bluuebunny/crossref_metadata_2025 huggingface dataset:
Device design and materials optimization of conformal coating for islets of Langerhans
Authors: Tomei Alice A. , Manzoli Vita , Fraker Christopher A. , Giraldo Jaime , Velluto Diana , Najjar Mejdi , Pileggi Antonello , Molano R. Damaris , Ricordi Camillo , Stabler Cherie L. , Hubbell Jeffrey A.
Published: July 2014

Significance

Cell encapsulation with biocompatible and permeable hydrogels may allow transplantation without immunosuppression. As an alternative to standard microencapsulation approaches that create single-sized capsules around cell clusters of different sizes, we have designed and optimized a novel approach for conformal coating of islets of Langerhans, resulting in thin, complete, and uniform coatings of similar thickness on differently sized islets. Coated islets exhibited no delay in glucose-stimulated insulin release or loss of function during culture, which is often observed with naked islets. The conformal coating reduces transplant volume relative to traditional encapsulation approaches. When transplanted in syngeneic diabetic mice, conformally coated islets restored and maintained euglycemia for more than 100 d with no foreign body reaction and normal revascularization.
Inactivity of human β,β-carotene-9′,10′-dioxygenase (BCO2) underlies retinal accumulation of the human macular carotenoid pigment
Authors: Li Binxing , Vachali Preejith P. , Gorusupudi Aruna , Shen Zhengqing , Sharifzadeh Hassan , Besch Brian M. , Nelson Kelly , Horvath Madeleine M. , Frederick Jeanne M. , Baehr Wolfgang , Bernstein Paul S.
Published: July 2014

Significance

Among mammals, only primates can accumulate high levels of xanthophyll carotenoids in their retinas. We discovered that, unlike most other mammals, the major xanthophyll cleavage enzyme β,β-carotene-9′,10′-dioxygenase (BCO2) is inactive in humans, explaining the unique accumulation of lutein, zeaxanthin, and
meso
-zeaxanthin in primate macula. We confirmed this discovery by demonstrating that BCO2 knockout mice accumulate zeaxanthin in their retinas.
A self-regulating template in human telomerase
Authors: Brown Andrew F. , Podlevsky Joshua D. , Qi Xiaodong , Chen Yinnan , Xie Mingyi , Chen Julian J.-L.
Published: July 2014

Significance

The telomerase enzyme is essential for maintaining the replicative capacity of highly prolific cells, such as stem cells and cancer, by synthesizing telomeric DNA onto chromosome ends. Telomerase functions as an RNA–protein complex with an integral telomerase RNA (TR) component. While the templates from all other reverse transcriptases (RTs) merely specify the sequence for nucleotide addition, we found that the human TR template is embedded with a single-residue pausing signal for regulating DNA synthesis. Mutation of this pausing signal alters the fundamental function of telomerase for synthesizing exact telomeric DNA repeats. This is the first instance, to our knowledge, of a single-residue pausing signal found in the RNA template of an RT.
CTSH
regulates β-cell function and disease progression in newly diagnosed type 1 diabetes patients
Authors: Fløyel Tina , Brorsson Caroline , Nielsen Lotte B. , Miani Michela , Bang-Berthelsen Claus Heiner , Friedrichsen Martin , Overgaard Anne Julie , Berchtold Lukas A. , Wiberg Anna , Poulsen Pernille , Hansen Lars , Rosinger Silke , Boehm Bernhard O. , Ram Ramesh , Nguyen Quang , Mehta Munish , Morahan Grant , Concannon Patrick , Bergholdt Regine , Nielsen Jens H. , Reinheckel Thomas , von Herrath Matthias , Vaag Allan , Eizirik Decio Laks , Mortensen Henrik B. , Størling Joachim , Pociot Flemming
Published: July 2014

Significance

In type 1 diabetes (T1D), the insulin-producing pancreatic β-cells are destroyed by the immune system. Both genetic and environmental factors contribute to T1D risk. Candidate genes for T1D identified by genome-wide association studies have been proposed to act at both the immune system and the β-cell levels. This study shows that the risk variant rs3825932 in the candidate gene cathepsin H (
CTSH
) predicts β-cell function in both model systems and human T1D. Collectively, our data indicate that higher
CTSH
expression in β-cells may protect against immune-mediated damage and preserve β-cell function, thereby representing a possible therapeutic target. Our study reinforces the concept that candidate genes for T1D may affect disease progression by modulating survival and function of the β-cells.
Primary cilia enhance kisspeptin receptor signaling on gonadotropin-releasing hormone neurons
Authors: Koemeter-Cox Andrew I. , Sherwood Thomas W. , Green Jill A. , Steiner Robert A. , Berbari Nicolas F. , Yoder Bradley K. , Kauffman Alexander S. , Monsma Paula C. , Brown Anthony , Askwith Candice C. , Mykytyn Kirk
Published: July 2014

Significance

Mammalian central neurons typically possess solitary appendages called primary cilia. These rod-shaped protrusions are enriched for signaling proteins, suggesting they respond to extracellular neuromodulators. Yet, neuronal ciliary signaling has never been observed and the roles of cilia in neuronal function are unclear. This study identifies Kiss1r, which plays a key role in reproductive function, as a novel ciliary receptor on gonadotropin-releasing hormone (GnRH) neurons. GnRH neurons possess multiple Kiss1r-positive cilia and the proportion of multiple cilia increases in parallel with pubertal maturation. Ablation of Kiss1r-positive cilia on GnRH neurons does not affect neuron migration or sexual maturation. However, kisspeptin-mediated increases in GnRH neuron firing rate are reduced in the absence of cilia. Thus, cilia enhance Kiss1r signaling on GnRH neurons.
Elite male faculty in the life sciences employ fewer women
Authors: Sheltzer Jason M. , Smith Joan C.
Published: July 2014

