bluuebunny/crossref_metadata_2025 huggingface dataset:
Extent of Intraprotocol and Intersite Variability of Thoracic Magnetic Resonance Acquisition Times at a Large Quaternary Institution
Authors: Ackman Jeanne B. , Nitiwarangkul Chayanin , Mercaldo Sarah F.
Published: March 2019

Purpose:

The purpose of this study was to describe Thoracic magnetic resonance (MR) acquisition time (AT) variability, associations, and technologist insights as to its causes at a large quaternary institution, by MR protocol and imaging site.

Materials and Methods:

A retrospective review of our 2017 QI database of 1.5 T MR imaging ATs for adults 19 years and above at the main hospital and outpatient (OPT) satellites was performed for all 5 Thoracic MR protocols. Summary statistics were calculated for ATs. Multivariable linear regression was adjusted for age, sex, body mass index, time of examination relative to shift change, technologist experience, and language interpreter. An anonymous REDCap survey of our MR technologists sought their assessment of reasons for AT variability and techniques that help reduce it.

Results:

A total of 174 adult OPT 1.5 T mediastinal, pleural, and lung MR examinations were analyzed, revealing high variability of median AT by protocol and site (P<0.001)—for example, mean, median, slowest, and fastest ATs for Thymus I− protocol (n=38) were 34, 32, 66, and 8 minutes, respectively. OPT site with fewest MR technologists and a single MR scanner had shortest mean AT across all protocols (35±15 min). Full Chest I− protocol had shortest AT across all sites (mean AT=33±13 min), compared with focused imaging protocols. All I−/I+ protocols had greater than expected AT, compared with the same protocol performed (I−). Surveyed MR technologists noted limited Thoracic MR training/experience, discomfort with thoracic anatomy and Thoracic MR performance, and AT-related benefit of effective communication with the radiologist with regard to lesion location.

Conclusions:

There was tremendous intraprotocol and intersite variability of Thoracic MR ATs. Increased technologist training, amplified experience, and a solid understanding of lesion location for focused examinations may all help reduce Thoracic MR AT.
ntEdit: scalable genome assembly polishing
Authors: Warren René L , Coombe Lauren , Mohamadi Hamid , Zhang Jessica , Jaquish Barry , Isabel Nathalie , Jones Steven JM , Bousquet Jean , Bohlmann Joerg , Birol Inanç
Published: March 2019

In the modern genomics era, genome sequence assemblies are routine practice. However, depending on the methodology, resulting drafts may contain considerable base errors. Although utilities exist for genome base polishing, they work best with high read coverage and do not scale well. We developed ntEdit, a Bloom filter-based genome sequence editing utility that scales to large mammalian and conifer genomes.

We first tested ntEdit and the state-of-the-art assembly improvement tools GATK, Pilon and Racon on controlled E. coli and C. elegans sequence data. Generally, ntEdit performs well at low sequence depths (<20X), fixing the majority (>97%) of base substitutions and indels, and its performance is largely constant with increased coverage. In all experiments conducted using a single CPU, the ntEdit pipeline executed in <14s and <3m, on average, on E. coli and C. elegans, respectively. We performed similar benchmarks on a sub-20X coverage human genome sequence dataset, inspecting accuracy and resource usage in editing chromosomes 1 and 21, and whole genome. ntEdit scaled linearly, executing in 30-40m on those sequences. We show how ntEdit ran in <2h20m to improve upon long and linked read human genome assemblies of NA12878, using high coverage (54X) Illumina sequence data from the same individual, fixing frame shifts in coding sequences. We also generated 17-fold coverage spruce sequence data from haploid sequence sources (seed megagametophyte), and used it to edit our pseudo haploid assemblies of the 20 Gbp interior and white spruce genomes in <4 and <5h, respectively, making roughly 50M edits at a (substitution+indel) rate of 0.0024.

Availability

https://github.com/bcgsc/ntedit

Supplemental material

available online.
SVCurator: A Crowdsourcing app to visualize evidence of structural variants for the human genome
Authors: Chapman Lesley M , Spies Noah , Pai Patrick , Lim Chun Shen , Carroll Andrew , Narzisi Giuseppe , Watson Christopher M. , Proukakis Christos , Clarke Wayne E. , Nariai Naoki , Dawson Eric , Jones Garan , Blankenberg Daniel , Brueffer Christian , Xiao Chunlin , Kolora Sree Rohit Raj , Alexander Noah , Wolujewicz Paul , Ahmed Azza , Smith Graeme , Shehreen Saadlee , Wenger Aaron M. , Salit Marc , Zook Justin M.
Published: March 2019

A high quality benchmark for small variants encompassing 88 to 90% of the reference genome has been developed for seven Genome in a Bottle (GIAB) reference samples. However a reliable benchmark for large indels and structural variants (SVs) is yet to be defined. In this study, we manually curated 1235 SVs which can ultimately be used to evaluate SV callers or train machine learning models. We developed a crowdsourcing app – SVCurator – to help curators manually review large indels and SVs within the human genome, and report their genotype and size accuracy.

