bluuebunny/crossref_metadata_2025 huggingface dataset:
Structural and elastic properties of ternary silicides ScTSi (T═Co, Ni, Cu, Ru, Rh, Pd, Ir, Pt) and of the equiatomic intermetallic compounds YTX (T═Ni, Ir and X═Si, Ge, Sn, Pb)
Authors: Sebehi N. , Bouamama Kh. , Djemia Ph. , Kassali K.
Published: August 2015

The structural and elastic properties of the ternary silicides ScTSi (T═Co, Ni, Cu, Ru, Rh, Pd, Ir, Pt) and of the equiatomic intermetallic YTX (T═Ni, Ir; X═Si, Ge, Sn, Pb) are studied using the density‐functional theory considering the orthorhombic TiNiSi crystal structure with space group Pnma. From the calculated formation energy, we obtained the following stability order of: ScCuSi < ScCoSi < ScRuSi < ScNiSi < ScPdSi < ScRhSi < ScIrSi < ScPtSi and YNiPb < YNiSn < YNiGe < YNiSi < YIrSn < YIrGe < YIrPb < YIrSi. The lattice parameters and atomic positions are in good agreement with experiments to ∼1%. All the alloys are mechanically stable using the Born elastic stability criteria. Ductile behavior inferred from the Pugh ratio, shear over bulk modulus G/B, and the Cauchy pressure, could be observed only for ScTSi (TRu, Pd, Pt) and YNiSn compounds and moderately for YIrX (Si, Ge, and Sn) and YNiPb. On the contrary, ScTSi (TCo, Ni, Cu, Rh, Ir), YNiGe, and YNiSi compounds are covalent or exhibit G/B > 0.5, leading to a brittle behavior in accordance with the Poisson ratio value. The Zener anisotropy factor is different from 1, indicating that these compounds are elastically anisotropic materials. A strong correlation is found between G and the Young's modulus E, slightly above G ∼(3/8)E widely found for polycrystalline metals.

Correlation trend between the Young's modulus E and the shear modulus G of intermetallic compounds.
Formation of Au nanocluster ordered array on Si(111)‐7 × 7 surface
Authors: Qu Bo , Hu Jin‐Hang , Li Hang , Li Wen‐Jie , Huang Miao‐Ling , Wu Qi‐Hui
Published: August 2015

We report a successful approach in fabricating well shape two‐dimensional Au nanocluster hexagonal array on an Si(111)‐7 × 7 surface. The size of Au hexagons is about 2.5 × 2.5 nm2. The easiness of preparing large scale Au hexagonal array makes this approach usable to fabricate a wide range of other metal nanocluster arrays. Further deposition of small amount of Au nanoclusters on the Au hexagonal array surface will prefer to occupy the holes of the hexagonal array at room temperature. Finally, a reasonable structure of formatting the hexagonal Au nanocluster array on the Si(111)‐7 × 7 surface was proposed. Copyright © 2015 John Wiley & Sons, Ltd.
Changes in the Surgical Management of Parastomal Hernias Over 15 Years: Results of 135 Cases
Authors: Köhler Gernot , Mayer Franz , Wundsam Helwig , Schrittwieser Rudolf , Emmanuel Klaus , Lechner Michael
Published: August 2015

Background

Over the years, various open and laparoscopic approaches toward the repair of parastomal hernias (PSH) have been described. The variety of published techniques itself can be seen as an indicator for the often low level of satisfaction reached with the surgical procedures.

Methods

From January 1999 to January 2014, we assessed all cases of PSH repair performed at the three participating surgical departments in a retrospective analysis. The results were evaluated with regard to different surgical techniques focusing on complications and recurrences.

Results

One hundred and thirty‐five individuals could be included in the analysis. They were operated on with eight different surgical techniques. Laparoscopic procedures were carried out in 46.7 % (63/135) of the cases. Median follow‐up was 54 months (12–146 months). We found 44 cases of recurrence (32.6 %) and 24 (17.8 %) of the patients experienced perioperative complications and 12 of them needed to return to theater. Fourteen of the 135 patients (10.4 %) were operated as emergency cases which were associated with a mortality of 28.6 % (4/14). In case of elective PSH repair, no mortality occured.

Conclusion

The results achieved by direct suture or the use of incised flat meshes for the repair of PSH were poor with these procedures having unacceptably high recurrence rates. With regard to the latter ostomy revision through three‐dimensional funnel‐shaped meshes and the laparoscopic sandwich technique showed the best results. Emergency procedures were linked to a dramatic increase in morbidity and mortality (p < 0.001).
Model-based evaluation of ferrous iron oxidation by acidophilic bacteria in chemostat and biofilm airlift reactors
Authors: Ebrahimi Sirous , Faraghi Neda , Hosseini Maryam
Published: August 2015

This article presents a model-based evaluation of ferrous iron oxidation in chemostat and biofilm airlift reactors inoculated with a mixed culture of Acidithiobacillus ferrooxidans and Leptospirillum ferrooxidans bacteria. The competition between the two types of bacteria in the chemostat and in the biofilm airlift reactors together with the distribution of both bacteria along the biofilm thickness at different time sections has been studied. The bacterial distribution profiles along the biofilm in the airlift reactor at different time scales show that in the beginning A. ferrooxidans bacteria are dominant, but when the reactor operates for a long time the desirable L. ferrooxidans species outcompete A. ferrooxidans as a result of the low Fe2+ and high Fe3+ concentrations. The results obtained from the simulation were compared with the experimental data of continuously operated internal loop airlift biofilm reactor. The model results are in good agreement with the experimental results.
Burnout and its Effect on Neurocognitive Performance
Authors: McInerney Shane , Rowan Michael , Lawlor Brian
Published: August 2015

Objectives: The hypothesis that burnout is predictive of performance deficits on a number of neuropsychological tests was examined among a group of psychiatric nurses. A possible relationship between burnout and state and trait anxiety levels was also investigated.

