bluuebunny/crossref_metadata_2025 huggingface dataset:
Oligonucleotide delivery to tumours using macromolecular carriers
Authors: Dass Crispin R.
Published: September 2004

Microparticles have been used for delivery of chemo‐ and radio‐therapeutic agents to tumours in patients for more than a decade, whereas cationic liposomes have been used for introducing nucleic acids into mammalian cells for the same period of time. This minireview discusses the potential of delivering therapeutic ONs (oligonucleotides) to tumours using cationic liposomes and the major obstacles to such delivery. It also proposes how microparticles can be utilized for delivery of ONs to solid tumours. Prospects for improved ON delivery using these carriers and ways to achieve this are discussed.
Subcellular localization of EEN/endophilin A2, a fusion partner gene in leukaemia
Authors: CHEUNG Ngai , SO Chi Wai , YAM Judy W. P. , SO C. K. C. , POON Randy Y. C. , JIN Dong-Yan , CHAN Li Chong
Published: September 2004

EEN (extra eleven nineteen), also known as EA2 (endophilin A2), a fusion partner of the MLL (mixed-lineage leukaemia) gene in human acute leukaemia, is a member of the endophilin A family, involved in the formation of endocytic vesicles. We present evidence to show that EEN/EA2 is localized predominantly in nuclei of various cell lines of haemopoietic, fibroblast and epithelial origin, in contrast with its reported cytoplasmic localization in neurons and osteoclasts, and that EEN/EA2 exhibits nucleocytoplasmic shuttling. During the cell cycle, EEN/EA2 shows dynamic localization: it is perichromosomal in prometaphase, co-localizes with the bipolar spindle in metaphase and anaphase and redistributes to the midzone and midbody in telophase. This pattern of distribution coincides with changes in protein levels of EEN/EA2, with the highest levels being observed in G2/M-phase. Our results suggest that distinct subcellular localization of the endophilin A family members probably underpins their diverse cellular functions and indicates a role for EEN/EA2 in the cell cycle.
Characterization of N-acetylneuraminic acid synthase isoenzyme 1 from Campylobacter jejuni
Authors: SUNDARAM Appavu K. , PITTS Lee , MUHAMMAD Kamilah , WU Jing , BETENBAUGH Michael , WOODARD Ronald W. , VANN Willie F.
Published: September 2004

Escherichia coli NeuNAc (N-acetylneuraminic acid) synthase catalyses the condensation of PEP (phosphoenolpyruvate) and ManNAc (N-acetylmannosamine) to form NeuNAc and is encoded by the neuB gene. Campylobacter jejuni has three neuB genes, one of which is very similar to the E. coli neuB gene. We have characterized the C. jejuni neuraminic acid synthase with respect to acylamino sugar specificity and stereochemistry of the PEP condensation. We determined the specificity of C. jejuni NeuNAc synthase for N-acetylmannosamine, N-butanoylmannosamine, N-propionoylmannosamine and N-pentanoylmannosamine. We find that, although this enzyme exhibits similar Km values for N-acylmannosamine molecules with different N-acyl groups, the kcat/Km values decreased with increasing chain length. NeuNAc synthase is a member of a PEP-utilizing family of enzymes that form oxo acids from PEP and a monosaccharide. This family includes KDO 8-P (2-keto-3-deoxy-D-manno-octulosonate 8-phosphate) synthase and DAH 7-P (2-keto-3-deoxy-D-arabino-heptulosonate 7-phosphate) synthase. Both enzymes catalyse the condensation of the re face of the aldehyde group of the monosaccharide with the si face of the PEP molecule. The C. jejuni NeuNAc synthase catalysed the condensation of Z- and E-[3-2H]PEP with ManNAc, yielding (3S)-3-deutero-NeuNAc and (3R)-3-deutero-NeuNAc respectively. The condensation of Z-[3-F]PEP and ManNAc yielded (3S)-3-fluoro-NeuNAc. Results of our studies suggest that the C. jejuni NeuNAc synthase, similar to KDO 8-P synthase and DAH 7-P synthase, catalyses the condensation of the si face of PEP with the aldehyde sugar. The present study is the first stereochemical analysis of the reaction catalysed by a bacterial NeuNAc synthase.
Sputa nerve growth factor forms a preferable substitute to mouse 7S-β nerve growth factor
Authors: KOH Dawn C.-I. , ARMUGAM A. , JEYASEELAN K.
Published: September 2004

The NGF (nerve growth factor) from Naja sputatrix has been purified by gel filtration followed by reversed-phase HPLC. The protein showed a very high ability to induce neurite formation in PC12 cells relative to the mouse NGF. Two cDNAs encoding isoforms of NGF have been cloned and an active recombinant NGF, sputa NGF, has been produced in Escherichia coli as a His-tagged fusion protein. Sputa NGF has been found to be non-toxic under both in vivo and in vitro conditions. The induction of neurite outgrowth by this NGF has been found to involve the high-affinity trkA–p75NTR complex of receptors. The pro-survival mechanism of p75NTR has been mediated by the activation of nuclear factor κB gene by a corresponding down-regulation of inhibitory κB gene. Real-time PCR and protein profiling (by surface-enhanced laser-desorption–ionization time-of-flight) have confirmed that sputa NGF up-regulates the expression of the endogenous NGF in PC12 cells. Preliminary microarray analysis has also shown that sputa NGF is capable of promoting additional beneficial effects such as the up-regulation of arginine vasopressin receptor 1A, voltage-dependent T-type calcium channel. Hence, sputa NGF forms a new and useful NGF.
DUK114, the Drosophila orthologue of bovine brain calpain activator protein, is a molecular chaperone
Authors: FARKAS Attila , NARDAI Gábor , CSERMELY Peter , TOMPA Peter , FRIEDRICH Peter
Published: September 2004