SVCurator is a Python Flask-based web platform that displays images from short, long, and linked read sequencing data from the GIAB Ashkenazi Jewish Trio son [NIST RM 8391/HG002], We asked curators to assign labels describing SV type (deletion or insertion), size accuracy, and genotype for 1235 putative insertions and deletions sampled from different size bins between 20 and 892,149 bp. The crowdsourced results were highly concordant with 37 out of the 61 curators having at least 78% concordance with a set of ‘expert’ curators, where there was 93% concordance amongst ‘expert’ curators. This produced high confidence labels for 935 events. When compared to the heuristic-based draft benchmark SV callset from GIAB, the SVCurator crowdsourced labels were 94.5% concordant with the benchmark set. We found that curators can successfully evaluate putative SVs when given evidence from multiple sequencing technologies.
The dot-compartment revealed? Diffusion MRI with ultra-strong gradients and spherical tensor encoding in the living human brain
Authors: Tax Chantal M.W. , Szczepankiewicz Filip , Nilsson Markus , Jones Derek K.
Published: March 2019

The so-called “dot-compartment” is conjectured in diffusion MRI to represent small spherical spaces, such as cell bodies, in which the diffusion is restricted in all directions. Previous investigations inferred its existence from data acquired with directional diffusion encoding which does not permit a straightforward separation of signals from ‘sticks’ (axons) and signals from ‘dots’. Here we combine isotropic diffusion encoding with ultra-strong diffusion gradients (240 mT/m) to achieve high diffusion-weightings with high signal to noise ratio, while suppressing signal arising from anisotropic water compartments with significant mobility along at least one axis (e.g., axons). A dot-compartment, defined to have apparent diffusion coefficient equal to zero and no exchange, would result in a non-decaying signal at very high b-values (b ≳ 7000 s/mm2). With this unique experimental setup, a residual yet slowly decaying, signal above the noise floor for b-values as high as 15000 s/mm2was seen clearly in the cerebellar grey matter (GM), and in several white matter (WM) regions to some extent. Upper limits of the dot-signal-fraction were estimated to be ~2% in cerebellar GM and ~0.2% in WM. By relaxing the assumption of zero diffusivity, the signal at high b-values in cerebellar GM could be represented more accurately by an isotropic water pool with a low apparent diffusivity of 0.11 µm2/ms and a substantial signal fraction of ~7-16%. This remaining signal at high b-values has potential to serve as a novel and simple marker for isotropically-restricted water compartments in cerebellar GM.
Herpesvirus infection reduces Pol II occupancy of host promoters but spares viral promoters
Authors: Hartenian Ella N , Glaunsinger Britt A
Published: March 2019

In mammalian cells, widespread acceleration of cytoplasmic mRNA degradation is linked to impaired RNA polymerase II (Pol II) transcription. This mRNA decay-induced transcriptional repression occurs during infection with gammaherpesviruses including Kaposi’s sarcoma-associated herpesvirus (KSHV) and murine gammaherpesvirus 68 (MHV68), which encode an mRNA endonuclease that initiates widespread RNA decay. Here, we show that MHV68-induced mRNA decay leads to a genome-wide reduction of Pol II occupancy at mammalian promoters. Viral genes, despite the fact that they require Pol II for transcription, escape this transcriptional repression. Protection is not governed by viral promoter sequences; instead, location on the viral genome is both necessary and sufficient to escape the transcriptional repression effects of mRNA decay. We hypothesize that the ability to escape from transcriptional repression is linked to the localization of viral DNA in replication compartments, providing a means for these viruses to counteract decay-induced viral transcript loss.
The PIWI/piRNA response is relaxed in a rodent that lacks mobilizing transposable elements
Authors: Vandewege Michael W. , Patt Roy N. , Merriman Dana K. , Ray David A. , Hoffmann Federico G.
Published: March 2019

Transposable elements (TEs) are genomic parasites that can propagate by inserting copies of themselves into host genomes. Mammalian genomes are typically dominated by LINE retrotransposons and their associated SINEs, and their mobilization in the germline is a challenge to genome integrity. There are genomic defenses against TE proliferation and the PIWI/piRNA defense is among the most well understood. However, the PIWI/piRNA system has been investigated largely in animals with abundant and actively mobilizing TEs and it is unclear how the PIWI/piRNA system functions in the absence of mobilizing TEs. The 13-lined ground squirrel provides an excellent opportunity to examine PIWI/piRNA and TE dynamics within the context of minimal, and possibly nonexistent, TE accumulation. We sequenced RNA and small RNAs pools from the testes of juvenile and adult squirrels and compared results to TE and PIWI/piRNA dynamics in the European rabbit and house mouse. Interestingly in squirrels, despite a lack of young insertions, TEs were still actively transcribed at higher levels compared to mouse and rabbit. All three PIWI proteins were either not expressed, or only minimally expressed, prior to P8 in squirrel testis, but there was little TE expression change with the onset of PIWI expression. We found PIWIs largely did not reduce TE transcription, and the ping-pong cycle was significantly reduced among squirrel LINEs and SINEs compared to the mouse and rabbit. We speculate that, although the PIWI/piRNA system is adaptable to novel TE threats, transcripts from TEs that are no longer threatening receive less attention from PIWI proteins.
Chloroplasts Require Glutathione Reductase to Balance Reactive Oxygen Species and Maintain Efficient Photosynthesis
Authors: Müller-Schüssele Stefanie J. , Wang Ren , Gütle Desirée D. , Romer Jill , Rodriguez-Franco Marta , Scholz Martin , Lüth Volker M. , Kopriva Stanislav , Dörmann Peter , Schwarzländer Markus , Reski Ralf , Hippler Michael , Meyer Andreas J.
Published: March 2019

Thiol-based redox-regulation is vital to coordinate chloroplast functions depending on illumination. Yet, how the redox-cascades of the thioredoxin and glutathione redox machineries integrate metabolic regulation and reactive oxygen species (ROS) detoxification remains largely unresolved. We investigate if maintaining a highly reducing stromal glutathione redox potential (EGSH) via glutathione reductase (GR) is necessary for functional photosynthesis and plant growth.