Methods: A sample of psychiatric nurses (n=45) completed the Maslach Burnout Inventory (MBI) and the State-Trait Anxiety Inventory (STAI). Their performance on a number of neurocognitive tests from the Cambridge Neuropsychological Test Automated Battery (CANTAB) was examined. The neurocognitive tests measured visual memory, verbal memory and attention.

Results: Multiple regression analyses were conducted between the burnout subscales and the results of the neuropsychological tests, and also between the burnout subscales and both state and trait anxiety levels. No overall significant relationship was found between neurocognitive performance and burnout. However, trait anxiety was significantly predictive of each of the burnout subscales of emotional exhaustion, depersonalisation and also of decreased personal accomplishment.

Conclusions: The levels of burnout were in the moderate range for all three subscales and so the population was suffering from burnout. The fact that the trait anxiety variable was predictive of all three of the burnout subscales suggests that predisposition towards anxiety may be a vulnerability factor among people who suffer from burnout.
N2-fixation, ammonium release and N-transfer to the microbial and classical food web within a plankton community
Authors: Adam Birgit , Klawonn Isabell , Svedén Jennie B , Bergkvist Johanna , Nahar Nurun , Walve Jakob , Littmann Sten , Whitehouse Martin J , Lavik Gaute , Kuypers Marcel M M , Ploug Helle
Published: August 2015

We investigated the role of N2-fixation by the colony-forming cyanobacterium, Aphanizomenon spp., for the plankton community and N-budget of the N-limited Baltic Sea during summer by using stable isotope tracers combined with novel secondary ion mass spectrometry, conventional mass spectrometry and nutrient analysis. When incubated with 15N2, Aphanizomenon spp. showed a strong 15N-enrichment implying substantial 15N2-fixation. Intriguingly, Aphanizomenon did not assimilate tracers of 15NH4+ from the surrounding water. These findings are in line with model calculations that confirmed a negligible N-source by diffusion-limited NH4+ fluxes to Aphanizomenon colonies at low bulk concentrations (<250 nm) as compared with N2-fixation within colonies. No N2-fixation was detected in autotrophic microorganisms <5 μm, which relied on NH4+ uptake from the surrounding water. Aphanizomenon released about 50% of its newly fixed N2 as NH4+. However, NH4+ did not accumulate in the water but was transferred to heterotrophic and autotrophic microorganisms as well as to diatoms (Chaetoceros sp.) and copepods with a turnover time of ~5 h. We provide direct quantitative evidence that colony-forming Aphanizomenon releases about half of its recently fixed N2 as NH4+, which is transferred to the prokaryotic and eukaryotic plankton forming the basis of the food web in the plankton community. Transfer of newly fixed nitrogen to diatoms and copepods furthermore implies a fast export to shallow sediments via fast-sinking fecal pellets and aggregates. Hence, N2-fixing colony-forming cyanobacteria can have profound impact on ecosystem productivity and biogeochemical processes at shorter time scales (hours to days) than previously thought.
Improved binding site assignment by high-resolution mapping of RNA–protein interactions using iCLIP
Authors: Hauer Christian , Curk Tomaz , Anders Simon , Schwarzl Thomas , Alleaume Anne-Marie , Sieber Jana , Hollerer Ina , Bhuvanagiri Madhuri , Huber Wolfgang , Hentze Matthias W. , Kulozik Andreas E.
Published: August 2015

Individual-nucleotide resolution crosslinking and immunoprecipitation (iCLIP) allows the determination of crosslinking sites of RNA-binding proteins (RBPs) on RNAs. iCLIP is based on ultraviolet light crosslinking of RBPs to RNA, reverse transcription and high-throughput sequencing of fragments terminating at the site of crosslinking. As a result, start sites of iCLIP fragments are expected to cluster with a narrow distribution, typically representing the site of direct interaction between the RBP and the RNA. Here we show that for several RBPs (eIF4A3, PTB, SRSF3, SRSF4 and hnRNP L), the start sites of iCLIP fragments show a fragment length-dependent broader distribution that can be shifted to positions upstream of the known RNA-binding site. We developed an analysis tool that identifies these shifts and can improve the positioning of RBP binding sites.
Whole-central nervous system functional imaging in larval Drosophila
Authors: Lemon William C. , Pulver Stefan R. , Höckendorf Burkhard , McDole Katie , Branson Kristin , Freeman Jeremy , Keller Philipp J.
Published: August 2015

Understanding how the brain works in tight concert with the rest of the central nervous system (CNS) hinges upon knowledge of coordinated activity patterns across the whole CNS. We present a method for measuring activity in an entire, non-transparent CNS with high spatiotemporal resolution. We combine a light-sheet microscope capable of simultaneous multi-view imaging at volumetric speeds 25-fold faster than the state-of-the-art, a whole-CNS imaging assay for the isolated Drosophila larval CNS and a computational framework for analysing multi-view, whole-CNS calcium imaging data. We image both brain and ventral nerve cord, covering the entire CNS at 2 or 5 Hz with two- or one-photon excitation, respectively. By mapping network activity during fictive behaviours and quantitatively comparing high-resolution whole-CNS activity maps across individuals, we predict functional connections between CNS regions and reveal neurons in the brain that identify type and temporal state of motor programs executed in the ventral nerve cord.
Dynamic markers based on blood perfusion fluctuations for selecting skin melanocytic lesions for biopsy
Authors: Lancaster Gemma , Stefanovska Aneta , Pesce Margherita , Marco Vezzoni Gian , Loggini Barbara , Pingitore Raffaele , Ghiara Fabrizio , Barachini Paolo , Cervadoro Gregorio , Romanelli Marco , Rossi Marco
Published: August 2015