Significance

Despite decades of progress, men still greatly outnumber women among biology faculty in the United States. Here, we show that high-achieving faculty members who are male train 10–40% fewer women in their laboratories relative to the number of women trained by other investigators. These skewed employment patterns may result from self-selection among female scientists or they may result from conscious or unconscious bias on the part of some faculty members. The dearth of women who are trained in these laboratories likely limits the number of female candidates who are most competitive for faculty job searches.
A single macrolichen constitutes hundreds of unrecognized species
Authors: Lücking Robert , Dal-Forno Manuela , Sikaroodi Masoumeh , Gillevet Patrick M. , Bungartz Frank , Moncada Bibiana , Yánez-Ayabaca Alba , Chaves José Luis , Coca Luis Fernando , Lawrey James D.
Published: July 2014

Significance

Macrolichens are considered to be well known, including the tropical montane basidiolichen fungus
Dictyonema glabratum
, also known as
Cora pavonia
, an important component of threatened paramo ecosystems, where it acts as a biological fertilizer due to its ability to fix atmospheric nitrogen. This lichen was long believed to represent a single species, but after revising this number to 16 in two genera (
Cora
and
Corella
), here we show that at least 126 phylogenetically and morphologically distinct species are contained within this group, with statistical analysis predicting more than 400. Our findings underline the importance of accurately documenting species richness for conservation purposes and support the notion of neotropical paramos as hotspots of recent diversification in plants, animals, and fungi.
Intravenously administered nanoparticles increase survival following blast trauma
Authors: Lashof-Sullivan Margaret M. , Shoffstall Erin , Atkins Kristyn T. , Keane Nickolas , Bir Cynthia , VandeVord Pamela , Lavik Erin B.
Published: July 2014

Significance

We have developed hemostatic nanoparticles that reduce bleeding and increase survival in both the short term and long term following the complex injuries sustained during blast trauma. This treatment has the potential to be deployed by first responders to save lives.
Evolution of the ribosome at atomic resolution
Authors: Petrov Anton S. , Bernier Chad R. , Hsiao Chiaolong , Norris Ashlyn M. , Kovacs Nicholas A. , Waterbury Chris C. , Stepanov Victor G. , Harvey Stephen C. , Fox George E. , Wartell Roger M. , Hud Nicholas V. , Williams Loren Dean
Published: July 2014

Significance

Ribosomes exist in every cell and are responsible for translation from mRNA to protein. The structure of the ribosomal common core is highly conserved in all living species, while the outer regions of the ribosome are variable. Ribosomal RNA of eukaryotes contains expansion segments accreted onto the surface of the core, which is nearly identical in structure to that in prokaryotic ribosomes. Comparing eukaryotic and prokaryotic ribosomes allows us to identify 3D insertion fingerprints of the expansion segments. Similar fingerprints allow us to analyze the common core and detect ancestral expansion segments within it. We construct a molecular model of ribosomal evolution starting from primordial biological systems near the dawn of life, culminating with relatively recent changes specific to metazoans.
Two miRNA clusters,
miR-34b/c
and
miR-449
, are essential for normal brain development, motile ciliogenesis, and spermatogenesis
Authors: Wu Jingwen , Bao Jianqiang , Kim Minkyung , Yuan Shuiqiao , Tang Chong , Zheng Huili , Mastick Grant S. , Xu Chen , Yan Wei
Published: July 2014

Significance

Most of the single miRNA gene knockouts display no developmental phenotype. Here, we report that simultaneous inactivation of two functionally overlapping miRNAs,
miR-34b/c
and
miR-449,
led to a sexually dimorphic partial perinatal lethality, growth retardation and sterility. Multiple underlying developmental defects, including underdevelopment of the basal forebrain structures, a lack of motile cilia in trachea and oviduct, severely disrupted spermatogenesis and oligoasthenoteratozoospermia, result from the dysregulation of ∼240 target genes that are mainly involved in three major cellular functions, including cell fate control, brain development and microtubule dynamics. This study provides physiological evidence demonstrating an essential role of
miR-34b/c
and
miR-449
in normal brain development, motile ciliogenesis and spermatogenesis.
Serial propagation of distinct strains of Aβ prions from Alzheimer’s disease patients
Authors: Watts Joel C. , Condello Carlo , Stöhr Jan , Oehler Abby , Lee Joanne , DeArmond Stephen J. , Lannfelt Lars , Ingelsson Martin , Giles Kurt , Prusiner Stanley B.
Published: July 2014

Significance

The amyloid-β (Aβ) peptide, which plays a central role in Alzheimer’s disease (AD) pathogenesis, exhibits many properties that are reminiscent of prions (self-propagating proteins that cause neurodegenerative disorders, such as mad cow disease). In the human prion diseases, distinct strains of prions can be distinguished, and therefore, we asked whether different strains of Aβ aggregates might exist in the brains of AD patients. Inoculation of transgenic mice with brain samples from patients with two different heritable forms of AD produced two distinct patterns of cerebral Aβ deposition, and these differences were maintained on serial passage. We conclude that distinct strains of Aβ can be discerned in AD patients, which may help to explain the clinical heterogeneity observed in the disease.
Genetic analysis of the Complexin trans-clamping model for cross-linking SNARE complexes in vivo
Authors: Cho Richard W. , Kümmel Daniel , Li Feng , Baguley Stephanie Wood , Coleman Jeff , Rothman James E. , Littleton J. Troy
Published: July 2014

