bluuebunny/crossref_metadata_2025 huggingface dataset:
The effect of copolymer composition on the dynamics of random copolymers in a homopolymer matrix
Authors: Kamath Sudesh Y. , Dadmun Mark D.
Published: September 2006

The effect of copolymer composition on the dynamics of random copolymers in a homopolymer matrix is studied using computer simulations within the framework of the bond-fluctuation model on blends containing low concentrations (10%) of A-B copolymers, where A and B are two different types of monomers, dispersed in a homopolymer matrix of chains with only A-type monomers. Four copolymer compositions were studied, ϕA=0.33, ϕA=0.5, ϕA=0.66, and ϕA=0.82, while maintaining a statistically random sequence distribution. For this study, we have only included intermolecular interactions between A and B monomers. Our results indicate, in agreement with experimental data, that copolymer composition has an impact on system dynamics. Analysis of the structure reveals that copolymers with majority A content are expanded in the homopolymer matrix, have fewer interchain copolymer-copolymer contacts, and are well dispersed in the homopolymer matrix. On the other hand, copolymers with lower A content form a more compact structure, have more interchain contacts, and form aggregates that are short lived. This in turn leads to slower system dynamics. Both the radius of gyration (Rg) and copolymer end-to-end vectors (Re) increase with increasing A content until ϕA=0.66 and then decrease. Copolymers with lower A content form more compact structures as the repulsive interactions between unlike species are minimized by the copolymers folding back on themselves and forming aggregates of copolymer chains. Thus, these results provide insight into the variation of copolymer dynamics with composition in the system by documenting the correlation between the thermodynamics of this mixture, the conformation of a copolymer chain in a homopolymer matrix, and the dynamics of both components in this blend.
Friction laws for lubricated nanocontacts
Authors: Buzio R. , Boragno C. , Valbusa U.
Published: September 2006

We have used friction force microscopy to probe friction laws for nanoasperities sliding on atomically flat substrates under controlled atmosphere and liquid environment, respectively. A power law relates friction force and normal load in dry air, whereas a linear relationship, i.e., Amontons’ law, is observed for junctions fully immersed in model lubricants, namely, octamethylciclotetrasiloxane and squalane. Lubricated contacts display a remarkable friction reduction, with liquid and substrate specific friction coefficients. Comparison with molecular dynamics simulations suggests that load-bearing boundary layers at junction entrance cause the appearance of Amontons’ law and impart atomic-scale character to the sliding process; continuum friction models are on the contrary of limited predictive power when applied to lubrication effects. An attempt is done to define general working conditions leading to the manifestation of nanoscale lubricity due to adsorbed boundary layers.
Kinetics and dynamic properties of equilibrium polymers
Authors: Huang C.-C. , Xu H. , Ryckaert J.-P.
Published: September 2006

The statistical mechanics and scission-recombination mechanism of self-assembling linear micelles are investigated by Brownian dynamics using a newly proposed mesoscopic model representing the micelles as equilibrium polymer chains. A semidilute concentration regime, yet dynamically unentangled, is considered over a wide range of scission/recombination rates. We focus on the analysis of short and long time behaviors of the scission and recombination mechanisms. Our results show that at time scales larger than the life time of the average chain length, the kinetics is in agreement with the mean-field kinetic model proposed by Cates and Candau [J. Phys.: Condens. Matter 2, 6869 (1990)] provided the kinetic constants are estimated as effective ones. These values do take into account through a transmission coefficient that a fraction of scission/recombination events is correlated over a short time (diffusion controlled mechanism) and thus turn out to be ineffective reactive events by annihilation effects. By studying macroscopic relaxation phenomena such as the average micelle length evolution after a T jump, the monomer diffusion, and the zero shear stress relaxation function, we confirm that the effective kinetic constants found are indeed the relevant parameters when macroscopic relaxation is coupled to the kinetics of micelles.
Accuracy and efficiency of electronic energies from systematic molecular fragmentation
Authors: Collins Michael A. , Deev Vitali A.
Published: September 2006

A systematic method for approximating the ab initio electronic energy of molecules from the energies of molecular fragments is tested on a large sample of typical organic molecular structures. The detailed methods, including some additional refinements for molecular rings and long range interactions, are described. The accuracy and computational efficiency of the systematic hierarchy of methods are reported.
Coupling Hydrophilic Amine-Containing Molecules to the Backbone of Poly(ε-Caprolactone)
Authors: Prime Emma L. , Cooper-White Justin J. , Qiao Greg G.
Published: September 2006

A poly(ϵ-caprolactone) (PCL) based biodegradable polymer containing robust, amine-reactive side chains has been successfully synthesized. The specific reactivity of the side chains allows for the coupling of unmodified amine-containing molecules such as poly(l-lysine) (PLL) to PCL to occur in the presence of other unprotected functional groups. The reactivity of this polymer has been demonstrated through successful coupling of both benzylamine (a model compound) and PLL. This novel amine-reactive polymer could have numerous applications in biomedical fields such as tissue engineering and drug delivery.
Miniemulsion Polymerization Stabilized by a Well-Defined, Amphiphilic Gradient Poly(styrene-co-acrylic acid) Copolymer
Authors: Lefay Catherine , Save Maud , Charleux Bernadette , Magnet Stéphanie
Published: September 2006