Skin malignant melanoma is a highly angiogenic cancer, necessitating early diagnosis for positive prognosis. The current diagnostic standard of biopsy and histological examination inevitably leads to many unnecessary invasive excisions. Here, we propose a non-invasive method of identification of melanoma based on blood flow dynamics. We consider a wide frequency range from 0.005–2 Hz associated with both local vascular regulation and effects of cardiac pulsation. Combining uniquely the power of oscillations associated with individual physiological processes we obtain a marker which distinguishes between melanoma and atypical nevi with sensitivity of 100% and specificity of 90.9%. The method reveals valuable functional information about the melanoma microenvironment. It also provides the means for simple, accurate, in vivo distinction between malignant melanoma and atypical nevi and may lead to a substantial reduction in the number of biopsies currently undertaken.
Integrated miRNA-mRNA analysis in the habenula nuclei of mice intravenously self-administering nicotine
Authors: Lee Sangjoon , Woo Jiwan , Kim Yong Sik , Im Heh-In
Published: August 2015

A considerable amount of evidence suggests that microRNAs (miRNAs) play crucial roles in the neuroadaptation of drug addiction. Habenula (Hb), one of the critical brain regions involved in reward and addiction, can be divided into two anatomically and transcriptionally distinct regions: medial habenula (MHb) and lateral habenula (LHb) nuclei. However, very few studies have compared the functional roles of these regions. Here, by using mirConnX integrator and KEGG pathway mapping, we simultaneously analysed the differential expression patterns of miRNAs and messenger RNA (mRNA) within MHb and LHb under nicotine addiction. Significantly altered miRNAs and mRNAs were found in the Hb of mice intravenously self-administering nicotine. Interestingly, some miRNAs were oppositely regulated between the MHb and the LHb and their potential targets included various genes of cell signalling pathways related to the degeneration of fasciculus retroflexus (FR). This study provides an improved insight into the differential regulation of habenular transcripts in nicotine addiction, as well as the potential functions of miRNAs in several biological pathways involved in the nicotine addiction.
miR-98 and its host gene Huwe1 target Caspase-3 in Silica nanoparticles-treated male germ cells
Authors: Xu Bo , Mao Zhilei , Ji Xiaoli , Yao Mengmeng , Chen Minjian , Zhang Xuemei , Hang Bo , Liu Yi , Tang Wei , Tang Qiusha , Xia Yankai
Published: August 2015

Silica nanoparticles (NP) is one of the most commonly used nanomaterials with potential health hazards. However, the effects of Silica NP on germ cells and the underlying mechanisms are still unclear. In this study, GC-2 and TM-4, which are two different types of male germ cells were exposed to Silica NP for 24h and then general cytotoxicity and multi-parameter cytotoxicity were evaluated. Our results showed that Silica NP could induce apoptosis in GC-2 cells. Transmission electron microscopy (TEM) results showed that Silica NP was localized in the lysosomes of GC-2 cells. High content screening (HCS) showed that Silica NP exposure could increased cell permeabilization and decreased mitochondrial membrane potential in GC-2 cells. The mRNA and protein levels of apoptosis markers (Bax, Caspase-3, Caspase-9) in GC-2 cells were significantly increased, while Bcl-2 was decreased. Accordingly, the expression level of miR-98, which can regulate Caspase-3, was significantly decreased. Huwe1, the host gene of miR-98, was positively associated with miR-98 expression after Silica NP exposure. Dual luciferase reporter assay suggested that miR-98 directly targets Caspase-3. These results suggest that Silica NP induces apoptosis via loss of mitochondrial membrane potential and Caspase-3 activation, while miR-98 plays key role in modulating this effect.
Highly Pathogenic Avian Influenza A(H5N1) Virus Struck Migratory Birds in China in 2015
Authors: Bi Yuhai , Zhang Zhenjie , Liu Wenjun , Yin Yanbo , Hong Jianmin , Li Xiangdong , Wang Haiming , Wong Gary , Chen Jianjun , Li Yunfeng , Ru Wendong , Gao Ruyi , Liu Di , Liu Yingxia , Zhou Boping , Gao George F. , Shi Weifeng , Lei Fumin
Published: August 2015

Approximately 100 migratory birds, including whooper swans and pochards, were found dead in the Sanmenxia Reservoir Area of China during January 2015. The causative agent behind this outbreak was identified as H5N1 highly pathogenic avian influenza virus (HPAIV). Genetic and phylogenetic analyses revealed that this Sanmenxia H5N1 virus was a novel reassortant, possessing a Clade 2.3.2.1c HA gene and a H9N2-derived PB2 gene. Sanmenxia Clade 2.3.2.1c-like H5N1 viruses possess the closest genetic identity to A/Alberta/01/2014 (H5N1), which recently caused a fatal respiratory infection in Canada with signs of meningoencephalitis, a highly unusual symptom with influenza infections in humans. Furthermore, this virus was shown to be highly pathogenic to both birds and mammals and demonstrate tropism for the nervous system. Due to the geographical location of Sanmenxia, these novel H5N1 viruses also have the potential to be imported to other regions through the migration of wild birds, similar to the H5N1 outbreak amongst migratory birds in Qinghai Lake during 2005. Therefore, further investigation and monitoring is required to prevent this novel reassortant virus from becoming a new threat to public health.
High ROR2 expression in tumor cells and stroma is correlated with poor prognosis in pancreatic ductal adenocarcinoma
Authors: Huang Jianfei , Fan Xiangjun , Wang Xudong , Lu Yuhua , Zhu Huijun , Wang Wei , Zhang Shu , Wang Zhiwei
Published: August 2015