Since absence of the plastid/mitochondrial GR is embryo-lethal in Arabidopsis thaliana, we used the model moss Physcomitrella patens to create knock-out lines. We dissect the role of GR in chloroplasts by in vivo monitoring stromal EGSH dynamics, and reveal changes in protein abundances by metabolic labelling.

Whereas stromal EGSH is highly reducing in wildtype and clearly responsive to light, the absence of GR leads to a partial oxidation, which is not rescued by light. Photosynthetic performance and plant growth are decreased with increasing light intensities, while ascorbate and zeaxanthin levels are elevated. An adjustment of chloroplast proteostasis is pinpointed by the induction of plastid protein repair and degradation machineries.

Our results indicate that the plastid thioredoxin and glutathione redox systems operate largely independently. They reveal a critical role of GR in maintaining efficient photosynthesis.
Scientific evaluation of negative exome sequencing followed by systematic scoring of candidate genes to decipher the genetics of neurodevelopmental disorders
Authors: Büttner Benjamin , Martin Sonja , Finck Anja , Arelin Maria , Baade-Büttner Carolin , Bartolomaeus Tobias , Bauer Peter , Bertsche Astrid , Bernhard Matthias K. , Biskup Saskia , Donato Nataliya Di , Elgizouli Magdeldin , Ewald Roland , Heine Constanze , Hellenbroich Yorck , Hentschel Julia , Hoffjan Sabine , Horn Susanne , Hornemann Frauke , Huhle Dagmar , Kamphausen Susanne B. , Kiess Wieland , Krey Ilona , Kuechler Alma , Liesfeld Ben , Merkenschlager Andreas , Mitter Diana , Muschke Petra , Pfäffle Roland , Polster Tilman , Schanze Ina , Schlump Jan-Ulrich , Syrbe Steffen , Wieczorek Dagmar , Zenker Martin , Lemke Johannes R. , Duc Diana Le , Platzer Konrad , Jamra Rami Abou
Published: March 2019

Background

Deciphering the monogenetic causes of neurodevelopmental disorders (NDD) is an important milestone to offer personalized care. But the plausibility of reported candidate genes in exome studies often remains unclear, which slows down progress in the field.

Methods

We performed exome sequencing (ES) in 198 cases of NDD. Cases that remained unresolved (n=135) were re-investigated in a research setting. We established a candidate scoring system (CaSc) based on 12 different parameters reflecting variant and gene attributes as well as current literature to rank and prioritize candidate genes.

Results

In this cohort, we identified 158 candidate variants in 148 genes with CaSc ranging from 2 to 11.7. Only considering the top 15% of candidates, 14 genes were already published or funneled into promising validation studies.

Conclusions

We promote that in an approach of case by case re-evaluation of primarily negative ES, systematic and standardized scoring of candidate genes can and should be applied. This simple framework enables better comparison, prioritization, and communication of candidate genes within the scientific community. This would represent an enormous benefit if applied to the tens of thousands of negative ES performed in routine diagnostics worldwide and speed up deciphering the monogenetic causes of NDD.
The Human Exonuclease-1 Interactome And Phosphorylation Sites
Authors: Eid Wassim , Hess Daniel , König Christiane , Gentili Christian , Ferrari Stefano
Published: March 2019

Error-free repair of DNA double-strand break is orchestrated by homologous recombination (HR) pathways and requires the concerted action of several factors. Among these, EXO1 and DNA2/BLM execute extensive resection of DNA ends to produce 3’-overhangs, which are key intermediates for downstream steps of HR. To help shedding light on regulatory aspects of DNA repair pathways in which EXO1 participates, we set out to identify proteins interacting with EXO1. Affinity purification of EXO1 followed by Orbitrap mass spectrometry led to the identification of novel partners that are involved in RNA processing or that are the causative agents of rare X-linked disorders. Depletion of a selected subset of EXO1 interacting proteins led to reduction of the DNA damage response. Among those, we examined the RRP5-homologue and NFκB-interacting protein PDCD11/ALG-4, which has roles in apoptosis and is a putative driver gene in cutaneous T-cell lymphoma. We provide evidence that depletion of PDCD11 decreased the formation of γH2AX foci and the phosphorylation of DNA damage response signaling intermediates in response to camptothecin or bleomycin, resulting in increased cellular resistance to DNA damage. Furthermore, extensive coverage of EXO1 sequence (>85%) by mass spectrometry allowed conducting an in-depth analysis of its phosphorylation sites, with the identification of 26 residues that are differentially modified in untreated conditions or upon induction of DNA damage.

As a whole, these results provide the basis for future in-depth studies on novel roles of EXO1 in genome stability and indicate targets for pharmacological inhibition of pathways of cancer development.

HIGHLIGHTS

Proteome-wide analysis of Exonuclease-1 (EXO1) interacting proteins revealed novel partners involved in RNA processing or that are the causative agents of rare X-linked disorders.

We provide evidence for a role of PDCD11 in the DNA Damage Response.