UK114, the goat liver tumour antigen, is a member of a widely distributed family of conserved low-molecular-mass proteins (YER057c/YjgF/UK114), the function of which is ill understood. To the various orthologues diverse functions have been ascribed, such as translation inhibition, regulation of purine repressor or calpain activation. Owing to a limited sequence similarity to Hsp90 (heat-shock protein 90), they have also been proposed to be molecular chaperones; however, this has never been tested. In the present paper, we report the cloning and characterization of the Drosophila orthologue, DUK114. In brief, DUK114 had no effect that would have qualified it as a calpain activator. In contrast, it proved to be a very potent molecular chaperone in in vitro assays. In a heat-aggregation test, it significantly decelerated the formation of citrate synthase aggregates. In a reverse assay, the recovery of the enzyme from urea- and heat-induced denatured states was accelerated almost 3-fold. On a molar basis, the chaperone activity of the 15-kDa DUK114 is comparable with that of Hsp90, the almost 6-times-larger archetypal molecular chaperone. In similar assays, DUK114 was ineffective with Drosophila calpain A or calpain B. To test for its chaperone activity in vivo, DUK114 was transfected into Schneider (S2) cells; after heat shock, the number of viable non-transfected cells started to increase after a lag time; in the presence of DUK114, cell proliferation started at once. Our work is the first experimental evidence that DUK114, and possibly other members of this family, are molecular chaperones.
Cyclophilin-D promotes the mitochondrial permeability transition but has opposite effects on apoptosis and necrosis
Authors: LI Yanmin , JOHNSON Nicholas , CAPANO Michela , EDWARDS Mina , CROMPTON Martin
Published: September 2004

Cyclophilin-D is a peptidylprolyl cis–trans isomerase of the mitochondrial matrix. It is involved in mitochondrial permeability transition, in which the adenine nucleotide translocase of the inner membrane is transformed from an antiporter to a non-selective pore. The permeability transition has been widely considered as a mechanism in both apoptosis and necrosis. The present study examines the effects of cyclophilin-D on the permeability transition and lethal cell injury, using a neuronal (B50) cell line stably overexpressing cyclophilin-D in mitochondria. Cyclophilin-D overexpression rendered isolated mitochondria far more susceptible to the permeability transition induced by Ca2+ and oxidative stress. Similarly, cyclophilin-D overexpression brought forward the onset of the permeability transition in intact cells subjected to oxidative stress. In addition, in the absence of stress, the mitochondria of cells overexpressing cyclophilin-D maintained a lower inner-membrane potential than those of normal cells. All these effects of cyclophilin-D overexpression were abolished by cyclosporin A. It is concluded that cyclophilin-D promotes the permeability transition in B50 cells. However, cyclophilin-D overexpression had opposite effects on apoptosis and necrosis; whereas NO-induced necrosis was promoted, NO- and staurosporine-induced apoptosis were inhibited. These findings indicate that the permeability transition leads to cell necrosis, but argue against its involvement in apoptosis.
RLP, a novel Ras-like protein, is an immediate-early transforming growth factor-β (TGF-β) target gene that negatively regulates transcriptional activity induced by TGF-β
Authors: PIEK Ester , van DINTHER Maarten , PARKS W. Tony , SALLEE John M. , BÖTTINGER Erwin P. , ROBERTS Anita B. , ten DIJKE Peter
Published: September 2004

We have described previously the use of microarray technology to identify novel target genes of TGF-β (transforming growth factor-β) signalling in mouse embryo fibroblasts deficient in Smad2 or Smad3 [Yang, Piek, Zavadil, Liang, Xie, Heyer, Pavlidis, Kucherlapati, Roberts and Böttinger (2003) Proc. Natl. Acad. Sci. U.S.A. 100, 10269–10274]. Among the TGF-β target genes identified, a novel gene with sequence homology to members of the Ras superfamily was identified, which we have designated as RLP (Ras-like protein). RLP is a Smad3-dependent immediate-early TGF-β target gene, its expression being induced within 45 min. Bone morphogenetic proteins also induce expression of RLP, whereas epidermal growth factor and phorbol ester PMA suppress TGF-β-induced expression of RLP. Northern-blot analysis revealed that RLP was strongly expressed in heart, brain and kidney, and below the detection level in spleen and skeletal muscles. At the protein level, RLP is approx. 30% homologous with members of the Ras superfamily, particularly in domains characteristic for small GTPases. However, compared with prototypic Ras, RLP contains a modified P-loop, lacks the consensus G2 loop and the C-terminal prenylation site and harbours amino acid substitutions at positions that render prototypic Ras oncogenic. However, RLP does not have transforming activity, does not affect phosphorylation of mitogen-activated protein kinase and is unable to bind GTP or GDP. RLP was found to associate with certain subtypes of the TGF-β receptor family, raising the possibility that RLP plays a role in TGF-β signal transduction. Although RLP did not interact with Smads and did not affect TGF-β receptor-induced Smad2 phosphorylation, it inhibited TGF-β-induced transcriptional reporter activation, suggesting that it is a novel negative regulator of TGF-β signalling.
The SARS coronavirus nucleocapsid protein induces actin reorganization and apoptosis in COS-1 cells in the absence of growth factors
Authors: SURJIT Milan , LIU Boping , JAMEEL Shahid , CHOW Vincent T. K. , LAL Sunil K.
Published: September 2004