RTK-like orphan receptor 2 (ROR2) is overexpressed in several cancers and has tumorigenic activity. However, the expression of ROR2 and its functional and prognostic significance have yet to be evaluated in pancreatic ductal adenocarcinoma (PDAC). Quantitative real-time polymerase chain reaction was used to characterize the expression of ROR2 mRNA in PDAC, corresponding peritumoral tissues and PDAC cell lines. Immunohistochemical analysis with tissue microarrays was used to evaluate ROR2 expression in PDAC and to investigate the relationship of this expression to clinicopathological factors and prognosis. The expression of ROR2 mRNA and protein was significantly higher in PDAC than in normal pancreatic tissues. High cytoplasmic ROR2 expression in cancer cells was significantly associated with a primary tumor, distant metastasis and TNM stage and high stromal ROR2 expression was significantly associated with regional lymph node metastasis and TNM stage. The Kaplan–Meier method and Cox regression analyses showed that high ROR2 expression in tumor cytoplasm or stromal cells was significantly associated with malignant attributes and reduced survival in PDAC. We present strong evidence that ROR2 could be used as an indicator of poor prognosis and could represent a novel therapeutic target for PDAC.
Spectroscopic evidence of ‘jumping and pecking’ of cholinium and H-bond enhanced cation–cation interaction in ionic liquids
Authors: Knorr Anne , Fumino Koichi , Bonsa Anne-Marie , Ludwig Ralf
Published: August 2015

Spectroscopic evidence for cation–cation interaction in ionic liquids. The repulsive electrostatic interaction is overcome by hydrogen bonding between ions of like charge.
A three-component synthesis of aryl(heteroaryl)acylamides
Authors: Loska Rafał , Bukowska Patrycja
Published: August 2015

α-Heteroarylamides of diverse structures are assembled in a three-component reaction of azole or azineN-oxides, 1,1-difluorostyrenes and amines.
Controlled micellar disassembly of photo- and pH-cleavable linear-dendritic block copolymers
Authors: Kalva Nagendra , Parekh Nimisha , Ambade Ashootosh V.
Published: August 2015

A biocompatible linear-dendritic copolymer comprising photo and pH-cleavable groups at the junction was synthesised and controlled release of drug into cells was demonstrated.
Cadmium ion sorption from aqueous solutions by high surface area ethylenediaminetetraacetic acid- and diethylene triamine pentaacetic acid-treated carbon nanotubes
Authors: Amiri Ahmad , Shanbedi Mehdi , Savari Maryam , Chew B. T. , Kazi S. N.
Published: August 2015

Friedel–Crafts acylation under microwave irradiation has been utilized for realizing functionalization of CNT with EDTA and DTPA. Significant increases in the specific surface area and sequestering metal ions are the most important advantages.
Marine inspired antiplasmodial thiazinoquinones: synthesis, computational studies and electrochemical assays
Authors: Imperatore C. , Persico M. , Aiello A. , Luciano P. , Guiso M. , Sanasi M. F. , Taramelli D. , Parapini S. , Cebrián-Torrejón G. , Doménech-Carbó A. , Fattorusso C. , Menna M.
Published: August 2015

An original approach, starting from marine derived compounds and combining chemical, computational and electrochemical methods, evidenced the thiazinoquinone scaffold as a new chemotype active againstP. falciparum.
Characterization of an exopolysaccharide from probiont Enterobacter faecalis MSI12 and its effect on the disruption of Candida albicans biofilm
Authors: Kiran G. Seghal , Priyadharshini S. , Anitha K. , Gnanamani Elumalai , Selvin Joseph
Published: August 2015

A study reporting the EPS from a probiotic strain Enterococcus faecalis MSI12 isolated from the marine environment and its potential as an antibiofilm agent and/or disruptor of pre-established biofilms of the most prominent human pathogen Candida albicans.
Inhibitory potential of some chalcones on cathepsins B, H and L
Authors: Garg Shweta , Raghav Neera
Published: August 2015

Cathepsins, intracellular proteases, are known to be involved in a number of physiological processes such as degradation of extracellular proteins, prohormone processing, progressions of atherosclerosis etc.
Preparation and properties of polystyrene nanocomposites containing dumbbell-shaped molecular nanoparticles based on polyhedral oligomeric silsesquioxane and [60]fullerene
Authors: Han Di , Guo Qing–Yun , Zhang Wen-Bin , Liu Liu-Xu , Fu Qiang
Published: August 2015

POSS–C60 as a novel nanofiller for polymer nanocomposites.
Generation of Cu–In alloy surfaces from CuInO2as selective catalytic sites for CO2electroreduction
Authors: Jedidi Abdesslem , Rasul Shahid , Masih Dilshad , Cavallo Luigi , Takanabe Kazuhiro
Published: August 2015

Selective electrocatalytic CO2reduction was achieved using CuIn alloy synthesized from CuInO2.
Downstream of the plant circadian clock: output pathways for the control of physiology and development
Authors: Adams Sally , Carré Isabelle A
Published: August 2015