The nitroxide-mediated controlled free-radical copolymerization of styrene (St) and acrylic acid (AA) was used to synthesize a well-defined poly(St30%-co-AA70%) amphiphilic gradient copolymer. The latter proved to be an efficient stabilizer in the 45 wt.-% solids content, batch miniemulsion polymerizations of St and of mixtures of methyl methacrylate/n-butyl acrylate (35/65 w/w). With 2,2´-azobisisobutyronitrile as an oil-soluble initiator, polystyrene latexes with a very narrow particle distribution were obtained, whereas the water-soluble initiator, potassium persulfate, led to broad, multimodal particle size distributions. Such results were explained by the contribution of two nucleation mechanisms: droplet nucleation and homogeneous nucleation. In contrast, the poly(methyl methacrylate-co-n-butyl acrylate) latexes exhibited larger particle size and narrower particle size distributions with persulfate initiator, than the polystyrene latexes.
Stem Cells Need Chemical Solutions
Authors: Parker Graham C.
Published: September 2006

Stem cell research finds itself at a critical juncture, having presented a promise and potential that has thus far gone substantially unrealized. Chemical science has technology and expertise that can solve some of these shortcomings. Recruiting chemical scientists to collaborate with stem cell biologists will advance significantly efforts to understand normal development of and achieve therapeutic applications using stem cells.
Structure - Property and Structure - Function Relations of Leafhopper (Kahaono montana) Silk
Authors: Chang Jung C. , Gurr Geoff M. , Fletcher Murray J. , Gilbert Robert G.
Published: September 2006

Kahaono montana Evans (Insecta: Hemiptera: Cicadellidae), an endemic Australian leafhopper, is unique among the insect order Hemiptera in producing a silk. In this study, the secondary structure of the protein comprising leafhopper silk, and the surface stretching mechanical properties of this biopolymer, were investigated using Fourier-transform infrared microscopy and atomic force microscopy, respectively. The curve-fitted amide I and amide III bands revealed a composition of 13.1% α-helix, 23.8% β-sheet, 25.5% random coil, and 37.6% aggregated side chains. The molecular stretching behaviour of raw and cleaned silk fibres differed markedly. Analysis of the AFM force curves showed an adhesive property of the raw silk, while the pure fibre showed only the presence of protein. These findings suggest that the silk fibres act as a structural support for other leafhopper secretions and together form a hydrophobic barrier that may protect the insects from rain and natural enemies. This is the first time such a use of silk has been found in a biological system.
Mechanistic Modelling and Network Properties of Ternary Thiol - Vinyl Photopolymerizations
Authors: Reddy Sirish K. , Cramer Neil B. , Kalvaitas Michael , Lee Tai Yeon , Bowman Christopher N.
Published: September 2006

Ternary thiol–vinyl polymerizations offer a unique platform for improved control over polymerization kinetics and network properties as compared to both binary thiol–vinyl systems and traditional (meth)acrylic systems. Therefore, this study seeks to improve the fundamental understanding of the complex ternary thiol–vinyl systems to enable enhanced control over polymerization kinetics, network evolution, and, ultimately, network properties. The polymerization kinetics and material properties afforded by thiol–triazine–methacrylate systems are investigated. The ternary kinetics are successfully predicted by understanding the reaction mechanisms of the corresponding binary components. In ternary thiol–ene–(meth)acrylate systems, the variation in stoichiometric ratios of thiol and ene does not significantly impact material properties as in thiol–ene- or thiol–(meth)acrylate systems. Further, the ternary systems also provide unique polymer properties such as high glass transition temperature with narrow transition widths.
Polymer Chemistry in Diabetes Treatment by Encapsulated Islets of Langerhans: Review to 2006
Authors: Lacík Igor
Published: September 2006

Polymeric materials have been successfully used in numerous medical applications because of their diverse properties. For example, development of a bioartificial pancreas remains a challenge for polymer chemistry. Polymers, as a form of various encapsulation device, have been proposed for designing the semipermeable membrane capable of long-term immunoprotection of transplanted islets of Langerhans, which regulate the blood glucose level in a diabetic patient. This review describes the current situation in the field, discussing aspects of material selection, encapsulation devices, and encapsulation protocols. Problems and unanswered questions are emphasized to illustrate why clinical therapies with encapsulated islets have not been realized, despite intense activity over the past 15 years. The review was prepared with the goal to address professionals in the field as well as the broad polymer community to help in overcoming final barriers to the clinical phase for transplantation of islets of Langerhans encapsulated in a polymeric membrane.
Accurately quantifying low-abundant targets amid similar sequences by revealing hidden correlations in oligonucleotide microarray data
Authors: Marcelino Luisa A. , Backman Vadim , Donaldson Andres , Steadman Claudia , Thompson Janelle R. , Preheim Sarah Pacocha , Lien Cynthia , Lim Eelin , Veneziano Daniele , Polz Martin F.
Published: September 2006