We conducted a comprehensive identification of EXO1 phosphorylation sites.
A Polybasic Domain in aPKC Mediates Par6-Dependent Control of Membrane Targeting and Kinase Activity
Authors: Dong Wei , Lu Juan , Zhang Xuejing , Wu Yan , Lettieri Kaela , Hammond Gerald R. , Hong Yang
Published: March 2019

Mechanisms coupling the atypical PKC (aPKC) kinase activity to its subcellular localization are essential for cell polarization. Unlike other members of the PKC family, aPKC has no well-defined plasma membrane (PM) or calcium binding domains, leading to the assumption that its subcellular localization relies exclusively on protein-protein interactions. Here we show that in both Drosophila and mammalian cells the pseudosubstrate region (PSr) of aPKC acts as a polybasic domain capable of targeting aPKC to the PM via electrostatic binding to PM PI4P and PI(4,5)P2. However, physical interaction between aPKC and Par-6 is required for the PM-targeting of aPKC, likely by allosterically exposing the PSr to bind PM. Binding of Par-6 also inhibits aPKC kinase activity and such inhibition can be relieved through Par-6 interaction with apical polarity protein Crumbs. Our data suggest a potential mechanism in which allosteric regulation of polybasic PSr by Par-6 couples the control of both aPKC subcellular localization and spatial activation of its kinase activity.

eTOC Summary

Dong et al. discover that the pseudo-substrate region (PSr) in aPKC is a polybasic domain capable of electrostatically targeting aPKC to plasma membrane. Allosteric regulation of PSr by Par-6 couples the control of both aPKC subcellular localization and spatial activation of kinase activity.
An RNA guanine quadruplex regulated pathway to TRAIL-sensitization by DDX21
Authors: McRae Ewan K.S. , Dupas Steven J. , Booy Evan P. , Piragasam Ramanaguru S. , Fahlman Richard P. , McKenna Sean A.
Published: March 2019

DDX21 is a newly discovered RNA G-quadruplex (rG4) binding protein with no known biological rG4 targets. In this study we identified 26 proteins that are expressed at significantly different levels in cells expressing wild type DDX21 relative to an rG4 binding deficient DDX21 (M4). From this list we validate MAGED2 as a protein that is regulated by DDX21 through rG4 in its 5’UTR. MAGED2 protein levels, but not mRNA levels, are reduced by half in cells expressing only DDX21 M4. MAGED2 has a repressive effect on TRAIL-R2 expression that is relieved under these conditions, resulting in elevated TRAIL-R2 mRNA and protein in cells expressing only DDX21 M4, and rendering previously resistant cells sensitive to TRAIL mediated apoptosis. Our work identifies the role of DDX21 in regulation at the translational level through biologically relevant rG4 and shows that MAGED2 protein levels are regulated, at least in part, by a rG4 forming potential in their 5’UTRs.
Ethanolamine regulates CqsR quorum-sensing signaling inVibrio cholerae
Authors: Watve Samit , Barrasso Kelsey , Jung Sarah A. , Davis Kristen J. , Hawver Lisa A. , Khataokar Atul , Palaganas Ryan G. , Neiditch Matthew B. , Perez Lark J. , Ng Wai-Leung
Published: March 2019

The pathogen that causes cholera,Vibrio cholerae, uses the cell-cell communication process known as quorum sensing (QS) to regulate virulence factor production and biofilm formation in response to changes in population density and complexity. QS is mediated through the detection of extracellular chemical signals called autoinducers. Four histidine kinases, LuxPQ, CqsS, CqsR and VpsS, have been identified as receptors to activate the key QS regulator LuxO at low cell density. At high cell density, detection of autoinducers by these receptors leads to deactivation of LuxO, resulting in population-wide gene expression changes. While the cognate autoinducers that regulate the activity of CqsS and LuxQ are known, the signals that regulate CqsR have not been determined. Here we show that the common metabolite ethanolamine specifically interacts with the ligand-binding CACHE domain of CqsRin vitroand induces the high cell-density QS response through CqsR kinase inhibition inV. choleraecells. We also identified residues in the CqsR CACHE domain important for ethanolamine detection and signal transduction. Moreover, mutations disrupting endogenous ethanolamine production inV. choleraedelay the onset of, but do not abolish, the high cell-density QS gene expression. Finally, we demonstrate that modulation of CqsR QS response by ethanolamine occurs inside animal hosts. Our findings suggest thatV. choleraeuses CqsR as a dual-function receptor to integrate information from the self-made signals as well as exogenous ethanolamine as an environmental cue to modulate QS response.

IMPORTANCE

Many bacteria use quorum sensing to regulate cellular processes that are important for their survival and adaptation to different environments. Quorum sensing usually depends on the detection on chemical signals called autoinducers made endogenously by the bacteria. We show here ethanolamine, a common metabolite made by various bacteria and eukaryotes, can modulate the activity of one of the quorum-sensing receptors inVibrio cholerae, the etiological agent of the disease cholera. Our results raise the possibility thatV. choleraeor other quorum-sensing bacteria can combine environmental sensing and quorum sensing to control group behaviors.
Both galactosaminogalactan and α-1,3-glucan contribute to aggregation ofAspergillus oryzaehyphae in liquid culture
Authors: Miyazawa Ken , Yoshimi Akira , Sano Motoaki , Tabata Fuka , Sugahara Asumi , Kasahara Shin , Koizumi Ami , Yano Shigekazu , Nakajima Tasuku , Abe Keietsu
Published: March 2019