In March 2003, a novel coronavirus was isolated from patients exhibiting atypical pneumonia, and was subsequently proven to be the causative agent of the disease now referred to as SARS (severe acute respiratory syndrome). The complete genome of the SARS-CoV (SARS coronavirus) has since been sequenced. The SARS-CoV nucleocapsid (SARS-CoV N) protein shares little homology with other members of the coronavirus family. In the present paper, we show that SARS-CoV N is capable of inducing apoptosis of COS-1 monkey kidney cells in the absence of growth factors by down-regulating ERK (extracellular-signal-regulated kinase), up-regulating JNK (c-Jun N-terminal kinase) and p38 MAPK (mitogen-activated protein kinase) pathways, and affecting their downstream effectors. SARS-CoV N expression also down-regulated phospho-Akt and Bcl-2 levels, and activated caspases 3 and 7. However, apoptosis was independent of the p53 and Fas signalling pathways. Furthermore, activation of the p38 MAPK pathway was found to induce actin reorganization in cells devoid of growth factors. At the cytoskeletal level, SARS-CoV N down-regulated FAK (focal adhesion kinase) activity and also down-regulated fibronectin expression. This is the first report showing the ability of the N protein of SARS-CoV to induce apoptosis and actin reorganization in mammalian cells under stressed conditions.
Association study of the I/D polymorphism and plasma angiotensin-converting enzyme (ACE) as risk factors for stent restenosis
Authors: RIBICHINI Flavio , FERRERO Valeria , MATULLO Giuseppe , FEOLA Mauro , VADO Antonello , CAMILLA Terenzio , GUARRERA Simonetta , CARTURAN Sonia , VASSANELLI Corrado , USLENGHI Eugenio , PIAZZA Alberto
Published: September 2004

The ID (insertion/deletion) polymorphism of the ACE (angiotensin-converting enzyme) gene controls plasma ACE levels. Both have been correlated with ISR (in-stent restenosis) in preliminary analyses, but not confirmed in larger studies. In the present study, baseline and 6-month quantitative coronary analysis were performed in 897 patients who had stent implantation and the ID polymorphism genotyped. Plasma ACE levels were measured in 848 patients (95%). Restenosis rates among genotypes were 31.2% DD, 25.5% ID and 28.8% II (not significant). Plasma ACE levels were significantly higher in restenotic patients compared with patients without restenosis (30.7±18.6 units/l compared with 22.8±12.8 units/l; P=0.0001) and a strong independent predictor of ISR [OR (odds ratio)=3.70; 95% CI (confidence interval), 2.40–5.71; P<0.0001], except in diabetics. In the subgroup of diabetics and patients with AMI (acute myocardial infarction), the DD genotypes actually had a lower risk of ISR than the II genotypes (diabetics, OR=0.16; 95% CI, 0.04–0.69; P=0.014; and patients with AMI, OR=0.21; 95% CI, 0.061–0.749; P=0.016). After exclusion of diabetics and patients with AMI, ISR rates for genotypes in 632 patients were 31.7% DD, 24.3% ID and 17.6% II (P=0.02; DD compared with non-DD OR=1.57; 95% CI, 1.09–2.25). The association between the D allele and ISR observed in selected populations does not hold with a larger sample size. Other than sample size, clinical variables can modulate the association between ID polymorphism and ISR. Plasma ACE level is a risk factor for ISR, independently of the ID genotype.
Invasive measurements of pulse wave velocity correlate with the degree of aortic valve calcification and severity associated with matrix metalloproteinases in elderly patients with aortic valve stenosis
Authors: LIU Ping-Yen , TSAI Wei-Chuan , LIN Chih-Chan , HSU Chih-Hsin , HAUNG Yao-Yi , CHEN Jyh-Hong
Published: September 2004

The aim of the present study was to assess the association between invasive PWV (pulse wave velocity), serum levels of MMPs (matrix metalloproteinases) and the echocardiographic severity and calcification score of degenerative AS (aortic stenosis). We enrolled 30 patients (16 males; age, 61.3±8.2 years) diagnosed with degenerative AS and an additional 30 age- and sex-matched control patients. Invasive PWV methods with a pigtail catheter and double-channel recording were performed in both groups in our catheterization laboratory. We scored the severity of calcification at the AV (aortic valve) during two-dimensional echocardiography. The association between the trans-valvular pressure gradient, the severity of calcification of the AV and the value of PWV were analysed. We also analysed the serum levels of MMP-9, MMP-3 and TIMP-1 (tissue inhibitor of metalloproteinases-1) in these patients. In the group with degenerative AS, mean AV pressure gradients (56.0 compared with 9.5 mmHg; P<0.001) and calcified AV scoring (3.3±1.2 compared with 2.1±0.9; P<0.001) were higher than in the control group. In addition, PWV was faster in the group with degenerative AS (15.5±3.8 compared with 8.0±2.7 m/s; P=0.001). After being adjusted for age, sex, mean blood pressure and left ventricular function, both the AV pressure gradient and the severity of calcification were strongly correlated with PWV (R=0.706, P<0.0001, and R=0.561, P=0.03 respectively). In addition, the serum levels of MMP-9, MMP-3 and TIMP-1 were all significantly higher and correlated with PWV in the group with AS (all P<0.05). With higher serum levels of MMPs and their inhibitors, we found that this invasive measurement of PWV was associated strongly with the pressure gradient and calcification of AV. More advanced degenerative changes in AV was probably associated with more severe aortic arteriosclerosis.
Carotid artery stiffening does not explain baroreflex impairment in pre-eclampsia
Authors: MERSICH Beatrix , RIGÓ János , LÉNÁRD Zsuzsanna , STUDINGER Péter , VISONTAI Zsuzsanna , KOLLAI Mark
Published: September 2004