Significance

Synaptic vesicle fusion at synapses is the primary mechanism by which neurons communicate. A highly conserved membrane fusion machine known as the SNARE complex mediates this process. In addition, neuronal SNARE-binding regulatory proteins have evolved to control the kinetics and speed of SNARE assembly at synapses. One such SNARE-binding protein, Complexin, has been found to inhibit synaptic vesicle fusion in the absence of an action potential and activate SNARE-mediated release during stimulation. Here we examine molecular models for how Complexin can mediate these unique effects on the SNARE fusion machine using genetic rescue experiments in
Drosophila
. We find that alternate SNARE-binding mechanisms by Complexin are likely to contribute to distinct inhibitory and activating functions in vivo.
Affinity maturation in an HIV broadly neutralizing B-cell lineage through reorientation of variable domains
Authors: Fera Daniela , Schmidt Aaron G. , Haynes Barton F. , Gao Feng , Liao Hua-Xin , Kepler Thomas B. , Harrison Stephen C.
Published: July 2014

Significance

An HIV vaccine must induce antibodies [broadly neutralizing antibodies (bnAbs)] that neutralize many viral variants. Determining pathways of antibody affinity maturation that have led to specific bnAbs and tracking coevolution in an infected individual of virus and antibody will define characteristics of immunogens that might elicit broad responses. We have followed, in an infected individual, one round of coevolution of viral envelope with antibodies from a single lineage. Insertions into a loop (V5) in gp120 of autologous viruses allowed escape from neutralization by antibodies in this lineage; antibody affinity maturation shifted the relative orientation of the light- and heavy-chain variable domains, allowing binding to envelopes with augmented V5. The results illustrate a mechanism of affinity maturation through mutation outside the antigen combining site.
Morphological variability at the morphosyntactic/semantic interface
Authors: Massery Laurie A. , Fuentes Claudio
Published: July 2014

Recent literature in second language acquisition shows that syntax-driven structures give way to successful modal interpretation and morphological production, while discourse-dependent environments do not (Sorace, 2005; Tsimpli & Sorace, 2006, Borganovo, Bruhn de Garavito, & Prévost, 2008; Iverson, Kempchinsky, & Rothman, 2008). It has also been suggested that discourse-dependent environments involve both structural and pragmatic knowledge of L2, which intersect at the syntax-discourse interface (Sorace, 2005; Iverson, Kempchinsky, & Rothman, 2008), thereby requiring a multi-layered understanding of the target language. The present study contributes to this line of research by further examining morphological variability (Prévost & White, 2000; Sorace, 2000; Sorace, 2005; Iverson, Kempchinsky, & Rothman, 2008; Slabakova, 2009) in L2 acquisition at the morphosyntactic-semantic interface, following the work of Borganovo, Bruhn de Garavito, & Prévost (2008). The results of our study reveal that learners, even at advanced stages of acquisition, perform poorly in epistemic environments where syntax and discourse intersect. In such environments, there appears to be an interaction with pragmatics (cf. Iverson, Kempchinsky, & Rothman, 2008) that causes learners to opt for the indicative mood, even when the subjunctive is prescriptively required. Unlike deontic modality, which is essentially syntax-driven, epistemic modality requires structural knowledge, as well as knowledge from other domains (Sorace, 2005). Our study reveals that learners at all levels of instruction performed better in “purely syntactic” environments of deontic modality than in pragmatically challenging epistemic environments.
C. elegans Anillin proteins regulate intercellular bridge stability and germline syncytial organization
Authors: Amini Rana , Goupil Eugénie , Labella Sara , Zetka Monique , Maddox Amy S. , Labbé Jean-Claude , Chartier Nicolas T.
Published: July 2014

Cytokinesis generally produces two separate daughter cells, but in some tissues daughter nuclei remain connected to a shared cytoplasm, or syncytium, through incomplete cytokinesis. How syncytia form remains poorly understood. We studied syncytial formation in the Caenorhabditis elegans germline, in which germ cells connect to a shared cytoplasm core (the rachis) via intercellular bridges. We found that syncytial architecture initiates early in larval development, and germ cells become progressively interconnected until adulthood. The short Anillin family scaffold protein ANI-2 is enriched at intercellular bridges from the onset of germ cell specification, and ANI-2 loss resulted in destabilization of intercellular bridges and germ cell multinucleation defects. These defects were partially rescued by depleting the canonical Anillin ANI-1 or blocking cytoplasmic streaming. ANI-2 is also required for elastic deformation of the gonad during ovulation. We propose that ANI-2 promotes germ cell syncytial organization and allows for compensation of the mechanical stress associated with oogenesis by conferring stability and elasticity to germ cell intercellular bridges.
Accurate phosphoregulation of kinetochore–microtubule affinity requires unconstrained molecular interactions
Authors: Zaytsev Anatoly V. , Sundin Lynsie J.R. , DeLuca Keith F. , Grishchuk Ekaterina L. , DeLuca Jennifer G.
Published: July 2014

Accurate chromosome segregation relies on dynamic interactions between microtubules (MTs) and the NDC80 complex, a major kinetochore MT-binding component. Phosphorylation at multiple residues of its Hec1 subunit may tune kinetochore–MT binding affinity for diverse mitotic functions, but molecular details of such phosphoregulation remain elusive. Using quantitative analyses of mitotic progression in mammalian cells, we show that Hec1 phosphorylation provides graded control of kinetochore–MT affinity. In contrast, modeling the kinetochore interface with repetitive MT binding sites predicts a switchlike response. To reconcile these findings, we hypothesize that interactions between NDC80 complexes and MTs are not constrained, i.e., the NDC80 complexes can alternate their binding between adjacent kinetochore MTs. Experiments using cells with phosphomimetic Hec1 mutants corroborate predictions of such a model but not of the repetitive sites model. We propose that accurate regulation of kinetochore–MT affinity is driven by incremental phosphorylation of an NDC80 molecular “lawn,” in which the NDC80–MT bonds reorganize dynamically in response to the number and stability of MT attachments.
The rigidity theorems of self-shrinkers
Authors: Ding Qi , Xin Y. L.
Published: July 2014