Microarrays have enabled the determination of how thousands of genes are expressed to coordinate function within single organisms. Yet applications to natural or engineered communities where different organisms interact to produce complex properties are hampered by theoretical and technological limitations. Here we describe a general method to accurately identify low-abundant targets in systems containing complex mixtures of homologous targets. We combined an analytical predictor of nonspecific probe–target interactions (cross-hybridization) with an optimization algorithm that iteratively deconvolutes true probe–target signal from raw signal affected by spurious contributions (cross-hybridization, noise, background, and unequal specific hybridization response). The method was capable of quantifying, with unprecedented specificity and accuracy, ribosomal RNA (rRNA) sequences in artificial and natural communities. Controlled experiments with spiked rRNA into artificial and natural communities demonstrated the accuracy of identification and quantitative behavior over different concentration ranges. Finally, we illustrated the power of this methodology for accurate detection of low-abundant targets in natural communities. We accurately identified
Vibrio
taxa in coastal marine samples at their natural concentrations (<0.05% of total bacteria), despite the high potential for cross-hybridization by hundreds of different coexisting rRNAs, suggesting this methodology should be expandable to any microarray platform and system requiring accurate identification of low-abundant targets amid pools of similar sequences.
A microrotary motor powered by bacteria
Authors: Hiratsuka Yuichi , Miyata Makoto , Tada Tetsuya , Uyeda Taro Q. P.
Published: September 2006

Biological molecular motors have a number of unique advantages over artificial motors, including efficient conversion of chemical energy into mechanical work and the potential for self-assembly into larger structures, as is seen in muscle sarcomeres and bacterial and eukaryotic flagella. The development of an appropriate interface between such biological materials and synthetic devices should enable us to realize useful hybrid micromachines. Here we describe a microrotary motor composed of a 20-μm-diameter silicon dioxide rotor driven on a silicon track by the gliding bacterium
Mycoplasma mobile
. This motor is fueled by glucose and inherits some of the properties normally attributed to living systems.
An aged host promotes the evolution of avirulent coxsackievirus into a virulent strain
Authors: Gay Raina T. , Belisle Sarah , Beck Melinda A. , Meydani Simin Nikbin
Published: September 2006

The emergence of new, more pathogenic viruses necessitates elucidation of factors that promote viral evolution. Aging, a potential factor, is associated with increased susceptibility to viral infections. We used the enterovirus coxsackievirus B3 (CVB3) to investigate the effects of host age on pathogenicity and viral gene sequence. Old mice infected with a normally amyocarditic strain of CVB3, CVB3/0, had significantly higher mean heart viral titers compared with CVB3/0-infected adult mice. To determine whether a change in the CVB3/0 viral population could contribute to the higher titers observed in the old infected mice, CVB3/0 was passed once through an old or adult host and the changes in pathogenicity and viral genome were examined after subsequent infection of old or adult mice. Adult mice infected with CVB3/0 that was passed through an old host (CVB3/0
Old
) exhibited significantly higher heart viral titers, pathology, and weight loss than adult mice infected with either stock CVB3/0 or CVB3/0 passed through an adult host (CVB3/0
Adult
). Sequence analysis of virus isolated from CVB3/0
Old
-infected mice revealed 13 specific and reproducible nucleotide changes. These changes result in a sequence that matches the virulent CVB3/20 strain and are associated with promoting cardiovirulence. In contrast, we observed only one nucleotide change, low heart viral titers, and no heart and liver pathology in adult mice infected with CVB3/0
Adult
. These results demonstrate that the aged host promotes rapid selection of a pathogenic variant of CVB3 from an avirulent strain and introduces a host–virus paradigm for studies of viral infection in the aged.
Human cytomegalovirus infection alters the substrate specificities and rapamycin sensitivities of raptor- and rictor-containing complexes
Authors: Kudchodkar Sagar B. , Yu Yongjun , Maguire Tobi G. , Alwine James C.
Published: September 2006

Signaling mediated by the mammalian target of rapamycin kinase (mTOR) is activated during human cytomegalovirus (HCMV) infection. mTOR is found in two complexes differing by the binding partner, rictor or raptor. Activated mTOR-raptor promotes cap-dependent translation through the hyperphosphorylation of the eIF4E-binding protein (4E-BP). This activity of the raptor complex is normally inhibited by cell stress responses or the drug rapamycin. However, we previously showed that this inhibition of mTOR signaling can be circumvented during HCMV infection such that hyperphosphorylation of 4E-BP is maintained. Here we show that HCMV infection also activates the rictor complex, as indicated by increased phosphorylation of Akt S473; this phosphorylation is insensitive to rapamycin but sensitive to caffeine in both uninfected and infected cells. By using short-hairpin RNAs to deplete rictor and raptor, we find that rictor is more significant than raptor for the viral infection. Surprisingly, the inhibitory effects of rapamycin on viral growth are primarily due to the presence of rictor, not raptor. Raptor and rictor depletion experiments show that in HCMV-infected cells, both raptor- and rictor-containing complexes can mediate the hyperphosphorylation of 4E-BP and the phosphorylation of p70S6 kinase. Under these conditions, the rictor complex is rapamycin-sensitive for the hyperphosphorylation of 4E-BP, but the raptor complex is not. These data suggest that, during HCMV infection, the rictor- and raptor-containing complexes are modified such that their substrate specificities and rapamycin sensitivities are altered. Our data also suggest that the present understanding of rapamycin's inhibitory effects is incomplete.
New building blocks in the 2/1 crystalline approximant of a Bergman-type icosahedral quasicrystal
Authors: Lin Qisheng , Corbett John D.
Published: September 2006