Stiffening of the barosensory vessel wall in hypertension has been suggested to play a role in the associated baroreflex impairment. The carotid distensibility–BRS (baroreflex sensitivity) relationship, however, has not been studied in pre-eclampsia, a condition where hypertension is spontaneously reversible. Twelve normotensive pregnant women and 12 patients with pre-eclampsia matched for maternal age and week of gestation were studied in the third trimester and 3 months postpartum. Carotid artery diastolic diameter and pulsatile distension was measured by echo-wall tracking and carotid pulse pressure by applanation tonometry, and the carotid distensibility coefficient was calculated. Spontaneous BRS was determined by the sequence and spectral methods from 10 min continuous recording of ECG and finger arterial blood pressure. In the third trimester, carotid distensibility was lower in patients with pre-eclampsia than in normotensive pregnant women (2.47±0.17 compared with 4.08±0.16×10−3/mmHg); postpartum, it increased moderately in patients, but remained below normotensive values (3.25±0.12 compared with 4.25±0.19×10−3/mmHg). In the third trimester, both patients and healthy pregnant women had equally low BRS values; postpartum, the various BRS indices increased markedly (by 60–190%) and to the same level in both groups. No correlation was found between changes in carotid artery distensibility and those in BRS from the third trimester to postpartum period in patients and healthy pregnant women. The lack of association between changes in carotid distensibility and BRS suggest that stiffening of the carotid artery in pre-eclampsia is not responsible for baroreflex dysfunction.
Identification of a SUMO-binding motif that recognizes SUMO-modified proteins
Authors: Song Jing , Durrin Linda K. , Wilkinson Thomas A. , Krontiris Theodore G. , Chen Yuan
Published: September 2004

Posttranslational modification by the ubiquitin homologue, small ubiquitin-like modifier 1 (SUMO-1), has been established as an important regulatory mechanism. However, in most cases it is not clear how sumoylation regulates various cellular functions. Emerging evidence suggests that sumoylation may play a general role in regulating protein-protein interactions, as shown in RanBP2/Nup358 and RanGAP1 interaction. In this study, we have defined an amino acid sequence motif that binds SUMO. This motif, V/I-X-V/I-V/I, was identified by NMR spectroscopic characterization of interactions among SUMO-1 and peptides derived from proteins that are known to bind SUMO or sumoylated proteins. This motif binds all SUMO paralogues (SUMO-1-3). Using site-directed mutagenesis, we also show that this SUMO-binding motif in RanBP2/Nup358 is responsible for the interaction between RanBP2/Nup358 and sumoylated RanGAP1. The SUMO-binding motif exists in nearly all proteins known to be involved in SUMO-dependent processes, suggesting its general role in sumoylation-dependent cellular functions.
Different relaxations in myoglobin after photolysis
Authors: Levantino Matteo , Cupane Antonio , Zimányi László , Ormos Pál
Published: September 2004

To clarify the interplay of kinetic hole-burning (KHB), structural relaxation, and ligand migration in myoglobin (Mb), we measured time-resolved absorption spectra in the Soret region after photolysis of carbon monoxide Mb (MbCO) in the temperature interval 120-260 K and in the time window 350 ns to 200 ms. The spectral contributions of both photolyzed (Mb
*
) and liganded Mb (MbCO) have been analyzed by taking into account homogeneous bandwidth, coupling to vibrational modes, and static conformational heterogeneity. We succeeded in separating the “time-dependent” spectral changes, and this work provides possibilities to identify the events in the process of ligand rebinding. KHB is dominant at
T
<190 K in both the Mb
*
and the MbCO components. For MbCO, conformational substates interconversion at higher temperatures tends to average out the KHB effect. At 230-260 K, whereas almost no shift is observed in the MbCO spectrum, a shift of the order of ≈80 cm
-1
is observed in Mb
*
. We attribute this shift to protein relaxation coupled to ligand migration. The time dependence of the Mb
*
spectral shift is interpreted with a model that enables us to calculate the highly nonexponential relaxation kinetics. Fits of stretched exponentials to this kinetics yield Kohlrausch parameter values of 0.25, confirming the analogy between proteins and glasses.
Sequence composition and genome organization of maize
Authors: Messing Joachim , Bharti Arvind K. , Karlowski Wojciech M. , Gundlach Heidrun , Kim Hye Ran , Yu Yeisoo , Wei Fusheng , Fuks Galina , Soderlund Carol A. , Mayer Klaus F. X. , Wing Rod A.
Published: September 2004

Zea mays
L. ssp.
mays
, or corn, one of the most important crops and a model for plant genetics, has a genome ≈80% the size of the human genome. To gain global insight into the organization of its genome, we have sequenced the ends of large insert clones, yielding a cumulative length of one-eighth of the genome with a DNA sequence read every 6.2 kb, thereby describing a large percentage of the genes and transposable elements of maize in an unbiased approach. Based on the accumulative 307 Mb of sequence, repeat sequences occupy 58% and genic regions occupy 7.5%. A conservative estimate predicts ≈59,000 genes, which is higher than in any other organism sequenced so far. Because the sequences are derived from bacterial artificial chromosome clones, which are ordered in overlapping bins, tagged genes are also ordered along continuous chromosomal segments. Based on this positional information, roughly one-third of the genes appear to consist of tandemly arrayed gene families. Although the ancestor of maize arose by tetraploidization, fewer than half of the genes appear to be present in two orthologous copies, indicating that the maize genome has undergone significant gene loss since the duplication event.
Spindle Pole Body Duplication in Fission Yeast Occurs at the G1/S Boundary but Maturation Is Blocked until Exit from S by an Event Downstream ofCdc10+
Authors: Uzawa Satoru , Li Fei , Jin Ye , McDonald Kent L. , Braunfeld Michael B. , Agard David A. , Cande W. Zacheus
Published: September 2004