The plant circadian clock controls many aspects of growth and development, allowing an individual to adapt its physiology and metabolism in anticipation of diurnal and seasonal environmental changes. Circadian regulation of hormone levels and hormonal signalling modulates many features of development, including daily growth patterns and the breaking of seed dormancy. The clock also plays a role in seasonal day-length perception, allowing plants to optimally time key development transitions, such as reproduction. Moreover, the clock restricts (gates) the sensitivity of a plant's response to environmental cues, such as light and stress, to specific times of the day, ensuring that the plant can distinguish between normal fluctuations and longer-term changes. The central oscillator controls many of these output pathways via rhythmic gene expression, with several of the core clock components encoding transcription factors. Post-transcriptional processes are also likely to make an important contribution to the circadian regulation of output pathways. The plant circadian clock plays a role in regulating fitness, hybrid vigour and numerous stress responses. Thus elucidating the complexities of the circadian output mechanisms and their regulation may provide new avenues for crop enhancement.
Studies in copper status and atherosclerosis
Authors: KINSMAN G. D. , HOWARD A. N. , STONE D. L. , MULLINS P. A.
Published: August 2015

Summary

Findings of this paper indicate that leucocyte copper has a significant link with the level of atherosclerosis found within the groups studied. Therefore copper may be involved in the mechanisms associated with ischaemic heart disease (IHD).
Rhizobium meliloti exopolysaccharides: genetic analyses and symbiotic importance
Authors: Reuber T. Lynne , Reed Jason , Glazebrook Jane , Glucksmann M. Alexandra , Ahmann Dianne , Marra Andrea , Walker Graham C.
Published: August 2015

Summary

Genetic experiments have indicated that succinoglycan (EPS I), the acidic Calcofluor-binding exopolysaccharide, of the nitrogen-fixing bacterium Rhizobium meliloti strain Rm1021 is required for nodule invasion and possibly for later events in nodule development on alfalfa and other hosts. Fourteen exo loci on the second megaplasmid have been identified that are required for, or affect, the synthesis of EPS I. Mutations in certain of these loci completely abolish the production of EPS I and result in mutants that form empty Fix- nodules. We have identified two loci, exoR and exoS, that are involved in the regulation of EPS I synthesis in the free-living state. Certain exo mutations which completely abolish EPS I production are lethal in an exoR95 or exoS96 background. Histochemical analyses of the expression of exo genes during nodulation using exo :: TnphoA fusions have indicated that the exo genes are expressed most strongly in the invasion zone. In addition, we have discovered that R. meliloti has a latent capacity to synthesize a second exopolysaccharide (EPS II) that can substitute for the role(s) of EPS I in nodulation of alfalfa but not of other hosts. Possible roles for exopolysaccharides in symbiosis are discussed.
The Association of the Adaptor Complexes AP1 and AP3 with GLUT4 Vesicles: Implications for GLUT4 Compartmentalisation
Authors: Gillingham Alison K. , Koumanov Françoise , Pryor Paul R. , Reaves Barbara J. , Holman Geoffrey D.
Published: August 2015

By using glycerol density gradients of post-plasma membrane supernatants from rat adipocytes, we have been able to separate the compartment containing GLUT4 from that containing transferrin receptors. The major pool of GLUT4 (≈ 80%) is localised to dense fractions, which also contain VAMP2, while the remainder is in lighter fractions, some of which also contain transferrin receptors. AP1 and AP3 complexes are detected in two fractions, the heavier of which co-sediments with the major pool of GLUT4. Nycodenz gradients have been used to examine the in vitro effects of GTP-γ-S on adaptor recruitment. On addition of GTPγS, GLUT4 vesicles fractionate as a heavier population of vesicles, which we suggest is due to a coating reaction. Under these conditions there is an increase in co-sedimentation of GLUT4 with API, but not with AP3. Western blotting of proteins associated with isolated GLUT4 vesicles shows the presence of AP1 and AP3 adaptor complexes. Cell free, in vitro association of the AP1 complex with GLUT4 vesicles is increased 3-fold by the addition of GTP-γ-S and an ATP regenerating system to the post-plasma membrane lysate. and 4-fold by carrying out the incubation at 37°C compared with 0°C. These data demonstrate that GLUT4 subcellular localisation can be studied in isolated fractions and that GLUT4 compartments are associated with at least two distinct adaptor complexes.
Studying metabolic regulation in human muscle
Authors: Kemp G. J.
Published: August 2015

Phosphorus magnetic resonance spectroscopy, despite some limitations, is a valuable non-invasive window on muscle metabolism in vivo, particularly oxidative ATP synthesis. A number of experiments have shown this to be dominated by closed-loop feedback mechanisms: a well-known model posits regulation by ADP, but there are others, difficult to distinguish experimentally. Moreover the contribution of open-loop control mechanisms (‘feed forward’ or ‘parallel activation’) in vivo remains controversial. Progress will require more precise data, better integrated with other measurements (e.g. muscle oxygenation), and improvement of the conceptual tools appropriate to such studies, where data are limited and steady-state assumptions frequently inapplicable.
Muscle fibres: applications for the study of the metabolic consequences of enzyme deficiencies in skeletal muscle
Authors: Vielhaber S. , Kudin A. , Schröder R. , Elger C. E. , Kunz W. S.
Published: August 2015