The refined x-ray crystal structure of the phase Mg
27
Al
10.7(2)
Zn
47.3(2)
(
Pa
3
) establishes it as the new 2/1 Bergman-type approximant of the icosahedral quasicrystal. The primitive cubic lattice consists of condensed triacontahedral and novel prolate rhombohedral (PR) clusters. Each triacontahedron encapsulates the traditional, multiply endohedral Bergman-type clusters, and each PR encapsulates an Al
2
dimer. This phase exhibits the same long-range order as recently established for the Tsai-type Sc–Mg–Zn 2/1 approximant crystal, with substantial geometric and atomic distribution differences between the two only in the short range orders. This common feature suggests that Bergman- and Tsai-type quasicrystals may be more similar than earlier conceived. Factors germane to the formation of, and the differences between, Bergman- vs. Tsai-type 1/1 and 2/1 approximate structures are considered, including notably different distributions of the more electropositive elements.
Chiral-selective aminoacylation of an RNA minihelix: Mechanistic features and chiral suppression
Authors: Tamura Koji , Schimmel Paul R.
Published: September 2006

Aminoacylation of RNA minihelices is speculated to be a key step in the transition from the putative RNA world to the theater of proteins. This reaction affords the opportunity to make chiral selection of an
l
- or
d
-amino acid and thus determine the ultimate chirality that is incorporated into proteins. Previous work showed chiral preference of aminoacylation with a nonprotein, nonribozyme, RNA-directed aminoacylation system. This preference was, in turn, determined by the preexisting chirality of the RNA. The α-amino group attached to the asymmetric α-carbon of the amino acid was an obvious candidate to play a role in chiral selectivity through interactions with the RNA. Also not clear was whether a simple manipulation could change the chiral selectivity, thereby giving insight into the basis of chiral selection in the first place. Here we show, surprisingly, no role for the free α-amino group in chiral selection. However, by a sequence manipulation, chiral preference was suppressed and partly reversed. This result and those with further RNA constructs support the idea that the chiral preference for an
l
-amino acid in these constructs depends on avoiding a sugar-pucker-sensitive steric clash between a pendant group of a base with the amino acid side chain.
Control of growth and positional information by the graded
vestigial
expression pattern in the wing of
Drosophila
melanogaster
Authors: Baena-Lopez L. A. , García-Bellido A.
Published: September 2006

The size and shape of organs depend on cellular processes such as cell proliferation, cell survival, and spatial arrangement of cells. In turn, all of these processes are a consequence of positional identity of individual cells in whole organs. Links of positional information with organ growth and pattern expression of genes is a little-addressed question. We show that differences in
vestigial
expression between neighboring cells of the wing blade autonomously and nonautonomously affect cell proliferation along the proximo-distal axis. On the other hand, uniform expression of
vestigial
inhibits cell proliferation and also perturbs the shape of wing blade altering the preferential orientation of cell divisions. Our observations provide evidence that local cell interactions, triggered by differences in
vestigial
expression between neighboring cells, confer positional values operating in the control of growth and shape of the wing.
Men accessing education: gendered aspirations
Authors: Burke Penny Jane
Published: September 2006

Raising aspirations has been identified as a key strategy for widening educational participation in lifelong learning policy. This article deconstructs the hegemonic discourse of raising aspirations through Economic and Social Research Council‐funded research on men, masculinities and higher educational access and participation. The article examines the ways that men students on access and foundation programmes talk about their aspirations and considers the multiple influences and practices that shape their decisions to participate in education. It is argued that a range of interlinked, and contradictory, masculine identifications are central to understanding the formation of aspirations, which are not fixed but shifting through different kinds of life and learning experiences, orientations and relations. The article suggests that widening participation policy and practice is too narrowly focused on simplistic notions of ‘raising aspirations’, leaving hidden intricate operations of power, privilege and inequality.
A locally adaptive statistical procedure (LAP) to identify differentially expressed chromosomal regions
Authors: Callegaro A. , Basso D. , Bicciato S.
Published: September 2006

Motivation: The systematic integration of expression profiles and other types of gene information, such as chromosomal localization, ontological annotations and sequence characteristics, still represents a challenge in the gene expression arena. In particular, the analysis of transcriptional data in context of the physical location of genes in a genome appears promising in detecting chromosomal regions with transcriptional imbalances often characterizing cancer.

Results: A computational tool named locally adaptive statistical procedure (LAP), which incorporates transcriptional data and structural information for the identification of differentially expressed chromosomal regions, is described. LAP accounts for variations in the distance between genes and in gene density by smoothing standard statistics on gene position before testing the significance of their differential levels of gene expression. The procedure smoothes parameters and computes p-values locally to account for the complex structure of the genome and to more precisely estimate the differential expression of chromosomal regions. The application of LAP to three independent sets of raw expression data allowed identifying differentially expressed regions that are directly involved in known chromosomal aberrations characteristic of tumors.

