BL‐increased gravitropic curvature completely, we investigated the possibility that BL may act on the gravitropic response by ways other than simply through enhanced ethylene production. We show that BL exhibits some of its stimulatory effect in the absence of ethylene. In addition, BL reduces the presentation time and lag period for the gravitropic response, whereas ethylene increases them. One possible mechanism of such action is that BL affects protein kinase activity, since the protein kinase inhibitors, staurosporine and H89, reduce BL‐increased gravitropic curvature. In summary, BL is involved in the gravitropic response in maize primary roots via ethylene production, but it acts in a way that differs somewhat from that of ethylene.