By using certain techniques developed in minimal submanifold theory, we study the rigidity problem for self-shrinkers in the present paper. We prove rigidity results for the squared norm of the second fundamental form of self-shrinkers, either under pointwise conditions or under integral conditions.
Material properties of Northeast Pacific zooplankton
Authors: Becker Kaylyn N. , Warren Joseph D.
Published: July 2014

We measured the density and sound speed contrasts relative to seawater of Northeast Pacific zooplankton. The density contrast (g) was measured for euphausiids, decapods (Sergestes similis), amphipods (Primno macropa, Phronima sp., and Hyperiid spp.), siphonophore bracts, chaetognaths, larval fish, crab megalopae, larval squid, and medusae. Morphometric data (length, width, and height) were collected for these taxa. Density contrasts varied within and between zooplankton taxa. The mean and standard deviation (s.d.) for euphausiid density contrast were 1.059 ± 0.009. Relationships between zooplankton density contrast and morphometric measurements, geographic location, and environmental conditions were investigated. Site had a significant effect on euphausiid density contrast. Density contrasts of euphausiids collected in the same geographic area ∼4–10 d apart were significantly higher (p< 0.001). Sound speed contrast (h) was measured for euphausiids and pelagic decapods (S. similis) and it varied between taxa. The mean and s.d. for euphausiid sound speed were 1.019 ± 0.009. Euphausiid mass was calculated from measured density and volume, and a relationship between euphausiid mass and length was produced. We determined that euphausiid volume could be accurately estimated from two-dimensional measurements of animal body shape, and that biomass (or biovolume) could be accurately calculated from digital photographs of animals. Data from this study can improve the accuracy of theoretical acoustic scattering models for these taxa, resulting in more accurate estimates of zooplankton biomass in this region.
Dynamics of T-cell IFN-γ and miR-29a expression during active pulmonary tuberculosis
Authors: Afum-Adjei Awuah Anthony , Ueberberg Bianca , Owusu-Dabo Ellis , Frempong Margaret , Jacobsen Marc
Published: July 2014

IFN-γ is crucial for protection against Mycobacterium tuberculosis. miR-29 was recently shown to non-redundantly inhibit IFN-γ. Here, we investigated IFN-γ and miR-29a expression dynamics of CD4+ T cells from patients during active tuberculosis (TB) (n = 32) and in household contacts who were latently M. tuberculosis infected (n = 19) from Ghana. Whereas M. tuberculosis-specific IFN-γ expression was similar during TB chemotherapy, superantigen stimulation indicated generally impaired IFN-γ expression in TB patients. No interdependency between miR-29a and IFN-γ expression of T cells was observed. However, miR-29a was differentially expressed in T cells during chemotherapy. We concluded that differential miR-29a expression in active TB was not causative for impaired IFN-γ expression.
A fully enzymatic method for site-directed spin labeling of long RNA
Authors: Lebars Isabelle , Vileno Bertrand , Bourbigot Sarah , Turek Philippe , Wolff Philippe , Kieffer Bruno
Published: July 2014

Site-directed spin labeling is emerging as an essential tool to investigate the structural and dynamical features of RNA. We propose here an enzymatic method, which allows the insertion of a paramagnetic center at a specific position in an RNA molecule. The technique is based on a segmental approach using a ligation protocol with T4 RNA ligase 2. One transcribed acceptor RNA is ligated to a donor RNA in which a thio-modified nucleotide is introduced at its 5′-end by in vitro transcription with T7 RNA polymerase. The paramagnetic thiol-specific reagent is subsequently attached to the RNA ligation product. This novel strategy is demonstrated by introducing a paramagnetic probe into the 55 nucleotides long RNA corresponding to K-turn and Specifier Loop domains from the Bacillus subtilis tyrS T-Box leader RNA. The efficiency of the coupling reaction and the quality of the resulting spin-labeled RNA were assessed by Mass Spectrometry, Electron Paramagnetic Resonance (EPR) and Nuclear Magnetic Resonance (NMR). This method enables various combinations of isotopic segmental labeling and spin labeling schemes, a strategy that will be of particular interest to investigate the structural and dynamical properties of large RNA complexes by NMR and EPR spectroscopies.
Redox Systems in Botrytis cinerea: Impact on Development and Virulence
Authors: Viefhues Anne , Heller Jens , Temme Nora , Tudzynski Paul
Published: July 2014

The thioredoxin system is of great importance for maintenance of cellular redox homeostasis. Here, we show that it has a severe influence on virulence of Botrytis cinerea, demonstrating that redox processes are important for host-pathogen interactions in this necrotrophic plant pathogen. The thioredoxin system is composed of two enzymes, the thioredoxin and the thioredoxin reductase. We identified two genes encoding for thioredoxins (bctrx1, bctrx2) and one gene encoding for a thioredoxin reductase (bctrr1) in the genome of B. cinerea. Knockout mutants of bctrx1 and bctrr1 were severely impaired in virulence and more sensitive to oxidative stress. Additionally, Δbctrr1 showed enhanced H2O2 production and retarded growth. To investigate the impact of the second major cellular redox system, glutathione, we generated deletion mutants for two glutathione reductase genes. The effects were only marginal; deletion of bcglr1 resulted in reduced germination and, correspondingly, to retarded infection as well as reduced growth on minimal medium, whereas bcglr2 deletion had no distinctive phenotype. In summary, we showed that the balanced redox status maintained by the thioredoxin system is essential for development and pathogenesis of B. cinerea, whereas the second major cellular redox system, the glutathione system, seems to have only minor impact on these processes.
Multi-scale and multi-domain computational astrophysics
Authors: van Elteren Arjen , Pelupessy Inti , Zwart Simon Portegies
Published: July 2014