Filamentous fungi generally form aggregated hyphal pellets in liquid culture. We previously reported that α-1,3-glucan-deficient mutants ofAspergillus nidulansdid not form hyphal pellets and their hyphae were fully dispersed, and we suggested that α-1,3-glucan functions in hyphal aggregation. Yet,Aspergillus oryzaeα-1,3-glucan-deficient (AGΔ) mutants still form small pellets; therefore, we hypothesized that another factor responsible for forming hyphal pellets remains in these mutants. Here, we identified an extracellular matrix polysaccharide galactosaminogalactan (GAG) as such a factor. To produce a double mutant ofA. oryzae(AG-GAGΔ), we disrupted the genes required for GAG biosynthesis in an AGΔ mutant. Hyphae of the double mutant were fully dispersed in liquid culture, suggesting that GAG is involved in hyphal aggregation inA. oryzae. Addition of partially purified GAG fraction to the hyphae of the AG-GAGΔ strain resulted in formation of mycelial pellets. Acetylation of the amino group in galactosamine of GAG weakened GAG aggregation, suggesting that hydrogen bond formation by this group is important for aggregation. Genome sequences suggest that α-1,3-glucan, GAG, or both are present in many filamentous fungi and thus may function in hyphal aggregation in these fungi. We also demonstrated that production of a recombinant polyesterase, CutL1, was higher in the AG-GAGΔ strain than in the wild-type and AGΔ strains. Thus, controlling hyphal aggregation factors of filamentous fungi may increase productivity in the fermentation industry.

Importance

Production using filamentous fungi is an important part of the fermentation industry, but hyphal aggregation in these fungi in liquid culture limits productivity compared with that of yeast or bacterial cells. We found that galactosaminogalactan and α-1,3-glucan both function in hyphal aggregation inAspergillus oryzae, and that the hyphae of a double mutant deficient in both polysaccharides become fully dispersed in liquid culture. We also revealed the relative contribution of α-1,3-glucan and galactosaminogalactan to hyphal aggregation. Recombinant protein production was higher in the double mutant than in the wild-type strain. Our research provides a potential technical innovation for the fermentation industry that uses filamentous fungi, as regulation of the growth characteristics ofA.oryzaein liquid culture may increase productivity.
Shh induces symmetry breaking in the presomitic mesoderm by inducing tissue shear and orientated cell rearrangements
Authors: Yin J. , Saunders T. E.
Published: March 2019

Future boundaries of skeletal muscle segments are determined in the presomitic mesoderm (PSM). Within the PSM, future somitic cells undergo significant changes in both morphology and position. How such large-scale cellular changes are coordinated and the effect on the future border formation is unknown. We find that cellular rearrangements differ between cell populations within the PSM. In contrast to lateral somitic cells, which display less organized rearrangement, the adaxial cell layer undergoes significant tissue shearing with dorsal and ventral cells sliding posteriorly. This shear is generated by orientated intercalations of dorsally and ventrally located adaxial cells, which induces a chevron-like pattern. We find Shh signaling is required for the tissue shear and morphogenesis of adaxial cells. In particular, we observe Shh-dependent polarized recruitment of non-muscle myosin IIA drives apical constrictions, and thus the intercalations and shear. This reveals a novel role for Shh in regulating cell mechanics in the PSM.
Harmonious genetic combinations rewire regulatory networks and flip gene essentiality
Authors: New Aaron M. , Lehner Ben
Published: March 2019

We lack an understanding of how the full range of genetic variants that occur in individuals can interact. To address this shortcoming, here we combine diverse mutations between genes in a model regulatory network, the galactose (GAL) switch of budding yeast. The effects of thousands of pairs of mutations fall into a limited number of phenotypic classes. While these effects are mostly predictable using simple rules that capture the ‘stereotypical’ genetic interactions of the network, some double mutants have unexpected outcomes including constituting alternative functional switches. Each of these ‘harmonious’ genetic combinations exhibits altered dependency on other regulatory genes. These cases illustrate how both pairwise and higher epistasis determines gene essentiality and how combinations of mutations rewire regulatory networks. Together, our results provide an overview of how broad spectra of mutations interact, how these interactions can be predicted, and how diverse genetic solutions can achieve ‘wild-type’ phenotypic behavior.
Efficient and specific oligo-based depletion of rRNA
Authors: Kraus Amelie J. , Brink Benedikt G. , Siegel T. Nicolai
Published: March 2019

In most organisms, ribosomal RNA (rRNA) contributes to >85% of total RNA. Thus, to obtain useful information from RNA-sequencing (RNA-seq) analyses at reasonable sequencing depth, typically, mature polyadenylated transcripts are enriched or rRNA molecules are depleted. Targeted depletion of rRNA or other highly abundant transcripts is particularly useful when studying transcripts lacking a poly(A) tail, such as some non-coding RNAs (ncRNAs), most bacterial RNAs and partially degraded or immature transcripts. While several commercially available kits allow effective rRNA depletion, their efficiency relies on a high degree of sequence homology between oligonucleotide probes and the target RNA. This restricts the use of such kits to a limited number of organisms with conserved rRNA sequences.

In this study we describe the use of biotinylated oligos and streptavidin-coated paramagnetic beads for the efficient and specific depletion of trypanosomal rRNA. Our approach reduces the levels of the most abundant rRNA transcripts to less than 5% with minimal off-target effects.