Availability: Functions in R for implementing the LAP method are available at

Contact:  silvio.bicciato@unipd.it

Supplementary Information:
Selective constraint and genetic differentiation in geographically distant barley yellow dwarf virus populations
Authors: Hall Gerod
Published: September 2006

Numerous studies have documented molecular variability in plant virus populations, but few have assessed the relative contribution of natural selection and genetic drift in generating the observed pattern of diversity. To this end, gene function, environment and phylogenetic history were examined to observe the effect on genetic diversity and population structure of the PAV and PAS species ofBarley yellow dwarf virus(familyLuteoviridae). Three functional classes of gene were analysed: transcription-related (RdRp), structural (CP) and movement-related (MP). The results indicate that there were no inherent differences, in terms of total diversity or diversity at synonymous or non-synonymous nucleotide sites, between functional classes of genes or populations. Rather, selective constraints on a gene may be more or less relaxed depending on its function and the phylogenetic history of the population sampled. The CP of the PAS species, but not the PAV species, was differentiated genetically between regions. This is probably due to genetic drift, as there was no evidence that any gene deviated from a neutral model of evolution or is under positive selection. In general, the MP was under considerably less functional constraint than structural or replication-related proteins and four positively selected codon sites were identified. Mutations at these sites differentiate species and geographical subpopulations, so presumably they have aided the virus in adaptation to its host environment and contributed to intra- and interspecies diversification.
Formerly unclassified, acid-stable equine picornaviruses are a third equine rhinitis B virus serotype in the genus Erbovirus
Authors: Black Wesley D. , Studdert Michael J.
Published: September 2006

Acid-stable equine picornaviruses (ASPs) were identified as a distinct serotype of equine picornaviruses that were isolated from nasal swabs taken from horses with acute febrile respiratory disease in the UK and Japan, and were placed in the group of unclassified picornaviruses. The nucleotide sequence of the P1 region, encoding the capsid proteins, was determined for three ASP isolates from the UK and the sequences were aligned with published sequences of Equine rhinitis B virus (ERBV), genus Erbovirus, including acid-labile ERBV1 and ERBV2 and the recently identified acid-stable ERBV1. The ASPs belong to the same phylogenetic group, composed of five acid-stable ERBV1 isolates. ERBV1 rabbit antiserum neutralized the ASP isolates at approximately 1/10 titre relative to acid-stable and acid-labile ERBV1 isolates, supporting prior findings that ASPs are a distinct serotype, albeit cross-neutralizing weakly with ERBV1. The genus Erbovirus therefore presently comprises three serotypes: ERBV1, ERBV2 and the proposed ERBV3.
Endotheliotropic elephant herpesvirus, the first betaherpesvirus with a thymidine kinase gene
Authors: Ehlers Bernhard , Dural Güzin , Marschall Manfred , Schregel Vera , Goltz Michael , Hentschke Jochen
Published: September 2006

Endotheliotropic elephant herpesvirus (elephantid herpesvirus 1; ElHV-1) is apathogenic for African elephants (Loxodonta africana), but causes fatal haemorrhagic disease in Asian elephants (Elephas maximus). This is thought to occur through transmission from African elephants in places where both species are housed, such as zoological gardens. The virus has caused considerable losses in North American and European zoological gardens and thus severely impedes breeding of the endangered Asian elephant. Previously, the ultrastructural and genetic characterization of ElHV-1 from a male Asian elephant that died from the disease at the Berlin zoological gardens in 1998 have been reported. Here, a partial characterization of the ElHV-1 genome is presented. A 60 kbp locus, spanning 34 open reading frames, was analysed. Most of the detected genes were found to be conserved among the herpesviruses and showed an overall arrangement most similar to that of betaherpesviruses, in particular Human herpesvirus 6 and Human herpesvirus 7. Most importantly, in addition to a protein kinase gene that is homologous to the human cytomegalovirus UL97 gene, a thymidine kinase (TK) gene was found, which is generally missing in betaherpesvirus genomes. Thus, ElHV-1 is the only known betaherpesvirus to encode a TK gene. This peculiarity might contribute to the fulminant pathogenicity of ElHV-1, but also provide a crucial enzymic activity for developing an efficient antiviral therapy with currently available nucleoside analogues.
The 3′ end of the foot-and-mouth disease virus genome establishes two distinct long-range RNA–RNA interactions with the 5′ end region
Authors: Serrano Paula , Pulido Miguel Rodriguez , Sáiz Margarita , Martínez-Salas Encarnacion
Published: September 2006

The untranslated regions (UTRs) of the foot-and-mouth disease virus (FMDV) genome contain multiple functional elements. In the 5′ UTR, the internal ribosome entry site (IRES) element governs cap-independent translation initiation, whereas the S region is presumably involved in RNA replication. The 3′ UTR, composed of two stem–loops and a poly(A) tract, is required for viral infectivity and stimulates IRES activity. Here, it was found that the 3′ end established two distinct strand-specific, long-range RNA–RNA interactions, one with the S region and another with the IRES element. These interactions were not observed with the 3′ UTR of a different picornavirus. Several results indicated that different 3′ UTR motifs participated in IRES or S region interactions. Firstly, a high-order structure adopted by both the entire IRES and the 3′ UTR was essential for RNA interaction. In contrast, the S region interacted with each of the stem–loops. Secondly, S–3′ UTR interaction but not IRES–3′ UTR interaction was dependent on a poly(A)-dependent conformation. However, no other complexes were observed in mixtures containing the three transcripts, suggesting that these regions did not interact simultaneously with the 3′ UTR probe. Cellular proteins have been found to bind the S region and one of these also binds to the 3′ UTR in a competitive manner. Our data suggest that 5′–3′-end bridging through both direct RNA–RNA contacts and RNA–protein interactions may play an essential role in the FMDV replication cycle.
Genetic and experimental comparison of porcine circovirus type 2 (PCV2) isolates from cases with and without PCV2-associated lesions provides evidence for differences in virulence
Authors: Opriessnig T. , McKeown N. E. , Zhou E.-M. , Meng X.-J. , Halbur P. G.
Published: September 2006