Astronomical phenomena are governed by processes on all spatial and temporal scales, ranging from days to the age of the Universe (13.8 Gyr) as well as from kilometre size up to the size of the Universe. This enormous range in scales is contrived, but as long as there is a physical connection between the smallest and largest scales it is important to be able to resolve them all, and for the study of many astronomical phenomena this governance is present. Although covering all these scales is a challenge for numerical modellers, the most challenging aspect is the equally broad and complex range in physics, and the way in which these processes propagate through all scales. In our recent effort to cover all scales and all relevant physical processes on these scales, we have designed the Astrophysics Multipurpose Software Environment (AMUSE). AMUSE is a Python-based framework with production quality community codes and provides a specialized environment to connect this plethora of solvers to a homogeneous problem-solving environment.
Spirosoma oryzae sp. nov., isolated from rice soil and emended description of the genus Spirosoma
Authors: Ahn Jae-Hyung , Weon Hang-Yeon , Kim Soo-Jin , Hong Seung-Beom , Seok Soon-Ja , Kwon Soon-Wo
Published: July 2014

A bacterial strain, designated RHs22T, was isolated from a soil sample cultivated with rice in the Suwon region of South Korea. The cells were aerobic, Gram-stain-negative, non-spore-forming, non-flagellated rods or occasionally filaments. The strain grew at 10–37 °C (optimum, 28–30 °C), at pH 5.0–10.0 (optimum, 7.0) and in the presence of 0–1 % (w/v) NaCl (optimum, 0 %). Phylogenetically, the strain was closely related to members of the genus

Spirosoma

, as its 16S rRNA gene sequence had similarity of 90.3–92.1 % with respect to those of members of the genus

Spirosoma

, showing the highest sequence similarity with

Spirosoma panaciterrae

DSM 21099T. Strain RHs22T revealed relatively low sequence similarities of less than 90 % with all the other species with validly published names. It contained MK-7 as the predominant menaquinone and summed feature 3 (C16 : 1ω6c and/or C16 : 1ω7c), C16 : 1ω5c, iso-C15 : 0 and iso-C17 : 0 3-OH as the main fatty acids. The polar lipids of strain RHs22T were phosphatidylethanolamine, one unknown aminolipid, two unknown aminophospholipids, one unknown phospholipid and five unknown lipids. The DNA G+C content was 57.0 mol%. Phylogenetic, phenotypic and chemotaxonomic data obtained in this study indicate that strain RHs22T represents a novel species of the genus

Spirosoma

, for which the name Spirosoma oryzae sp. nov. is proposed. The type strain is RHs22T ( = KACC 17324T = DSM 28354T). An emended description of the genus

Spirosoma

is also proposed.
Pseudoruegeria sabulilitoris sp. nov., isolated from seashore sand
Authors: Park Sooyeon , Jung Yong-Taek , Won Sung-Min , Yoon Jung-Hoon
Published: July 2014

A Gram-stain-negative, aerobic, non-spore-forming, non-flagellated and ovoid or rod-shaped bacterial strain, GJMS-35T, was isolated from seashore sand at Geoje Island, South Korea. Strain GJMS-35T grew optimally at 28–30 °C, at pH 7.0–8.0 and in the presence of 2.0 % (w/v) NaCl. Phylogenetic trees based on 16S rRNA gene sequences revealed that strain GJMS-35T clustered with type strains of species of the genus

Pseudoruegeria

, with which it exhibited 96.97–98.42 % 16S rRNA gene sequence similarity. Sequence similarities to the type strains of other recognized species were less than 96.39 %. Strain GJMS-35T contained Q-10 as the predominant ubiquinone and C18 : 1ω7c as the major fatty acid. The major polar lipids of strain GJMS-35T were phosphatidylcholine, phosphatidylglycerol, phosphatidylethanolamine, one unidentified glycolipid, one unidentified aminolipid and one unidentified lipid. The DNA G+C content of strain GJMS-35T was 64.1 mol% and its mean DNA–DNA relatedness values with type strains of three species of the genus

Pseudoruegeria

were 11–21 %. Its differential phenotypic properties, together with its phylogenetic and genetic distinctiveness, revealed that strain GJMS-35T is set apart from other species of the genus

Pseudoruegeria

. On the basis of the data presented, strain GJMS-35T is considered to represent a novel species of the genus

Pseudoruegeria

, for which the name Pseudoruegeria
sabulilitoris sp. nov. is proposed. The type strain is GJMS-35T ( = KCTC 42111T = NBRC 110380T).
Molecular characterization of Streptococcus pyogenes group A isolates from a tertiary hospital in Lebanon
Authors: Karaky Nathalie M. , Araj George F. , Tokajian Sima T.
Published: July 2014

Streptococcus pyogenes[Group AStreptococcus(GAS)] is one of the most important human pathogens, responsible for numerous diseases with diverse clinical manifestations. As the epidemiology of GAS infections evolves, a rapid and reliable characterization of the isolates remains essential for epidemiological analysis and infection control. This study investigated the epidemiological patterns and genetic characteristics of 150 GAS isolates from a tertiary hospital in Lebanon byemmtyping, superantigens (SAgs) detection, PFGE and antibiotic profiling. The results revealed 41 distinctemmtypes, the most prevalent of which wereemm89 (16 %),emm12 (10 %),emm2 (9 %) andemm1 (8 %). Testing for the presence of superantigens showed thatspeB(87 %),ssa(36 %) andspeG(30 %) were predominant. PFGE detected 39 pulsotypes when a similarity cut-off value of 80 % was implemented. Antibiotic-susceptibility testing against seven different classes of antibiotics showed that 9 % of the isolates were resistant to clindamycin, 23 % were resistant to erythromycin and 4 % showed the macrolide–lincosamide–streptogramin B (MLSB) phenotype. The emergence of tetracycline-resistant strains (37 %) was high when compared with previous reports from Lebanon. This study provided comprehensive evidence of the epidemiology of GAS in Lebanon, highlighting the association betweenemmtypes and toxin genes, and providing valuable information about the origin and dissemination of this pathogen.
Structural constraints in the packaging of bluetongue virus genomic segments
Authors: Burkhardt Christiane , Sung Po-Yu , Celma Cristina C. , Roy Polly
Published: July 2014