The regulation and timing of spindle pole body (SPB) duplication and maturation in fission yeast was examined by transmission electron microscopy. When cells are arrested at G1 by nitrogen starvation, the SPB is unduplicated. On release from G1, the SPBs were duplicated after 1–2 h. In cells arrested at S by hydroxyurea, SPBs are duplicated but not mature. In G1 arrest/release experiments with cdc2.33 cells at the restrictive temperature, SPBs remained single, whereas in cells at the permissive temperature, SPBs were duplicated. In cdc10 mutant cells, the SPBs seem not only to be duplicated but also to undergo partial maturation, including invagination of the nuclear envelope underneath the SPB. There may be an S-phase–specific inhibitor of SPB maturation whose expression is under control of cdc10+. This model was examined by induction of overreplication of the genome by overexpression of rum1p or cdc18p. In cdc18p-overexpressing cells, the SPBs are duplicated but not mature, suggesting that cdc18p is one component of this feedback mechanism. In contrast, cells overexpressing rum1p have large, deformed SPBs accompanied by other features of maturation and duplication. We propose a feedback mechanism for maturation of the SPB that is coupled with exit from S to trigger morphological changes.
Endocytosis and Sorting of ErbB2 and the Site of Action of Cancer Therapeutics Trastuzumab and Geldanamycin
Authors: Austin Cary D. , De Mazière Ann M. , Pisacane Paul I. , van Dijk Suzanne M. , Eigenbrot Charles , Sliwkowski Mark X. , Klumperman Judith , Scheller Richard H.
Published: September 2004

ErbB2 is a transmembrane tyrosine kinase whose surface overexpression is linked to tumorigenesis and poor prognosis in breast cancer patients. Two models have emerged that account for the high surface distribution of ErbB2. In one model, the surface pool is dynamic and governed by a balance between endocytosis and recycling, whereas in the other it is retained, static, and excluded from endocytosis. These models have contrasting implications for how ErbB2 exerts its biological function and how cancer therapies might down-regulate surface ErbB2, such as the antibody trastuzumab (Herceptin) or the Hsp90 inhibitor geldanamycin. Little is known, however, about how these treatments affect ErbB2 endocytic trafficking. To investigate this issue, we examined breast carcinoma cells by immunofluorescence and quantitative immunoelectron microscopy and developed imaging and trafficking kinetics assays using cell surface fluorescence quenching. Surprisingly, trastuzumab does not influence ErbB2 distribution but instead recycles passively with internalized ErbB2. By contrast, geldanamycin down-regulates surface ErbB2 through improved degradative sorting in endosomes exclusively rather than through increased endocytosis. These results reveal substantial dynamism in the surface ErbB2 pool and clearly demonstrate the significance of endosomal sorting in the maintenance of ErbB2 surface distribution, a critical feature of its biological function.
Stepwise detection of recombination breakpoints in sequence alignments
Authors: Graham Jinko , McNeney Brad , Seillier-Moiseiwitsch Françoise
Published: September 2004

Motivation: We propose a stepwise approach to identify recombination breakpoints in a sequence alignment. The approach can be applied to any recombination detection method that uses a permutation test and provides estimates of breakpoints.

Results: We illustrate the approach by analyses of a simulated dataset and alignments of real data from HIV-1 and human chromosome 7. The presented simulation results compare the statistical properties of one-step and two-step procedures. More breakpoints are found with a two-step procedure than with a single application of a given method, particularly for higher recombination rates. At higher recombination rates, the additional breakpoints were located at the cost of only a slight increase in the number of falsely declared breakpoints. However, a large proportion of breakpoints still go undetected.

Availability: A makefile and C source code for phylogenetic profiling and the maximum χ2 method, tested with the gcc compiler on Linux and WindowsXP, are available at http://stat-db.stat.sfu.ca/stepwise/

Contact:  jgraham@stat.sfu.ca
Akkermansia muciniphila gen. nov., sp. nov., a human intestinal mucin-degrading bacterium
Authors: Derrien Muriel , Vaughan Elaine E. , Plugge Caroline M. , de Vos Willem M.
Published: September 2004

The diversity of mucin-degrading bacteria in the human intestine was investigated by combining culture and 16S rRNA-dependent approaches. A dominant bacterium, strain MucT, was isolated by dilution to extinction of faeces in anaerobic medium containing gastric mucin as the sole carbon and nitrogen source. A pure culture was obtained using the anaerobic soft agar technique. Strain MucTwas a Gram-negative, strictly anaerobic, non-motile, non-spore-forming, oval-shaped bacterium that could grow singly and in pairs. When grown on mucin medium, cells produced a capsule and were found to aggregate. Strain MucTcould grow on a limited number of sugars, includingN-acetylglucosamine,N-acetylgalactosamine and glucose, but only when a protein source was provided and with a lower growth rate and final density than on mucin. The G+C content of DNA from strain MucTwas 47·6 mol%. 16S rRNA gene sequence analysis revealed that the isolate was part of the divisionVerrucomicrobia. The closest described relative of strain MucTwasVerrucomicrobium spinosum(92 % sequence similarity). Remarkably, the 16S rRNA gene sequence of strain MucTshowed 99 % similarity to three uncultured colonic bacteria. According to the data obtained in this work, strain MucTrepresents a novel bacterium belonging to a new genus in subdivision 1 of theVerrucomicrobia; the nameAkkermansia muciniphilagen. nov., sp. nov. is proposed; the type strain is MucT(=ATCC BAA-835T=CIP 107961T).
Mycobacterium psychrotolerans sp. nov., isolated from pond water near a uranium mine
Authors: Trujillo Martha E. , Velázquez Encarna , Kroppenstedt Reiner M. , Schumann Peter , Rivas Raúl , Mateos Pedro F. , Martínez-Molina Eustoquio
Published: September 2004