There are marked differences in the clinical expression of diseases associated with porcine circovirus type 2 (PCV2) in the field. The objective of this study was to compare the sequences and pathogenicity of PCV2 isolates from field cases with and without PCV2-associated lesions. Forty-two specific-pathogen-free (SPF) pigs were assigned randomly to three groups of 14 pigs each. At 7 weeks of age, group 1 pigs were mock-inoculated with saline, group 2 pigs were inoculated with PCV2-4838 (isolated from a pig with no evidence of PCV2-associated lymphoid lesions) and group 3 pigs were inoculated with PCV2-40895 (isolated from a pig with PCV2-associated lymphoid lesions and disease). The PCV2-4838 and PCV2-40895 isolates shared approximately 98.9 % nucleotide sequence identity across the entire genome. A total of nine amino acid changes in ORF2 and two amino acid changes in ORF1 were identified between the two isolates. PCV2-4838-inoculated pigs had significantly more genomic copy numbers of PCV2 in their sera at 7 days post-inoculation (p.i.) (P<0.0001) and significantly fewer genomic copy numbers at 14, 21 and 28 days p.i. (P<0.05) compared with pigs inoculated with the PCV2-40895 isolate. Microscopic lesions in lymphoid tissues were significantly less severe (P<0.05) and the amount of PCV2 antigen associated with these lesions was significantly lower (P<0.05) in pigs inoculated with PCV2-4838. The results of this study suggest that PCV2 isolates from the USA differ in virulence in an SPF pig model.
Molecular characterization of the largest mycoviral-like double-stranded RNAs associated with Amasya cherry disease, a disease of presumed fungal aetiology
Authors: Kozlakidis Z. , Covelli L. , Di Serio F. , Citir A. , Açıkgöz S. , Hernández C. , Ragozzino A. , Flores R. , Coutts R. H. A.
Published: September 2006

The sequence of the four large (L) double-stranded RNAs (dsRNAs) associated with Amasya cherry disease (ACD), which has a presumed fungal aetiology, is reported. ACD L dsRNAs 1 (5121 bp) and 2 (5047 bp) potentially encode proteins of 1628 and 1620 aa, respectively, that are 37 % identical and of unknown function. ACD L dsRNAs 3 (4458 bp) and 4 (4303 bp) potentially encode proteins that are 68 % identical and contain the eight motifs conserved in RNA-dependent RNA polymerases (RdRp) of dsRNA mycoviruses, having highest similarity with those of members of the family Totiviridae. Both terminal regions share extensive conservation in all four RNAs, suggesting a functional relationship between them. As ACD L dsRNAs 1 and 2 do not encode RdRps, both are probably replicated by those from either ACD L dsRNA 3 or 4. Partial characterization of the equivalent L dsRNAs 3 and 4 associated with cherry chlorotic rusty spot revealed essentially identical sequences.
Transformation of Tetrahymena thermophila by Electroporation
Authors: Sweet Melody T. , Allis C. David
Published: September 2006

INTRODUCTION

This protocol describes a method for transformation of the Tetrahymena using electroporation. The vector is electroporated into cells after mating, where it is incorporated into the DNA of developing macronuclei. Because T. thermophila can be propagated indefinitely without conjugation, transformation of the macronucleus provides a way to obtain stable somatic transformants. DNA vectors transformed using this protocol include those containing drug-resistant versions of Tetrahymena genes (which replace endogenous genes via homologous recombination) as well as those containing rDNA replication origins. Cotransformation using these two vector types is also possible: Electroporated cells are selected for drug resistance conferred by the replicating vector for 10-15 generations, then screened for the gene replacement using another drug-resistant marker. Usually a few percent of the cells that are transformed by the replicating vector are cotransformed by the gene replacement vector.
Fragmentation of DNA by Nebulization: Figure 1.
Authors: Sambrook Joseph , Russell David W.
Published: September 2006

INTRODUCTION

DNA fragmentation is often necessary prior to library construction or subcloning for DNA sequencing. This protocol describes a method for DNA fragmentation by nebulization, in which the fine mist created by forcing a DNA solution through a small hole in the nebulizer unit is collected. The size of the fragments obtained by nebulization is determined chiefly by the speed at which the DNA solution passes through the hole, altering the pressure of the gas blowing through the nebulizer, the viscosity of the solution, and the temperature. Nebulization is easy, quick, and requires only small amounts of DNA (0.5-5 μg). The resulting DNA fragments are distributed over a narrow range of sizes (700-1330 bp). It requires ligation of DNA before nebulization and end-repair afterward.
Molecular Identification of an ArabidopsisS-Adenosylmethionine Transporter. Analysis of Organ Distribution, Bacterial Expression, Reconstitution into Liposomes, and Functional Characterization
Authors: Palmieri Luigi , Arrigoni Roberto , Blanco Emanuela , Carrari Fernando , Zanor Maria Ines , Studart-Guimaraes Claudia , Fernie Alisdair R. , Palmieri Ferdinando
Published: September 2006