The mechanism used by bluetongue virus (BTV) to ensure the sorting and packaging of its 10 genomic segments is still poorly understood. In this study, we investigated the packaging constraints for two BTV genomic segments from two different serotypes. Segment 4 (S4) of BTV serotype 9 was mutated sequentially and packaging of mutant ssRNAs was investigated by two newly developed RNA packaging assay systems, one in vivo and the other in vitro. Modelling of the mutated ssRNA followed by biochemical data analysis suggested that a conformational motif formed by interaction of the 5′ and 3′ ends of the molecule was necessary and sufficient for packaging. A similar structural signal was also identified in S8 of BTV serotype 1. Furthermore, the same conformational analysis of secondary structures for positive-sense ssRNAs was used to generate a chimeric segment that maintained the putative packaging motif but contained unrelated internal sequences. This chimeric segment was packaged successfully, confirming that the motif identified directs the correct packaging of the segment.
DNA replication and transcription programs respond to the same chromatin cues
Authors: Lubelsky Yoav , Prinz Joseph A. , DeNapoli Leyna , Li Yulong , Belsky Jason A. , MacAlpine David M.
Published: July 2014

DNA replication is a dynamic process that occurs in a temporal order along each of the chromosomes. A consequence of the temporally coordinated activation of replication origins is the establishment of broad domains (>100 kb) that replicate either early or late in S phase. This partitioning of the genome into early and late replication domains is important for maintaining genome stability, gene dosage, and epigenetic inheritance; however, the molecular mechanisms that define and establish these domains are poorly understood. The modENCODE Project provided an opportunity to investigate the chromatin features that define the Drosophila replication timing program in multiple cell lines. The majority of early and late replicating domains in the Drosophila genome were static across all cell lines; however, a small subset of domains was dynamic and exhibited differences in replication timing between the cell lines. Both origin selection and activation contribute to defining the DNA replication program. Our results suggest that static early and late replicating domains were defined at the level of origin selection (ORC binding) and likely mediated by chromatin accessibility. In contrast, dynamic domains exhibited low ORC densities in both cell types, suggesting that origin activation and not origin selection governs the plasticity of the DNA replication program. Finally, we show that the male-specific early replication of the X chromosome is dependent on the dosage compensation complex (DCC), suggesting that the transcription and replication programs respond to the same chromatin cues. Specifically, MOF-mediated hyperacetylation of H4K16 on the X chromosome promotes both the up-regulation of male-specific transcription and origin activation.
Diversity of miRNAs, siRNAs, and piRNAs across 25 Drosophila cell lines
Authors: Wen Jiayu , Mohammed Jaaved , Bortolamiol-Becet Diane , Tsai Harrison , Robine Nicolas , Westholm Jakub O. , Ladewig Erik , Dai Qi , Okamura Katsutomo , Flynt Alex S. , Zhang Dayu , Andrews Justen , Cherbas Lucy , Kaufman Thomas C. , Cherbas Peter , Siepel Adam , Lai Eric C.
Published: July 2014

We expanded the knowledge base for Drosophila cell line transcriptomes by deeply sequencing their small RNAs. In total, we analyzed more than 1 billion raw reads from 53 libraries across 25 cell lines. We verify reproducibility of biological replicate data sets, determine common and distinct aspects of miRNA expression across cell lines, and infer the global impact of miRNAs on cell line transcriptomes. We next characterize their commonalities and differences in endo-siRNA populations. Interestingly, most cell lines exhibit enhanced TE-siRNA production relative to tissues, suggesting this as a common aspect of cell immortalization. We also broadly extend annotations of cis-NAT-siRNA loci, identifying ones with common expression across diverse cells and tissues, as well as cell-restricted loci. Finally, we characterize small RNAs in a set of ovary-derived cell lines, including somatic cells (OSS and OSC) and a mixed germline/somatic cell population (fGS/OSS) that exhibits ping-pong piRNA signatures. Collectively, the ovary data reveal new genic piRNA loci, including unusual configurations of piRNA-generating regions. Together with the companion analysis of mRNAs described in a previous study, these small RNA data provide comprehensive information on the transcriptional landscape of diverse Drosophila cell lines. These data should encourage broader usage of fly cell lines, beyond the few that are presently in common usage.
Evolution of H3K27me3-marked chromatin is linked to gene expression evolution and to patterns of gene duplication and diversification
Authors: Arthur Robert K. , Ma Lijia , Slattery Matthew , Spokony Rebecca F. , Ostapenko Alexander , Nègre Nicolas , White Kevin P.
Published: July 2014

Histone modifications are critical for the regulation of gene expression, cell type specification, and differentiation. However, evolutionary patterns of key modifications that regulate gene expression in differentiating organisms have not been examined. Here we mapped the genomic locations of the repressive mark histone 3 lysine 27 trimethylation (H3K27me3) in four species of Drosophila, and compared these patterns to those in C. elegans. We found that patterns of H3K27me3 are highly conserved across species, but conservation is substantially weaker among duplicated genes. We further discovered that retropositions are associated with greater evolutionary changes in H3K27me3 and gene expression than tandem duplications, indicating that local chromatin constraints influence duplicated gene evolution. These changes are also associated with concomitant evolution of gene expression. Our findings reveal the strong conservation of genomic architecture governed by an epigenetic mark across distantly related species and the importance of gene duplication in generating novel H3K27me3 profiles.
PiggyBac Transposon-Mediated Cellular Transgenesis in Mammalian Forebrain by In Utero Electroporation
Authors: Chen Fuyi , Maher Brady J. , LoTurco Joseph J.
Published: July 2014