By adjusting the sequence of the oligonucleotide probes, our approach can be used to deplete rRNAs or other abundant transcripts independent of species. Thus, our protocol provides a useful alternative for rRNA removal where enrichment of polyadenylated transcripts is not an option and commercial kits for rRNA are not available.
The FH2 domain of formin proteins is critical for platelet cytoskeletal dynamics
Authors: Green Hannah L.H. , Zuidscherwoude Malou , Thomas Steven G.
Published: March 2019

Reorganisation of the actin cytoskeleton is required for proper functioning of platelets following activation in response to vascular damage. Formins are a family of proteins which regulate actin polymerisation and cytoskeletal organisation. Several formin protein are expressed in platelets and so we used an inhibitor of formin mediated actin polymerisation (SMIFH2) to uncover the role of these proteins in platelet spreading. Pre-treatment with SMIFH2 completely blocks platelet spreading in both mouse and human platelets through effects on the organisation and dynamics of actin and microtubules. However, platelet aggregation and secretion are unaffected. SMIFH2 also caused a decrease in resting platelet size and disrupted the balance of tubulin post-translational modification. These data therefore demonstrated an important role for formin mediated actin polymerisation in platelet spreading and highlighted their importance in cross talk between the actin and tubulin cytoskeletons.

Key Points

Inhibition of FH2 domains blocks platelet spreading and disrupts actin and microtubule organisation

Inhibition of FH2 domains causes a reduction in resting platelet size but not by microtubule coil depolymerisation

FH2 domains play a role in the post-translational modification of microtubules

Visual abstract
Stoichiometric constraints modulate the effects of temperature and nutrients on biomass distribution and community stability
Authors: Sentis Arnaud , Haegeman Bart , Montoya José M.
Published: March 2019

Abstract

Temperature and nutrients are two of the most important drivers of global change. Both can modify the elemental composition (i.e. stoichiometry) of primary producers and consumers. Yet their combined effect on the stoichiometry, dynamics, and stability of ecological communities remains largely unexplored. To fill this gap, we extended the Rosenzweig-MacArthur consumer-resource model by including thermal dependencies, nutrient dynamics, and stoichiometric constraints on both the primary producer and the consumer. We found that stoichiometric constraints dampen the paradox of enrichment and increased persistence at high nutrient levels. Nevertheless, they also reduced consumer persistence at extreme temperatures. Finally, we also found that stoichiometric constraints can strongly influence biomass distribution across trophic levels by modulating consumer assimilation efficiency and resource growth rates along the environmental gradients. In the Rosenzweig-MacArthur model, consumer biomass exceeded resource biomass for most parameter values whereas, in the stoichiometric model, consumer biomass was strongly reduced and sometimes lower than resource biomass. Our findings highlight the importance of accounting for stoichiometric constraints as they can mediate the temperature and nutrient impact on the dynamics and functioning of ecological communities.
Cell shape and antibiotic resistance is maintained by the activity of multiple FtsW and RodA enzymes inListeria monocytogenes
Authors: Rismondo Jeanine , Halbedel Sven , Gründling Angelika
Published: March 2019

Rod-shaped bacteria have two modes of peptidoglycan synthesis: lateral synthesis and synthesis at the cell division site. These two processes are controlled by two macromolecular protein complexes, the elongasome and divisome. Recently, it has been shown that theBacillus subtilisRodA protein, which forms part of the elongasome, has peptidoglycan glycosyltransferase activity. The cell division specific RodA homolog FtsW fulfils a similar role at the divisome. The human pathogenListeria monocytogenesencodes up to six FtsW/RodA homologs, however their functions have not yet been investigated. Analysis of deletion and depletion strains led to the identification of the essential cell division-specific FtsW protein, FtsW1. Interestingly,L. monocytogenesencodes a second FtsW protein, FtsW2, which can compensate for the lack of FtsW1, when expressed from an inducible promoter.L. monocytogenesalso possesses three RodA homologs, RodA1, RodA2 and RodA3 and their combined absence is lethal. Cells of arodA1/rodA3double mutant are shorter and have increased antibiotic and lysozyme sensitivity, probably due to a weakened cell wall. Results from promoter activity assays revealed that expression ofrodA3andftsW2is induced in the presence of antibiotics targeting penicillin binding proteins. Consistent with this, arodA3mutant was more susceptible to the β-lactam antibiotic cefuroxime. Interestingly, overexpression of RodA3 also led to increased cefuroxime sensitivity. Our study highlights thatL. monocytogenesencodes a multitude of functional FtsW and RodA enzymes to produce its rigid cell wall and that their expression needs to be tightly regulated to maintain growth, cell division and antibiotic resistance.

Importance

The human pathogenListeria monocytogenesis usually treated with high doses of β-lactam antibiotics, often combined with gentamicin. However, these antibiotics only act bacteriostatically onL. monocytogenesand the immune system is needed to clear the infection. Therefore, individuals with a compromised immune system are at risk to develop a severe form ofListeriainfection, which can be fatal in up to 30% of cases. The development of new strategies to treatListeriainfections is therefore necessary. Here we show that the expression of some of the FtsW and RodA enzymes ofL. monocytogenesis induced by the presence of β-lactam antibiotics and their combined absence makes bacteria more susceptible to this class of antibiotics. The development of antimicrobials that inhibit the activity or production of FtsW/RodA enzymes might therefore help to improve the treatment ofListeriainfections and thereby lead to a reduction in mortality.
Characterization and application of Bragg-edge transmission imaging for strain measurement and crystallographic analysis on the IMAT beamline
Authors: Ramadhan Ranggi S. , Kockelmann Winfried , Minniti Triestino , Chen Bo , Parfitt David , Fitzpatrick Michael E. , Tremsin Anton S.
Published: March 2019