Mitochondrial function in saponin-permeabilized muscle fibres can be studied by high-resolution respirometry, laser-excited fluorescence spectroscopy and fluorescence microscopy. We applied these techniques to study metabolic effects of changes in the pattern of mitochondrial enzymes in skeletal muscle of patients with chronic progressive external ophthalmoplegia or Kearns-Sayre syndrome harbouring large-scale deletions of mitchondrial DNA (mtDNA). In all patients combined deficiencies of respiratory chain enzymes containing mitochondrially encoded subunits were observed. The citrate synthase-normalized activity ratios of these enzymes decreased linearly with increasing mtDNA heteroplasmy. This indicates the absence of any well-defined mutation thresholds for mitochondrial enzyme activities in the entire skeletal muscle. We applied metabolic control analysis to perform a quantitative estimation of the metabolic influence of the observed enzyme deficiencies. For patients with degrees of mtDNA heteroplasmy below about 60% we observed at almost normal maximal rates of respiration an increase in flux control coefficients of complexes I and IV. Permeabilized skeletal-muscle fibres of patients with higher degrees of mtDNA heteroplasmy and severe enzyme deficiencies exhibited additionally decreased maximal rates of respiration. This finding indicates the presence of a ‘metabolic threshold’ which can be assessed by functional studies of muscle fibres providing the link to the phenotypic expression of the mtDNA mutation in skeletal muscle.
Structure—function relations in the human placenta
Authors: Dockery P. , Bermingham J. , Jenkins D.
Published: August 2015

The human haemochorial placenta is a complex and dynamic interface between embryonic and maternal tissues. A myriad array of compounds has been identified at this interface, some of which exert local effects which might be important in maintaining the integrity of the organ. These compounds are diverse in nature and function; they include enzymes, hormones and bioactive peptides. Successful nidation requires the synchronization of endometrial maturation and embryonic development. The complex nature of this interface requires the application of sound sampling strategies. The new stereological methods have thrown fresh light on the growth and development of the human placenta. These methods permit the objective, quantitative description of morphology by efficient design-based methods. This approach has permitted a better definition of the functional morphology of the placenta. Applications of these methodologies are providing a spatial and temporal framework on which to lay the new physiological and molecular information. Here we review the essential features of the stereological approach, identify useful structural quantities and provide some examples of their application. The problems associated with the quantification of immunocytochemistry are illustrated with the use of immunoreactivity to insulin-like growth factor I receptor in normal placentae and in pre-eclampsia. Although stereology can provide useful quantitative information about the structure of this dynamic tissue, other anatomical methods that could be applied to better define the relationships between structure and function will be discussed. These include confocal microscopy, to examine the dynamic physiological interactions of the different tissue compartments, and low-temperature electron microscopy techniques such as cryosubstitution, to allow better access to the biochemical information resident in the tissue. The complex and dynamic nature of the tissue requires a multidisciplinary approach; central to these investigations is a comprehensive understanding of its fine structure.
Serine proteases of the complement system
Authors: Sim R. B. , Laich A.
Published: August 2015

The complement system in blood plasma is a major mediator of innate immune defence. The function of complement is to recognize, then opsonize or lyse, particulate materials, including bacteria, yeasts and other microrganisms, host cell debris and altered host cells. Recognition occurs by binding of complement proteins to charge or saccharide arrays. After recognition, a series of serine proteases is activated, culminating in the assembly of complex unstable proteases called C3/C5 convertases. These activate the complement protein C3, which acts as an opsonin. The complement serine proteases include the closely related Clr, Cls, MASPs 1–3 (80–90 kDa), C2 and Factor B (100 kDa), Factor D (25 kDa) and Factor 1 (85 kDa). Each of these has unusually restricted specificity and low enzymic activity. The C1r, C1s and MASP group occur as proenzymes. When activated, they are regulated, like many plasma serine proteases, by a serpin, C1-inhibitor. C2 and Factor B, however, have complex multiple regulation by a group of complement proteins called the Regulation of Complement Activation (or RCA) proteins, whereas Factors I and D appear to have no natural inhibitors. Advances in structure determination and protein-protein interaction properties are leading to a more detailed understanding of the complement-system proteases, and are indicating possible new routes for potential therapeutic control of complement.
Making light of it: the role of plant haem oxygenases in phytochrome chromophore synthesis
Authors: Terry M. J. , Linley P. J. , Kohchi T.
Published: August 2015

The haem oxygenase (HO) enzyme catalyses the oxidation of haem to biliverdin IXα, CO and Fe2+, and performs a wide variety of roles in Nature, including degradation of haem from haemoglobin, iron acquisition and phycobilin biosynthesis. In plants, HOs are required for the synthesis of the chromophore of the phytochrome family of photoreceptors. There are four HO genes in the Arabidopsis genome. Analysis of a mutant deficient in HO1 (the hy1 mutant) has demonstrated that this plastid-localized protein is the major HO in the phytochrome chromophore synthesis pathway. HO2 may also have a minor role in this pathway, but our understanding of the divergent roles of this small gene family is still far from complete.
Binding sites of lipophilic quinone and quinone analogue inhibitors in the cytochrome b6f complex of oxygenic photosynthesis
Authors: Cramer W.A. , Zhang H. , Yan J. , Kurisu G.
Published: August 2015

The main structural features of the cytochrome b6f complex, solved to 3.0–3.1 Å (1 Å=10−10 m) in the cyanobacterium Mastigocladus laminosus and the green alga Chlamydomonas reinhardtii are discussed. The discussion is focused on the binding sites of plastoquinone and quinone analogue inhibitors discerned in the structure. These sites mark the pathway(s) of quinone translocation across the complex.
Quantification of the effects of melittin on liposome structure
Authors: Popplewell J. , Freeman N. , Carrington S. , Ronan G. , McDonnell C. , Ford R.C.
Published: August 2015

An optical technique, dual-polarization interferometry, has been used to examine lipid structures at the solid/liquid interface. Changes in the lipid structures, in real time, were examined as a consequence of challenging them with a peptide (melittin) that is known to induce liposome rupture. This work suggests that it should be possible to obtain a better understanding of the detail of the melittin rupture process.
The myo-inositol-1-phosphate synthase gene is essential in Trypanosoma brucei
Authors: Martin K.L. , Smith T.K.
Published: August 2015