In utero electroporation (IUE) is an effective transfection method for delivering plasmid DNA into neural progenitor cells and neurons of mammalian neocortex in vivo. Although IUE is effective at delivering multiple DNA plasmids into populations of cells, unfortunately plasmids delivered into neural progenitor cells remain largely episomal and often get inactivated or lost after cell division. This results in a form of “birthdate” labeling in which only the cell types that do not undergo a second cell division continue to express the transfected plasmids. This limits the application of IUE with standard plasmids and precludes its use in experiments where manipulating or labeling the complete cell lineage of a progenitor is desired. To circumvent this episomal loss of plasmid in IUE, we have used a binary piggyBac transposon system to induce nonviral genomic integration of transgenes. These transgenes do not appear to inactivate after cell division, and this results in stable somatic cellular transgenesis of neurons and glia. Like standard IUE, the system can be used with multiple combinations of plasmids to achieve multicolor labeling and both loss-of-function and gain-of-function manipulations. In this protocol, we describe the method for delivering a binary piggyBac transposon plasmid system by IUE.
Subcapsular Transplantation of Tissue in the Kidney
Authors: Shultz Leonard D. , Goodwin Neal , Ishikawa Fumihiko , Hosur Vishnu , Lyons Bonnie L. , Greiner Dale L.
Published: July 2014

Multiple sites can be used for the engraftment of primary human cells and tissues into murine hosts. For example, leukemias are usually best engrafted intravenously, but they can also be engrafted directly into the bone marrow cavity. Some solid tumors such as colon tumors grow successfully following subcutaneous engraftment, sometimes requiring provision of a Matrigel artificial basement membrane. In certain cases (e.g., human bladder cancer and ductal breast carcinoma), the use of the autochthonous site (bladder and mammary duct, respectively) is often most efficient, whereas the tumors can grow poorly when transplanted subcutaneously or heterochthonously. Here, we present a protocol for the surgical implantation of tissues under the kidney capsule. The kidney is especially suited for the transplantation of normal as well as malignant cells and tissues. It is very accessible, and transplanted tissues are well contained under the renal capsule in a highly vascularized site. Furthermore, the retroperitoneal location of the kidney, together with its separation from other organs, is advantageous both for imaging and biopsy.
Preparation of Cytosolic Extracts and Activation of Caspases by Cytochrome c
Authors: McStay Gavin P. , Green Douglas R.
Published: July 2014

It can be useful to explore the caspase activation process in an in vitro setting. In this protocol, cytosolic extracts prepared from cell culture are incubated with cytochrome c and adenosine triphosphate (dATP), leading to the oligomerization of apoptotic protease activating factor-1 (APAF-1) and the formation of the apoptosome. The apoptosome serves as an activation platform for caspase-9, which binds to the apoptosome through heterodimeric caspase recruitment domain (CARD) interactions and then dimerizes. This leads to cleavage of the executioner, caspase-3. These extracts contain highly active caspases that can be analyzed using a variety of biochemical assays.
Northern Blots for Small RNAs and MicroRNAs
Authors: Rio Donald C.
Published: July 2014

This protocol describes the detection of small RNAs (∼10–200 nucleotides) by blot hybridization. The RNA samples, denatured in formamide, are separated by denaturing polyacrylamide gel electrophoresis. Because high-percentage polyacrylamide gels are required to separate RNAs in this size range, it is necessary to perform electrophoretic transfer to positively charged nylon membranes. After transfer, the immobilized RNAs are subjected to hybridization with a 32P-radiolabeled DNA or RNA probe and detected by phosphorimaging or autoradiography. This procedure is commonly used to detect small, U-rich spliceosomal small nuclear RNAs (snRNAs) and miRNAs. It should be possible also to detect most miRNAs using high-percentage (e.g., 15%) urea–polyacrylamide gel electrophoresis.
Measuring the Ca2+-Binding Kinetics of Proteins
Authors: Faas Guido C. , Mody Istvan
Published: July 2014

Ca2+-binding proteins (CBPs) are instrumental in the control of Ca2+ signaling. For the transduction of a change in intracellular Ca2+ concentration into a cellular biochemical or biophysical action, it is necessary for Ca2+ to bind specific Ca2+-binding proteins (CBPs) that relay the Ca2+ signal. The competition for Ca2+ between the various CBPs plays an essential and direct role in this transduction and in the resulting biochemical message. Therefore, a thorough understanding of Ca2+ signaling necessitates appreciating the kinetic properties of all the relevant CBPs. Unfortunately, most conventional techniques used to measure Ca2+-binding kinetics are too slow to determine accurately the fast binding kinetics of most CBPs. To address this problem, we have developed an ultrafast in vitro technique for measuring the Ca2+-binding properties of CBPs following flash photolysis of caged Ca2+. We introduce here the protocols that are necessary for the data collection associated with this technique.
Two-Photon Imaging of Neural Activity in Awake Mobile Mice
Authors: Dombeck Daniel , Tank David
Published: July 2014