This paper presents a series of experiments to characterize the performance of the new IMAT beamline at the ISIS pulsed neutron source and provides examples to showcase the potential applications of Bragg-edge transmission imaging on the instrument. The characterization includes determination of the IMAT spectral and spatial resolutions through calibration measurements, and also determination of the precision and the accuracy of Bragg-edge analysis for lattice parameters of ceramics, metals and textured engineering alloys through high-temperature measurements. A novel Bragg-edge analysis method based on the cross-correlation of different Bragg edges has been developed to provide an estimate of the change in lattice parameter, which is especially useful for measurements of textured samples. Three different applications of the Bragg-edge transmission imaging technique are presented, including strain mapping, texture mapping and obtaining crystallographic information, i.e. the dependence on temperature of the Debye–Waller factor. The experimental results demonstrate the ability of the IMAT beamline to provide accurate strain measurements with uncertainties as low as 90 µɛ with reasonable measurement time, while characteristic materials parameters can be mapped across the sample with a spatial resolution of 300–600 µm for a strain map and down to ∼90 µm for a texture map.
A 1:1:1 co-crystal solvate comprising 2,2′-dithiodibenzoic acid, 2-chlorobenzoic acid and N,N-dimethylformamide: crystal structure, Hirshfeld surface analysis and computational study
Authors: Tan Sang Loon , Tiekink Edward R. T.
Published: March 2019

The asymmetric unit of the three-component title compound, 2,2′-dithiodibenzoic acid–2-chlorobenzoic acid–N,N-dimethylformamide (1/1/1), C14H10O4S2·C7H5ClO2·C3H7NO, contains a molecule each of 2,2′-dithiodibenzoic acid (DTBA), 2-chlorobenzoic acid (2CBA) and dimethylformamide (DMF). The DTBA molecule is twisted [the C—S—S—C torsion angle is 88.37 (17)°] and each carboxylic group is slightly twisted from the benzene ring to which it is connected [CO2/C6 dihedral angles = 7.6 (3) and 12.5 (3)°]. A small twist is evident in the molecule of 2CBA [CO2/C6 dihedral angle = 4.4 (4)°]. In the crystal, the three molecules are connected by hydrogen bonds with the two carboxylic acid residues derived from DTBA and 2CBA forming a non-symmetric eight-membered {...HOCO}2 synthon, and the second carboxylic acid of DTBA linked to the DMF molecule via a seven-membered {...HOCO...HCO} heterosynthon. The three-molecule aggregates are connected into a supramolecular chain along the a axis via DTBA-C—H...O(hydroxyl-2CBA), 2CBA-C—H...O(hydroxyl-DTBA) and DTBA-C—H...S(DTBA) interactions. Supramolecular layers in the ab plane are formed as the chains are linked via DMF-C—H...S(DTBA) contacts, and these inter-digitate along the c-axis direction without specific points of contact between them. A Hirshfeld surface analysis points to additional but, weak contacts to stabilize the three-dimensional architecture: DTBA-C=O...H(phenyl-DTBA), 2CBA-Cl...H(phenyl-DTBA), as well as a π–π contact between the delocalized eight-membered {...HOC=O}2 carboxylic dimer and the phenyl ring of 2CBA. The latter was confirmed by electrostatic potential (ESP) mapping.
(E,E)-3-Methyl-2,5-bis(4-methylbenzylidene)cyclopentanone: synthesis, characterization, Hirshfeld surface analysis and antibacterial activity
Authors: Mahdi Fatiha , Sid Assia , Bouchene Rafika , Mosset Paul , Roisnel Thierry
Published: March 2019

The title compound, (E,E)-3-methyl-2,5-bis(4-methylbenzylidene)cyclopentanone (MBMCP), C22H22O, was obtained by Claisen–Schmidt condensation of 4-methylbenzaldehyde with 3-methylcyclopentanone in good yield. The structure of MBMCP was studied using UV, FT–IR and Raman spectroscopy, single-crystal X-ray diffraction (XRD) measurements, and 1H and 13C nuclear magnetic resonance (NMR) spectroscopy. The molecular structure of MBMCP is fully extended in the E,E configuration. C—H...π stacking interactions play a significant role in the stabilization of the molecular packing. Hirshfeld surface analysis was used to quantify the non-covalent interactions in the crystal lattice. Microbiological studies were performed to investigate the antimicrobial activity of this new product.
Quantitative lean assessment of line of balance schedules’ quality
Authors: Ungureanu Lucian Constantin , Hartmann Timo , Serbanoiu Ion
Published: March 2019

Purpose

The line of balance (LOB) method is a suitable scheduling technique that managers can use to support lean planning efforts for projects composed of repetitive activities such as high-rise building construction. Like any other method, LOB has certain disadvantages that create a set of practical limitations in its application. An LOB schedule gives insights about how continuous and synchronized single resources are scheduled and how uniform these resources are distributed over the project duration. However, these three characteristics have to be visually checked, which makes the evaluation and the comparison of different schedule alternatives difficult. To overcome this problem, the purpose of this paper is to present a quantitative method to calculate quality degrees for the continuity, the synchronization and the uniformity of an LOB schedule that can be applied to assess an LOB schedule globally.

Design/methodology/approach

The paper introduces a set of global indicators, termed quality degrees, which allow for a quick quantitative evaluation of LOB schedules at the global level. These quality degrees are quantitative indicators for the: degree of continuity, degree of synchronization and degree of uniformity within a specific LOB alternative. A mathematical model was developed to calculate the quality degrees for LOB schedules. This model was validated using a well-known case study extracted from literature, and its practical implementation was exemplified on two real Romanian projects.