The de novo synthesis of myo-inositol occurs via a two-step process: first, glucose 6-phosphate is converted into inositol 1-phosphate by an INO1 (myo-inositol-1-phosphate synthase; EC 5.5.1.4); then, it is dephosphorylated by an inositol monophosphatase. The myo-inositol can then be incorporated into PI (phosphatidylinositol), which is utilized in a variety of cellular functions, including the biosynthesis of GPI (glycosylphosphatidylinositol) anchors. A putative INO1 was identified in the Trypanosoma brucei genome database and, by recombinant expression in Escherichia coli, was shown to be a catalytically active INO1. To investigate the importance of INO1, we created a conditional knockout, which, under non-permissive conditions, showed that INO1 is an essential gene in bloodstream form T. brucei and that the de novo synthesized myo-inositol is used for the formation of PI and GPI anchors.
A molecular understanding of the catalytic cycle of the nucleotide-binding domain of the ABC transporter HlyB
Authors: Zaitseva J. , Jenewein S. , Oswald C. , Jumpertz T. , Holland I.B. , Schmitt L.
Published: August 2015

The ABC transporter (ATP-binding-cassette transporter) HlyB (haemolysin B) is the central element of a type I secretion machinery, dedicated to the secretion of the toxin HlyA in Escherichia coli. In addition to the ABC transporter, two other indispensable elements are necessary for the secretion of the toxin across two membranes in a single step: the transenvelope protein HlyD and the outer membrane protein TolC. Despite the fact that the hydrolysis of ATP by HlyB fuels secretion of HlyA, the essential features of the underlying transport mechanism remain an enigma. Similar to all other ABC transporters, ranging from bacteria to man, HlyB is composed of two NBDs (nucleotide-binding domains) and two transmembrane domains. Here we summarize our detailed biochemical, biophysical and structural studies aimed at an understanding of the molecular principles of how ATP-hydrolysis is coupled to energy transduction, including the conformational changes occurring during the catalytic cycle, leading to substrate transport. We have obtained individual crystal structures for each single ground state of the catalytic cycle. From these and other biochemical and mutational studies, we shall provide a detailed molecular picture of the steps governing intramolecular communication and the utilization of chemical energy, due to ATP hydrolysis, in relation to resulting structural changes within the NBD. These data will be summarized in a general model to explain how these molecular machines achieve translocation of molecules across biological membranes.
Crystallographic and single-particle analyses of native- and nucleotide-bound forms of the cystic fibrosis transmembrane conductance regulator (CFTR) protein
Authors: Awayn N.H. , Rosenberg M.F. , Kamis A.B. , Aleksandrov L.A. , Riordan J.R. , Ford R.C.
Published: August 2015

Cystic fibrosis, one of the major human inherited diseases, is caused by defects in the CFTR (cystic fibrosis transmembrane conductance regulator), a cell-membrane protein. CFTR acts as a chloride channel which can be opened by ATP. Low-resolution structural studies of purified recombinant human CFTR are described in the present paper. Localization of the C-terminal decahistidine tag in CFTR was achieved by Ni2+-nitriloacetate nanogold labelling, followed by electron microscopy and single-particle analysis. The presence of the gold label appears to improve the single-particle-alignment procedure. Projection structures of CFTR from two-dimensional crystals analysed by electron crystallography displayed two alternative conformational states in the presence of nucleotide and nanogold, but only one form of the protein was observed in the quiescent (nucleotide-free) state.
Do protein–lipid interactions determine the recognition of transmembrane helices at the ER translocon?
Authors: White S.H. , von Heijne G.
Published: August 2015

Membrane-protein integration, folding and assembly processes in vivo depend on complex targeting, translocation, chaperoning, and sorting machineries that somehow read the ‘molecular code’ built into the nascent polypeptide, ultimately producing a properly folded protein integrated into the correct target membrane. Although the main molecular constituents and the basic mechanistic principles of many of these machines are known in outline, the codes remain poorly defined and there is little quantitative information on how protein sequence affects the final structure of membrane proteins. By carefully designing model protein constructs, we have derived the first true biological hydrophobicity scale and have been able to get a first impression of how the position of a given type of residue within a transmembrane segment affects its ability to promote membrane insertion.
Diet-induced obesity in the Sprague–Dawley rat: dietary manipulations and their effect on hypothalamic neuropeptide energy balance systems
Authors: Mercer J.G. , Archer Z.A.
Published: August 2015

The SD (Sprague–Dawley) rat model of DIO (diet-induced obesity) is reported to exhibit a clear segregation into susceptible and resistant subpopulations shortly after transfer to a HE (high energy) diet. This does not appear to be the case for rats sourced in the U.K., where body weight gain on obesogenic HE diet is normally distributed, as might be anticipated for a polygenic trait in an outbred population. Many of the energy balance effects of dietary manipulation in this model (e.g. supplementation of HE diet with the liquid diet, Ensure; energy intake and defence of body weight following withdrawal of obesogenic diet) appear to be characteristics of the diets being manipulated rather than subject traits. The activities of energy balance-related hypothalamic signals are affected by diet and the development of DIO, but may not be able to differentiate between different diets and the relative levels of obesity that develop.
α-Synuclein aggregation in neurodegenerative diseases and its inhibition as a potential therapeutic strategy
Authors: Paleologou K.E. , Irvine G.B. , El-Agnaf O.M.A.
Published: August 2015