A method for functional, cellular-resolution imaging of neural populations in awake and mobile mice is presented here. The method is based on the use of a spherical treadmill, head restraint, and motion correction software that facilitate neural imaging in the awake brain with a fixed upright two-photon microscope. These approaches have proven to be applicable to a wide range of brain regions and should help further our understanding of how neuronal population activity within the brain’s microcircuitry is connected to animal behavior.
Crystal structure of 1,1-diacetylferrocene dihydrazone
Authors: Shikhaliyev Namig G. , Gurbanov Atash V. , Muzalevsky Vasily M. , Nenajdenko Valentine G. , Khrustalev Victor N.
Published: July 2014

The title compound, [Fe(C7H9N2)2], crystallizes with two crystallographically independent molecules in the unit cell. These represent the chiral atropoisomers distinguished by the mutual arrangement of the two acetyl–hydrazone groups with acisconformation of the C=N bonds. The two cyclopentadienyl (Cp) rings are planar and nearly parallel, the tilt between the two rings being 3.16 (16)° [4.40 (18)° for the second independent molecule]. The conformation of the Cp rings is close to eclipsed, the twist angle being 0.1 (2)° [3.3 (2)°]. The two acetyl–hydrazone substituents are also planar and are inclined at 13.99 (15)/9.17 (16)° [6.83 (17)/14.59 (15)°] relative to the Cp rings. The Fe—C bond lengths range from 2.035 (3) to 2.065 (2) Å, with an average of 2.050 (3) Å [2.036 (3) to 2.069 (2), average 2.046 (3) Å], which agrees well with those reported for most ferrocene derivatives. In the crystal, the molecules form dimersviatwo strong N—H...N hydrogen bonds. The dimers are linked into a three-dimensional framework by weak N—H...N hydrogen bonds.
Poly[[hexaaquasesqui(μ-benzene-1,2,4,5-tetracarboxylato)dicopper(II)disodium] monohydrate]
Authors: Camara Magatte , Keita Mohamed Fadel , Cisse Cherif Cheikh Samsidine , Daiguebonne Carole , Guillou Olivier
Published: July 2014

In the title compound, {[Cu2Na2(C10H2O8)1.5(H2O)6]·H2O}n, the Cu2+ion is hexacoordinated by five O atoms from benzene-1,2,4,5-tetracarboxylate (btec4−) ligands and one water molecule. The Na+ion is also hexacoordinated, by four O atoms from btec4−ligands and two water molecules. One of the two btec4−molecules sits on a crystallographic inversion centre. CuO6and NaO6octahedra are connected, forming bi-dimensional layers. These layers, which extend parallel to theacplane, are further interconnected by μ10- or μ11-bridging btec4−ligands and by O—H...O hydrogen bonds, involving both btec4−ligands and water molecules, forming a three-dimensional network.
7-Chloro-4-oxo-4H-chromene-3-carbaldehyde
Authors: Ishikawa Yoshinobu
Published: July 2014

In the title compound, C10H5ClO3, a chlorinated 3-formylchromone derivative, all atoms are essentially coplanar (r.m.s. deviation = 0.0592 Å for all non-H atoms), with the largest deviation from the least-squares plane [0.1792 (19) Å] being for the chromone-ring carbonyl O atom. In the crystal, molecules are linked through C—H...O hydrogen bonds to form tetrads, which are assembled by stacking interactions [centroid–centroid distance between the pyran rings = 3.823 (3) Å] and van der Waals contacts between the Cl atoms [Cl...Cl = 3.4483 (16) Å and C—Cl...Cl = 171.73 (7)°] into a three-dimensional architecture.
3β-Hydroxy-28-norolea-12,17-dien-11-one
Authors: Seebacher Werner , Weis Robert , Faist Johanna , Saf Robert , Belaj Ferdinand
Published: July 2014

The title compound, C29H44O2, was formed by treatment of 11-oxooleanolic acid under strong alkaline conditions. The absolute structure of the chiral molecules could not be determined reliably from the diffraction data, but is known from other triterpenes. The asymmetric unit consists of two molecules, 1 and 2. In both molecules, ringsAandBshow chair conformations. The other rings show mixed forms between envelope and half-chair conformations with atoms in positions 8, 15 and 21 forming the flaps in ringsC,DandE, respectively. RingsDandEof molecule 2 are disordered over two orientations, with occupancies of 0.557 (4) and 0.443 (4), which differ in the direction of the flap in ringE. In the crystal, molecules 1, as well as the molecules 2, are linked by O—H...O hydrogen bonds, forming chains parallel to thebaxis.
3-(2,4-Dichlorophenoxy)-1-(4-methoxybenzyl)-4-(4-nitrophenyl)azetidin-2-one
Authors: Atioğlu Zeliha , Akkurt Mehmet , Jarrahpour Aliasghar , Heiran Roghayeh , Özdemir Namık
Published: July 2014

The β-lactam ring of the title compound, C23H18Cl2N2O5, is nearly planar [maximum deviation = 0.019 (2) Å for the N atom] and its mean plane makes dihedral angles of 56.86 (15), 68.83 (15) and 83.75 (15)° with the dichloro-, nitro- and methoxy-substituted benzene rings, respectively. In the crystal, molecules are linked by pairs of C—H...O hydrogen bonds, forming inversion dimers withR22(10) loops. The dimers are linked by further C—H...O hydrogen bonds, forming sheets lying parallel to (001). The molecular packing is further stabilized by C—H...π interactions.
6-Fluoroindan-1-one
Authors: Slaw Benjamin R. , Tanski Joseph M.
Published: July 2014

The title compound, C9H7FO, crystallizes with two independent molecules in the asymmetric unit, in which corresponding bond lengths are the same within experimental error. The five-membered ring in each molecule is almost planar, with r.m.s. deviations of 0.016 and 0.029 Å. In the crystal, molecules form sheets parallel to (1 0 0)viaC—H...O and C—H...F interactions with F...F contacts [3.1788 (16) and 3.2490 (16) Å] between the sheets.