Despite much study of the role of S-adenosylmethionine (SAM) in the methylation of DNA, RNA, and proteins, and as a cofactor for a wide range of biosynthetic processes, little is known concerning the intracellular transport of this essential metabolite. Screening of the Arabidopsis (Arabidopsis thaliana) genome yielded two potential homologs of yeast (Saccharomyces cerevisiae) and human SAM transporters, designated as SAMC1 and SAMC2, both of which belong to the mitochondrial carrier protein family. The SAMC1 gene is broadly expressed at the organ level, although only in specialized tissues of roots with high rates of cell division, and appears to be up-regulated in response to wounding stress, whereas the SAMC2 gene is very poorly expressed in all organs/tissues analyzed. Direct transport assays with the recombinant and reconstituted SAMC1 were utilized to demonstrate that this protein displays a very narrow substrate specificity confined to SAM and its closest analogs. Further experiments revealed that SAMC1 was able to function in uniport and exchange reactions and characterized the transporter as highly active, but sensitive to physiologically relevant concentrations of S-adenosylhomocysteine, S-adenosylcysteine, and adenosylornithine. Green fluorescent protein-based cell biological analysis demonstrated targeting of SAMC1 to mitochondria. Previous proteomic analyses identified this protein also in the chloroplast inner envelope. In keeping with these results, bioinformatics predicted dual localization for SAMC1. These findings suggest that the provision of cytosolically synthesized SAM to mitochondria and possibly also to plastids is mediated by SAMC1 according to the relative demands for this metabolite in the organelles.
Diacidic Motif Is Required for Efficient Transport of the K+ Channel KAT1 to the Plasma Membrane
Authors: Mikosch Melanie , Hurst Annette C. , Hertel Brigitte , Homann Ulrike
Published: September 2006

For a number of mammalian ion channels, trafficking to the plasma membrane was found to be controlled by intrinsic sequence motifs. Among these sequences are diacidic motifs that function as endoplasmic reticulum (ER) export signals. So far it is unclear if similar motifs also exist in plant ion channels. In this study we analyzed the function of four diacidic DXE/DXD motifs of the plant K+ channel KAT1. Mutation of the first diacidic DXE motif resulted in a strong reduction of the KAT1 conductance in both guard cell protoplasts and HEK293 cells (human embryonic kidney cells). Confocal fluorescence microscopy of guard cells expressing the mutated KAT1 fused to green fluorescent protein revealed localization of the mutated channel only in intracellular structures around the nucleus. These structures could be identified as part of the ER via coexpression of KAT1 fused to yellow fluorescent protein with an ER-retained protein (HDEL) fused to cyan fluorescent protein. Block of vesicle formation from the ER by overexpression of the small GTP-binding protein Sar1 fixed in its GDP-bound form led to retention of wild-type KAT1 in similar parts of the ER. Mutation of the three other diacidic motifs had no effect. Together, the results demonstrate that one diacidic motif of KAT1 is essential for ER export of the functional channel in both guard cell protoplasts and HEK293 cells. This suggests that trafficking of plant plasma membrane ion channels is controlled via a conserved mechanism.
Functional characterization of a novel insulin receptor mutation contributing to Rabson–Mendenhall syndrome
Authors: Tuthill A. , Semple R. K. , Day R. , Soos M. A. , Sweeney E. , Seymour P. J. , Didi M. , O’Rahilly S.
Published: September 2006

Objective/Patients  Rabson–Mendenhall syndrome (RMS) is a rare, recessively inherited disorder of extreme insulin resistance due to mutations in the insulin receptor gene. We have identified a pair of siblings with RMS attributable to compound heterozygosity for two insulin receptor mutations, one previously unreported, and have characterized the novel receptor mutation functionally.

Measurements  Insulin receptor sequencing was performed to identify the mutations. Expression levels of the mature receptor were determined in lymphoblastoid cells from the affected subjects. Further studies of immortalized cell lines transfected with mutant and wild type (WT) receptors were undertaken to characterize the effects of the novel mutation on [125I]‐labelled insulin binding, proreceptor processing and insulin‐stimulated receptor autophosphorylation.

Results  Sequencing of the insulin proreceptor coding sequence revealed both siblings to be compound heterozygotes for the missense mutations Arg209His and Gly359Ser in the mature insulin receptor. The former mutation has been described in homozygous form in Donohue syndrome, while the latter is novel. Insulin receptor expression in lymphoblastoid cell lines was present at only 10–30% of that in control cells; studies of immortalized cells transfected with mutant and WT receptors confirmed the reduced expression of the mutant. The degree of impairment of insulin binding and insulin‐stimulated receptor autophosphorylation were commensurate with the decrease in expression of the mature receptor.