An acid-fast, rapidly growing, psychrotolerant short rod was isolated from pond water near a uranium mine. Phylogenetic analysis of the 16S rRNA gene sequence grouped this strain with the rapidly growing mycobacteria. The 16S rRNA gene sequence of isolate WA101Tshowed highest similarity to that ofMycobacterium sphagniDSM 44076T; however, DNA–DNA relatedness between the two strains was less than 30 %. Chemotaxonomic analyses, which included fatty acid and mycolic acid patterns, confirmed the classification of strain WA101Tin the genusMycobacterium. Physiological data, including antibiotic resistance, NaCl tolerance, carbon sources, temperature growth range and enzyme activities, were also determined. Based on the genotypic and phenotypic results it is proposed that isolate WA101Trepresents a novelMycobacteriumspecies. The nameMycobacterium psychrotoleranssp. nov. is proposed, with type strain WA101T(=DSM 44697T=LMG 21953T).
Desulfovibrio bastinii sp. nov. and Desulfovibrio gracilis sp. nov., moderately halophilic, sulfate-reducing bacteria isolated from deep subsurface oilfield water
Authors: Magot Michel , Basso Odile , Tardy-Jacquenod Christèle , Caumette Pierre
Published: September 2004

Two moderately halophilic, mesophilic, sulfate-reducing bacteria were isolated from production-water samples from Emeraude Oilfield, Congo. Motile, vibrioid cells of SRL4225T grew optimally at a concentration of 4 % NaCl, at pH 5·8–6·2, with a minimal pH for growth of 5·2, showing that it is a moderately acidophilic bacterium. Cells of SRL6146T were motile, curved or vibrioid, long and thin rods. Optimal growth was obtained at a concentration of 5–6 % NaCl, at pH 6·8–7·2. The nutritional requirements showed that many of the characteristics of these strains overlap with those of known Desulfovibrio species. On the basis of 16S rRNA gene sequence analysis and DNA–DNA hybridization studies, both strains are members of the genus Desulfovibrio. However, they are not closely related to any species of the genus that have validly published names. It is therefore proposed that the two strains are members of two novel species of the genus Desulfovibrio with the names Desulfovibrio bastinii sp. nov. (type strain SRL4225T=DSM 16055T=ATCC BAA-903T) and Desulfovibrio gracilis sp. nov. (type strain SRL6146T=DSM 16080T=ATCC BAA-904T).
Diversity and evolution of Bdellovibrio-and-like organisms (BALOs), reclassification of Bacteriovorax starrii as Peredibacter starrii gen. nov., comb. nov., and description of the Bacteriovorax–Peredibacter clade as Bacteriovoracaceae fam. nov.
Authors: Davidov Yaacov , Jurkevitch Edouard
Published: September 2004

A phylogenetic analysis of Bdellovibrio-and-like organisms (BALOs) was performed. It was based on the characterization of 71 strains and on all consequent 16S rRNA gene sequences available in databases, including clones identified by data-mining, totalling 120 strains from very varied biotopes. Amplified rDNA restriction analysis (ARDRA) accurately reflected the diversity and phylogenetic affiliation of BALOs, thereby providing an efficient screening tool. Extensive phylogenetic analysis of the 16S rRNA gene sequences revealed great diversity within the Bdellovibrio (>14 % divergence) and Bacteriovorax (>16 %) clades, which comprised nine and eight clusters, respectively, exhibiting more than 3 % intra-cluster divergence. The clades diverged by more than 20 %. The analysis of conserved 16S rRNA secondary structures showed that Bdellovibrio contained motifs atypical of the δ-Proteobacteria, suggesting that it is ancestral to Bacteriovorax. While none of the Bdellovibrio strains were of marine origin, Bacteriovorax included separate soil/freshwater and marine-specific groups. On the basis of their extensive diversity and the large distance separating the groups, it is proposed that Bacteriovorax starrii be placed into a new genus, Peredibacter gen. nov., with Peredibacter starrii A3.12T (=ATCC 15145T=NCCB 72004T) as its type strain. Also proposed is a redefinition of the Bdellovibrio and the Bacteriovorax–Peredibacter lineages as two different families, i.e. ‘Bdellovibrionaceae’ and a new family, Bacteriovoracaceae. Also, a re-evaluation of oligonucleotides targeting BALOs is presented, and the implications of the large diversity of these organisms and of their distribution in very different environments are discussed.
Colwellia piezophila sp. nov., a novel piezophilic species from deep-sea sediments of the Japan Trench
Authors: Nogi Yuichi , Hosoya Shoichi , Kato Chiaki , Horikoshi Koki
Published: September 2004

Two strains of obligately piezophilic bacteria were isolated from sediment collected from the bottom surface of a small canyon on the seaward slope of the Japan Trench at a depth of 6278 m. The isolated strains, Y223GTand Y251E, are closely affiliated with members of the genusColwelliaon the basis of 16S rRNA gene sequence analysis. The G+C contents of both strains were about 39 mol%. DNA–DNA hybridization values between these strains andColwelliareference strains were significantly lower than those accepted as the phylogenetic definition of a species. The novel strains are Gram-negative, polarly flagellated and facultatively anaerobic. The optimal pressure for growth was 60 MPa at both 4 and 10 °C; the most rapid growth rate was observed at 10 °C and 60 MPa. No growth occurred at 15 °C under any pressure studied. The major isoprenoid quinone is Q-8. The predominant cellular fatty acids are C16 : 0 and C16 : 1. Based on the taxonomic differences observed, the isolated strains appear to represent a novel obligately piezophilicColwelliaspecies. The nameColwellia piezophilasp. nov. (type strain Y223GT=JCM 11831T=ATCC BAA-637T) is proposed.
Characterization of Pseudoalteromonas distincta-like sea-water isolates and description of Pseudoalteromonas aliena sp. nov.
Authors: Ivanova Elena P. , Gorshkova Nataliya M. , Zhukova Natalia V. , Lysenko Anatolii M. , Zelepuga Elena A. , Prokof’eva Nina G. , Mikhailov Valery V. , Nicolau Dan V. , Christen Richard
Published: September 2004