There is strong evidence for the involvement of α-synuclein in the pathologies of several neurodegenerative disorders, including PD (Parkinson's disease). Development of disease appears to be linked to processes that increase the rate at which α-synuclein forms aggregates. These processes include increased protein concentration (via either increased rate of synthesis or decreased rate of degradation), and altered forms of α-synuclein (such as truncations, missense mutations, or chemical modifications by oxidative reactions). Aggregated forms of the protein are toxic to cells and one therapeutic strategy would be to reduce the rate at which aggregation occurs. To this end we have designed several peptides that reduce α-synuclein aggregation. A cell-permeable version of one such peptide was able to inhibit the DNA damage induced by Fe(II) in neuronal cells transfected with α-synuclein (A53T), a familial PD-associated mutation.
Negative feedback in NO/cGMP signalling
Authors: Koesling D. , Mullershausen F. , Lange A. , Friebe A. , Mergia E. , Wagner C. , Russwurm M.
Published: August 2015

Most of the effects of the signalling molecule nitric oxide (NO) are mediated by the stimulation of the NO-sensitive GC (guanylate cyclase) and the subsequent increase in cGMP formation. The enzyme contains a prosthetic haem group, which mediates NO stimulation. In addition to the physiological activator NO, NO-sensitizers like the substance YC-1 sensitize the enzyme towards NO and may therefore have important pharmacological implications. Two isoforms of NO-sensitive GC have been identified to date that share regulatory properties, but differ in the subcellular localization. The more ubiquitously expressed α1β1 heterodimer and the α2β1 isoform are mainly expressed in brain. In intact cells, NO-induced cGMP signalling not only depends on cGMP formation, but is also critically determined by the activity of the enzymes responsible for cGMP degradation, e.g. PDE5 (phosphodiesterase 5). Recently, direct activation of PDE5 by cGMP was demonstrated, limiting the cGMP increase and thus functioning as a negative feedback. As the cGMP-induced PDE5 activation turned out to be sustained, in the range of hours, it is probably responsible for the NO-induced desensitization observed within NO/cGMP signalling.
Mitochondrial fatty acid synthesis and maintenance of respiratory competent mitochondria in yeast
Authors: Hiltunen J.K. , Okubo F. , Kursu V.A.S. , Autio K.J. , Kastaniotis A.J.
Published: August 2015

Mitochondrial FAS (fatty acid synthesis) of type II is a widely conserved process in eukaryotic organisms, with particular importance for respiratory competence and mitochondrial morphology maintenance in Saccharomyces cerevisiae. The recent characterization of three missing enzymes completes the pathway. Etr1p (enoyl thioester reductase) was identified via purification of the protein followed by molecular cloning. To study the link between FAS and cell respiration further, we also created a yeast strain that has FabI enoyl-ACP (acyl-carrier protein) reductase gene from Escherichia coli engineered to carry a mitochondrial targeting sequence in the genome, replacing the endogenous ETR1 gene. This strain is respiratory competent, but unlike the ETR1 wild-type strain, it is sensitive to triclosan on media containing only non-fermentable carbon source. A colony-colour-sectoring screen was applied for cloning of YHR067w/RMD12, the gene encoding mitochondrial 3-hydroxyacyl-ACP dehydratase (Htd2/Yhr067p), the last missing component of the mitochondrial FAS. Finally, Hfa1p was shown to be the mitochondrial acetyl-CoA carboxylase.
Signalling functions for sphingolipid long-chain bases in Saccharomyces cerevisiae
Authors: Liu K. , Zhang X. , Sumanasekera C. , Lester R.L. , Dickson R.C.
Published: August 2015

Over the past several years, studies of sphingolipid functions in the baker's yeast Saccharomyces cerevisiae have revealed that the sphingoid LCBs (long-chain bases), dihydrosphingosine and PHS (phytosphingosine), are important signalling molecules or second messengers under heat stress and during non-stressed conditions. LCBs are now recognized as regulators of AGC-type protein kinase (where AGC stands for protein kinases A, G and C) Pkh1 and Pkh2, which are homologues of mammalian phosphoinositide-dependent protein kinase 1. LCBs were previously shown to activate Pkh1 and Pkh2, which then activate the downstream protein kinase Pkc1. We have recently demonstrated that PHS stimulates Pkh1 to activate additional downstream kinases including Ypk1, Ypk2 and Sch9. We have also found that PHS acts downstream of Pkh1 and partially activates Ypk1, Ypk2 and Sch9. These kinases control a wide range of cellular processes including growth, cell wall integrity, stress resistance, endocytosis and aging. As we learn more about the cellular processes controlled by Ypk1, Ypk2 and Sch9, we will have a far greater appreciation of LCBs as second messengers.
SH3 domain-containing proteins and the actin cytoskeleton in yeast
Authors: Mirey G. , Soulard A. , Orange C. , Friant S. , Winsor B.
Published: August 2015

SH3 (Src homology-3) domains are involved in protein–protein interactions through proline-rich domains. Many SH3-containing proteins are implicated in actin cytoskeleton organization. The aim of our ongoing work is to study the functions of the SH3-containing proteins in actin cytoskeleton regulation. The yeast Saccharomyces cerevisiae proteome includes 29 SH3 domains distributed in 25 proteins. We have examined the direct involvement of these SH3 domains in actin polymerization using an in vitro polymerization assay on GST (glutathione S-transferase)–SH3-coated beads. As expected, not all SH3 domains show polymerization activity, and many recruit distinct partners as assessed by microscopy and pull-down experiments. One such partner, Las17p, the yeast homologue of WASP (Wiskott–Aldrich syndrome protein), was assayed because it stimulates actin nucleation via the Arp2/3 (actin-related protein 2/3) complex. Ultimately, proteins involved in specific biological processes, such as membrane trafficking, may also be recruited by some of these SH3 domains, shedding light on the SH3-containing proteins and actin cytoskeleton functions in these processes.