Findings

The paper illustrates this contribution using two case studies that confirm that the proposed method can be used to evaluate different schedule alternatives. In particular, the paper shows that quality indicators can be used to analyze and control interdependencies between cost and time.

Research limitations/implications

The main limitation of the proposed method is that it cannot indicate the desired level of continuity, synchronization or uniformity to be achieved. Further studies need to explore this possibility, as well the relationship between indicators.

Practical implications

The presented quality indicators contribute to existing LOB methods as they allow for the quick analysis and assessment of schedules without an in-depth visual analysis.

Originality/value

The paper proposes an innovative method, mathematically formulated, to quantitatively assess the quality aspects of continuity, synchronization and uniformity for LOB schedules.
Work integrated learning internships in transnational education
Authors: Bilsland Christine , Carter Leanne , Wood Leigh N.
Published: March 2019

Purpose

Research into employability initiatives such as work integrated learning (WIL) in transnational education (TNE) is scarce, and the alumni voice in TNE is largely unreported. The purpose of this paper is to contribute to TNE research by investigating the value of internship electives in the TNE campus location.

Design/methodology/approach

A grounded theory approach employed semi-structured interviews with local business undergraduate alumni in Vietnam.

Findings

Internships were instrumental to local graduate employment transitions. University support of WIL internships was a valuable differentiator in the Vietnamese university context, where internships lack formal support mechanisms. Alumni regarded internships as transformational learning journeys, rather than simply as pathways to post-graduate jobs.

Research limitations/implications

The scope of the study was Vietnam. Findings imply the importance of incorporating local stakeholder perspectives into TNE, particularly regarding WIL.

Practical implications

Universities that operate in transnational environments must meet local stakeholder needs by providing authentic, industry-related learning activities. The findings support the integration of WIL internships into TNE programmes in Vietnam and further research relevant to other TNE contexts.

Originality/value

The study contributes to underdeveloped TNE research around employability in general, and more specifically about the particular value of internships in TNE campus locations. Alumni stakeholders constitute uniquely valuable feedback sources based on their shared experience as TNE students, interns and graduate employees in local work environments. Their insights enable universities to facilitate locally relevant learning outcomes.
Project manager’s role in safety performance of Saudi construction
Authors: Khawam Abdullah A. , Bostain Nancy S.
Published: March 2019

Purpose

The purpose of this paper is to address the primary research question, which is what is the relationship between the project manager’s (PM) possession of the Project Management Professional (PMP) certification and the level of the safety culture present in the construction project the PM manages.

Design/methodology/approach

The research was based on a survey of a purposive sample, 109 engineers and first-line supervisors worked in 23 construction projects of which ten were led by PMP-certified PMs and 13 were led by PMs lacking PMP certification. Each PM completed a demographic questionnaire for the predictor variables of PMP certification controlled for age and experience. To assess the criterion variable of safety culture total score, engineers and first-line supervisors working in the same project completed the questionnaire of safety culture values and practices.

Findings

Results of this study indicated the level of safety culture was significantly different, and improved, for engineers and first-line supervisors who work under PMs with PMP certification compared to the level of safety culture in projects managed by PMs with no PMP certification. Although alignment of safety culture perceptions among different levels in the organization helps to achieve a positive safety culture, the role of the PM in transferring, implementing and maintaining the safety culture in the construction project is fundamental, particularly in small-to-medium sized enterprises (SMEs).

Originality/value

This study addressed the role of the PMs managerial skills in the safety performance of Saudi Arabian SMEs. The principal finding was that PMs with managerial skills perform better regarding safety performance in SME construction projects than PMs lacking managerial skills. The primary recommendation is that leaders in construction projects must carefully evaluate engineers’ managerial skills before hiring the individuals as PMs. A PM’s promotion model developed in this study provides a suitable framework and business process component for construction leaders seeking to maintain safety performance successfully.
Segmenting elder German grocery shoppers based on shopping motivations
Authors: Oeser Gerald , Aygün Tanju , Balan Claudia-Livia , Paffrath Rainer , Schuckel Marcus Thomas
Published: March 2019

Purpose

Elder German grocery shoppers are a growing, heterogeneous, and highly relevant and attractive, but under-researched market segment. In order to understand them and their grocery shopping motivations better and target them efficiently and effectively, the purpose of this paper is to identify dimensions of their shopping motivations and segment them based on these dimensions.

Design/methodology/approach

In total, 26 grocery store-choice criteria were identified in a thorough literature review and focus-group interviews with 36 elder German consumers aged 65 and older. In a subsequent survey, the importance of these criteria was rated by 1,288 German shoppers of the same age group. A principal component and cluster analysis were performed to identify dimensions of store-choice criteria and segments of elder German grocery shoppers. Multivariate analysis of variance, analysis of variance and discriminant analysis were used to test for statistically significant differences between the clusters.

Findings

Basic quality considerations, shopping experience and social interaction, service and assistance, price consciousness, product orientation, convenient location and quick service and packaging requirements influence the grocery store choice of elder German consumers in decreasing order of variance explained. The cluster analysis revealed indifferent, leisure, convenience, assistance-oriented, no frills, product-oriented and service-oriented elder German shoppers, which differ in their shopping motivations statistically and significantly. These clusters are described and contrasted in detail to derive managerial implications.

Originality/value

This research provides the first store-choice component analysis and cluster analysis for elder German grocery shoppers. This can help food retail to reach this attractive target group more efficiently and effectively and improve the food supply of elder German consumers.