Seven melanogenicPseudoalteromonas distincta-like strains, KMM 3562T, KMM 3536, KMM 3537, KMM 3538, KMM 3539, KMM 3615 and KMM 3629, which expressed tyrosinases were isolated from sea-water samples collected from different locations in Amursky Bay (Sea of Japan, Pacific Ocean) and characterized to clarify their taxonomic position. By 16S rRNA gene sequence analysis, the bacteria were shown to belong to the genusPseudoalteromonas. The G+C content of the DNAs of the strains was 41–43 mol%. The level of DNA similarity among these strains was conspecific (92–97 %), indicating that they represented a single genospecies. However, DNA from the strains isolated from sea water showed only 63–65 % genetic relatedness with the DNA of the type strainP. distincta. The novel organisms grew mainly between 4 and 30 °C, were neutrophilic and slightly halophilic (four strains had a narrow range of growth between 3 and 6 % NaCl, w/v), were haemolytic and cytotoxic and were able to degrade starch, gelatin and Tween 80. The predominant fatty acids, including 16 : 0, 16 : 1ω7, 17 : 1ω7 and 18 : 1ω7, were typical of the genusPseudolateromonas. The phylogenetic, genetic and physiological properties of the seven strains placed them within a novel species,Pseudoalteromonas alienasp. nov., the type strain of which is SW19T(=KMM 3562T=LMG 22059T).
Psychrobacter maritimus sp. nov. and Psychrobacter arenosus sp. nov., isolated from coastal sea ice and sediments of the Sea of Japan
Authors: Romanenko Lyudmila A. , Lysenko Anatoly M. , Rohde Manfred , Mikhailov Valery V. , Stackebrandt Erko
Published: September 2004

Four strains of Gram-negative, aerobic, psychrotolerant, non-motile, non-pigmented bacteria were isolated from coastal sea-ice and sediment samples. These strains displayed the general chemotaxonomic and phenotypic features of members of the genusPsychrobacter. 16S rRNA gene sequencing positioned the three isolates KMM 3646T, KMM 3643 and KMM 3645 and isolate KMM 3659Tin two distinct lineages within the genusPsychrobacter, displaying less than 98·5 % sequence similarity to the type strains of otherPsychrobacterspecies. Genomic distinctness was supported by phenotypic differences in growth temperatures, salinity range for growth and metabolic properties. Based on a combination of phenotypic and biochemical characteristics and phylogenetic position, it is proposed that the members of these two distinct lineages represent two novel species, for which the namesPsychrobacter maritimussp. nov. (type strain Pi2-20T=KMM 3646T=DSM 15387T) andPsychrobacter arenosussp. nov. (type strain R7T=KMM 3659T=DSM 15389T) are proposed.
Rubrobacter taiwanensis sp. nov., a novel thermophilic, radiation-resistant species isolated from hot springs
Authors: Chen Mao-Yen , Wu Shih-Hsiung , Lin Guang-Huey , Lu Chun-Ping , Lin Yung-Ting , Chang Wen-Chang , Tsay San-San
Published: September 2004

Two novel bacteria, with an optimum growth temperature of approximately 60 °C, were isolated from Lu-shan hot springs in the central region of Taiwan. These isolates were aerobic, thermophilic, halotolerant, pink-pigmented, heterotrophic and resistant to gamma-radiation. Both pleomorphic, short, rod-shaped cells and coccoid cells were observed. Strains LS-286 (=ATCC BAA-452=BCRC 17198) and LS-293T(=ATCC BAA-406T=BCRC 17173T) represented a novel species of the genusRubrobacter, according to a phylogenetic analysis of the 16S rRNA gene, DNA–DNA hybridization, biochemical features and fatty acid composition. The nameRubrobacter taiwanensissp. nov. is proposed for this novel species, with LS-293Tas the type strain.
Duganella violaceinigra sp. nov., a novel mesophilic bacterium isolated from forest soil
Authors: Li Wen-Jun , Zhang Yu-Qin , Park Dong-Jin , Li Chang-Tian , Xu Li-Hua , Kim Chang-Jin , Jiang Cheng-Lin
Published: September 2004

A mesophilic bacterium, designated strain YIM 31327T, was isolated from a forest soil sample collected from Yunnan Province, China, and was then investigated using a polyphasic approach. The strain grew optimally at 28–30 °C and pH 7·2. The cells were Gram-negative, short, rod-shaped, motile and non-spore-forming with flagella. The major ubiquinone was Q-8 and the cellular fatty acids were C16 : 0 and C12 : 0. The DNA G+C content of strain YIM 31327T was 62·8 mol%. Phylogenetic analysis revealed that strain YIM 31327T was a member of the β-Proteobacteria, being most closely related to Duganella zoogloeoides, with which it exhibited less than 96 % 16S rRNA gene sequence similarity. On the basis of the phenotypic and genotypic differences between strain YIM 31327T and D. zoogloeoides, a novel species, Duganella violaceinigra sp. nov., is proposed, with YIM 31327T (=CIP 108077T=KCTC 12193T) as the type strain.
Paracoccus haeundaensis sp. nov., a Gram-negative, halophilic, astaxanthin-producing bacterium
Authors: Lee Jae Hyung , Kim Yun Sook , Choi Tae-Jin , Lee Won Jae , Kim Young Tae
Published: September 2004

An aerobic, non-motile, Gram-negative, orange-pigmented, rod-shaped, astaxanthin-producing marine bacterium was isolated from the Haeundae Coast, Korea. This strain, BC74171T, produced carotenoids, mainly astaxanthin. All the type strains of the genus Paracoccus were compared with strain BC74171T using 16S rRNA gene sequence analysis, fatty acid patterns and physiological reaction profiles. Based on the results of these analyses, it is proposed that strain BC74171T represents a novel species, Paracoccus haeundaensis sp. nov. The type strain is BC74171T (=KCCM 10460T=LMG P-21903T).
Degradation of hDlg and MAGIs by human papillomavirus E6 is E6-AP-independent
Authors: Grm Helena Sterlinko , Banks Lawrence
Published: September 2004