Conclusions  Loss of function of the novel insulin receptor (INSR) G359S variant is largely accounted for by aberrant proreceptor processing rather than intrinsically impaired signal transduction by the mutant receptor.
Fine needle aspiration cytology of the thyroid: a comparison of 5469 cytological and final histological diagnoses
Authors: Sangalli G. , Serio G. , Zampatti C. , Bellotti M. , Lomuscio G.
Published: September 2006

Objective:  We evaluated the efficacy of fine needle aspiration cytology (FNAC) of the thyroid in a series of 5469 lesions with histological control and studied the causes of, and the possibility of reducing the limitations of the method.

Methods:  FNAC was always performed by a pathologist under the guidance of a clinician, using a 22‐gauge needle. Generally two aspirations were carried out, and usually four slides were obtained for each nodule; they were then stained with May–Grünwald–Giemsa and with Papanicolaou. The cytological diagnoses were classified in four groups: inadequate, benign, suspicious and malignant.

Results:  We obtained a complete sensitivity of 93.4%, a positive predictive value of malignancy of 98.6%, and a specificity of 74.9%. At histological control, the cytological diagnosis of Hurthle cell neoplasm corresponded to a significantly higher incidence of malignant neoplasms than the diagnosis of non‐Hurthle cell follicular neoplasm (32.1% versus 15.5%). There were 66 false‐negative findings, the main cause of diagnostic error (24 cases) being failure to recognize the follicular variant of papillary carcinoma. The number of inadequate FNACs was low (4.2%).

Conclusion:  Our study confirmed the great efficacy of thyroid FNAC. A cytological diagnosis of Hurthle cell neoplasm should be considered an indicator of high risk. Awareness that failure to recognize the follicular variant of papillary carcinoma was the main problem in the interpretation of thyroid FNAC should lead to a decrease of false‐negative diagnoses. The inadequate rate was very low, as it was the pathologist personally who performed the needle aspiration.
AW01 Blood Transfusion and Hemorrhage: Past, Present and Future
Authors: Dzik W.
Published: September 2006

Introduction  In the early years of the 19th century, James Blundell reported in the Lancet the earliest clinical application of blood transfusion for the treatment of hemorrhage. Although his initial experiments may appear to us to have burst upon the medical world, Blundell had in fact done years of pre‐clinical research using animal models to establish principles to be brought to the clinic.

Methods/Results  Today, blood transfusion remains the cornerstone of treatment for serious bleeding – not only to restore oxygen carrying capacity but also to improve hemostasis, arrest hemorrhage, and prevent bleeding. However, the indications for transfusion with FFP or Platelets for the prevention of bleeding remain ill‐defined. In particular, despite the enormous volumes of blood components transfused worldwide, the evidence that commonly used coagulation tests are reliable guides to transfusion with FFP is scant. Recent laboratory and clinical studies have provided some insights into the weaknesses of the coagulation testing as a guide to blood management. Carefully performed randomized clinical trials are needed in order to better understand how to use transfusion to arrest and prevent hemorrhage. One such trial has recently begun in the USA. In the future, improved understanding of hemostasis will likely bring about the use of new biotherapies that Blundell could not possibly have imagined. The application of genomics to hemostasis is expected to uncover genetic polymorphisms that will lead to improved diagnostics and more tailored medical therapeutics.

Conclusions  While such advances are anticipated, transfusion therapy remains fundamentally important as a life‐saving therapy for bleeding disorders. Moreover, despite our current advanced technologies, hemorrhage during childbirth (which Blundell sought to overcome) remains a devastating problem throughout the world.
SI13 Noninvasive MRI Monitoring of Engraftment of Cultured Bone Marrow Mesenchymal Stem Cell into the Damaged Heart
Authors: Watt S. M. , Stuckey D. J. , Carr C. A. , Martin‐Rendon E. , Hale S. J. M. , Tatton L. , Harding S. E. , Radda G. K. , Clarke K.
Published: September 2006

Stem cells offer a promising approach to the treatment of myocardial infarction and prevention of heart failure. We have used iron‐labelling of bone marrow stromal cells (BMSCs) to noninvasively track cell location in the infarcted rat heart over 16 weeks using cine‐MRI and to isolate the BMSCs from the grafted hearts using the magnetic properties of the donor cells. BMSCs were isolated from rat bone marrow, characterised by flow cytometry, transduced with lentiviral vectors expressing GFP and labelled with iron particles. BMSCs were injected into the infarct periphery immediately following coronary artery ligation and rat hearts were imaged at 1, 4, 10 and 16 weeks post‐infarction. Signal voids caused by the iron particles in the BMSCs were detected in all rats at all time points. In mildly infarcted hearts, the volume of the signal void decreased over the 16 weeks, whilst the signal void volume did not decrease significantly in severely infarcted hearts. High resolution 3D MR microscopy identified hypointense regions at the same position as in vivo. Donor cells containing iron particles and expressing GFP were identified in MR‐targeted heart sections after magnetic cell separation from digested hearts. In conclusion, MRI can be used to track cells labelled with iron particles in damaged tissue for at least 16 weeks after injection and to guide tissue sectioning by accurately identifying regions of cell engraftment. The magnetic properties of the iron labelled donor cells can be used for their isolation from host tissue to enable further characterisation.