An important characteristic of the E6 proteins derived from cancer-associated human papillomaviruses (HPVs) is their ability to target cellular proteins for ubiquitin-mediated degradation. Degradation of the p53 tumour suppressor protein by E6 is known to involve the cellular ubiquitin ligase, E6-AP; however, it is presently not known how E6 targets theDrosophiladiscs large (Dlg) tumour suppressor and the membrane-associated guanylate kinase inverted (MAGI) family of proteins for degradation. By using anin vitroE6-AP immunodepletion assay, these targets were tested for degradation in a E6-AP-dependent manner. The data showed clearly that E6 can direct the degradation of Dlg and the MAGI family of proteins in the absence of E6-AP in thisin vitrosystem. These results provide compelling evidence for the role of E6-associated ubiquitin ligases other than E6-AP in the degradation of certain E6 targets.
Analysis of the medium (M) segment sequence of Guaroa virus and its comparison to other orthobunyaviruses
Authors: Briese Thomas , Rambaut Andrew , Lipkin W. Ian
Published: September 2004

Guaroa virus(GROV), a segmented virus in the genusOrthobunyavirus, has been linked to the Bunyamwera serogroup (BUN) through cross-reactivity in complement fixation assays of S segment-encoded nucleocapsid protein determinants, and also to the California serogroup (CAL) through cross-reactivity in neutralization assays of M segment-encoded glycoprotein determinants. Phylogenetic analysis of the S-segment sequence supported a closer relationship to the BUN serogroup for this segment and it was hypothesized that the serological reaction may indicate genome-segment reassortment. Here, cloning and sequencing of the GROV M segment are reported. Sequence analysis indicates an organization similar to that of other orthobunyaviruses, with genes in the order Gn–nsm–gc, and mature proteins generated by protease cleavage at one, and by signalase at possibly three, sites. A potential role of motifs that are more similar to CAL than to BUN virus sequences with respect to the serological reaction is discussed. No discernable evidence for reassortment was identified.
Downregulation of major histocompatibility complex class I in bovine papillomas
Authors: Araibi E. H. , Marchetti B. , Ashrafi G. H. , Campo M. S.
Published: September 2004

Bovine papillomavirus (BPV) induces papillomas in cattle; in the great majority of cases, these regress due to the host immune response, but they can persist and progress to malignancy. Even in the absence of malignant transformation, BPV infection persists for a significant period of time before activation of the host immune system, suggesting that the host immune system is unaware of, or disabled by, BPV. E5 is the major oncoprotein of BPV, which, in addition to its transforming properties, downregulates the expression and transport to the cell surface of major histocompatibility complex class I (MHC I). Here, it is shown that co-expression of MHC I and E5 in papillomas caused by BPV-4 infection is mutually exclusive, in agreement with the inhibition of surface MHC I expression by E5 that is observed in vitro. The inhibition of MHC expression in E5-expressing papilloma cells could explain the long period that is required for activation of the immune response and has implications for the progression of papillomas to the malignant stage; absence of peptide presentation by MHC I to cytotoxic T lymphocytes would allow the infected cells to evade the host cellular immune response and allow the lesions to persist.
Genetic and phenotypic variations of isolates of shrimp Taura syndrome virus found in Penaeus monodon and Metapenaeus ensis in Taiwan
Authors: Chang Yun-Shiang , Peng Shao-En , Yu Hon-Tsen , Liu Feng-Chuan , Wang Chung-Hsiung , Lo Chu-Fang , Kou Guang-Hsiung
Published: September 2004

Distinct Taura syndrome virus (TSV) isolates were found in Metapenaeus ensis (isolate Tw2KMeTSV), Penaeus monodon (isolate Tw2KPmTSV) and Litopenaeus vannamei (isolate Tw02LvTSV). Nucleotide sequence analysis of these three isolates revealed differences in the TSV structural protein (capsid protein precursor) gene orf2. TSV ORF2 amino acid sequence comparison and phylogenetic analysis suggested a comparatively close relationship between these three Taiwanese isolates and the Hawaiian isolate HI94TSV. In P. monodon specimens that were naturally and experimentally infected with the Tw2KPmTSV isolate, the virus was contained and shrimps showed no clinical signs of infection. However, when P. monodon was challenged with the Tw2KMeTSV isolate, the virus replicated freely. The ORF2 amino acid sequence of the Tw2KMeTSV isolate differed from that of isolate Tw2KPmTSV in four positions and these differences may account for their phenotypic differences, at least in terms of their ability to replicate in specific hosts.
Selection pressures in the capsid genes of plant RNA viruses reflect mode of transmission
Authors: Chare Elizabeth R. , Holmes Edward C.
Published: September 2004

To determine the selection pressures faced by RNA viruses of plants, patterns of nonsynonymous (dN) and synonymous (dS) substitution in the capsid genes of 36 viruses with differing modes of transmission were analysed. This analysis provided strong evidence that the capsid proteins of vector-borne plant viruses are subject to greater purifying selection on amino acid change than those viruses transmitted by other routes and that virus–vector interactions impose greater selective constraints than those between virus and plant host. This could be explained by specific interactions between capsid proteins and cellular receptors in the insect vectors that are necessary for successful transmission. However, contrary to initial expectations based on phylogenetic relatedness, vector-borne plant viruses are subject to weaker selective constraints than vector-borne animal viruses. The results suggest that the greater complexity involved in the transmission of circulative animal viruses compared with non-circulative plant viruses results in more intense purifying selection.