bluuebunny/crossref_metadata_2025 huggingface dataset:
Mutational impact of culturing human pluripotent and adult stem cells
Authors: Kuijk Ewart , Jager Myrthe , Roest Bastiaan van der , Locati Mauro , Hoeck Arne Van , Korzelius Jerome , Janssen Roel , Besselink Nicolle , Boymans Sander , Boxtel Ruben van , Cuppen Edwin
Published: September 2018

Genetic changes acquired during in vitro culture pose a potential risk for the successful application of stem cells in regenerative medicine. To assess mutation accumulation risks induced by culturing, we determined genetic aberrations in individual human induced pluripotent stem cells (iPS cells) and adult stem cells (ASCs) by whole genome sequencing analyses. Individual iPS cells, intestinal ASCs and liver ASCs accumulated 3.5±0.5, 7.2±1.0 and 8.4±3.6 base substitutions per population doubling, respectively. The annual in vitro mutation accumulation rate of ASCs adds up to ∼1600 base pair substitutions, which is ∼40-fold higher than the in vivo rate of ∼40 base pair substitutions per year. Mutational analysis revealed a distinct in vitro induced mutational signature that is irrespective of stem cell type and distinct from the in vivo mutational signature. This in vitro signature is characterized by C to A changes that have previously been linked to oxidative stress conditions. Additionally, we observed stem cell-specific mutational signatures and differences in transcriptional strand bias, indicating differential activity of DNA repair mechanisms between stem cell types in culture. We demonstrate that the empirically defined mutation rates, spectra, and genomic distribution enable risk assessment by modelling the accumulation of specific oncogenic mutations during typical in vitro expansion, manipulation or screening experiments using human stem cells. Taken together, we have here for the first time accurately quantified and characterized in vitro mutation accumulation in human iPS cells and ASCs in a direct comparison. These results provide insights for further optimization of culture conditions for safe in vivo utilization of these cell types for regenerative purposes.
Fine-scale recombination landscapes between a freshwater and marine population of threespine stickleback fish
Authors: Shanfelter Alice F. , Archambeault Sophie L. , White Michael A.
Published: September 2018

Meiotic recombination is a highly conserved process that has profound effects on genome evolution. Recombination rates can vary drastically at a fine-scale across genomes and often localize to small recombination “hotspots” with highly elevated rates surrounded by regions with little recombination. Hotspot targeting to specific genomic locations is variable across species. In some mammals, hotspots have divergent landscapes between closely related species which is directed by the binding of the rapidly evolving protein, PRDM9. In many species outside of mammals, hotspots are generally conserved and tend to localize to regions with open chromatin such as transcription start sites. It remains unclear if the location of recombination hotspots diverge in taxa outside of mammals. Threespine stickleback fish (Gasterosteus aculeatus) are an excellent model to examine the evolution of recombination over short evolutionary timescales. Using an LD-based approach, we found recombination rates varied at a fine-scale across the genome, with many regions organized into narrow hotspots. Hotspots had divergent landscapes between stickleback populations, where only ~15% were shared, though part of this divergence could be due to demographic history. Additionally, we did not detect a strong association of PRDM9 with recombination hotspots in threespine stickleback fish. Our results suggest fine-scale recombination rates may be diverging between closely related populations of threespine stickleback fish and argue for additional molecular characterization to verify the extent of the divergence.
Escalation and constraints of antagonistic armaments in water striders
Authors: Crumière Antonin Jean Johan , Armisén David , Vargas-Lowman Aïdamalia , Kubarakos Martha , Moreira Felipe Ferraz Figueiredo , Khila Abderrahman
Published: September 2018

Sexual conflict may result in the escalating coevolution of sexually antagonistic traits. However, our understanding of the evolutionary dynamics of antagonistic traits and their role in association with sex-specific escalation remains limited. Here we study sexually antagonistic coevolution in a genus of water striders calledRhagovelia. We identified a set of male grasping traits and female anti-grasping traits used during pre-mating struggles and show that natural variation of these traits is associated with variation in mating performance in the direction expected for antagonistic co-evolution. Phylogenetic mapping detected signals of escalation of these sexually antagonistic traits suggesting an ongoing arms race. Moreover, their escalation appears to be constrained by a trade-off with dispersal through flight in both sexes. Altogether our results highlight how sexual interactions may have shaped sex-specific antagonistic traits and how constraints imposed by natural selection may have influenced their evolution.
Lovastatin, not simvastatin, corrects core phenotypes in the fragile X mouse model
Authors: Muscas Melania , Louros Susana R. , Osterweil Emily K.
Published: September 2018

The cholesterol-lowering drug lovastatin corrects neurological phenotypes in animal models of fragile X syndrome (FX), a commonly identified genetic cause of autism and intellectual disability. The therapeutic efficacy of lovastatin is being tested in clinical trials for FX, however the structurally similar drug simvastatin has been proposed as an alternative due to an increased potency and brain penetrance. Here, we perform a side-by-side comparison of the effects of lovastatin and simvastatin treatment on two core phenotypes in the Fmr1-/y mouse model. We find that while lovastatin normalizes excessive hippocampal protein synthesis and reduces audiogenic seizures (AGS) in the Fmr1-/y mouse, simvastatin does not correct either phenotype. These results caution against the assumption that simvastatin is a valid alternative to lovastatin for the treatment of FX.
Stage-specific transcriptomes and DNA methylomes indicate an early and transient loss of transposon control in Arabidopsis shoot stem cells
Authors: Gutzat Ruben , Rembart Klaus , Nussbaumer Thomas , Pisupati Rahul , Hofmann Falko , Bradamante Gabriele , Daubel Nina , Gaidora Angelika , Lettner Nicole , Donà Mattia , Nordborg Magnus , Nodine Michael , Scheid Ortrun Mittelsten
Published: September 2018

In contrast to animals, postembryonic development in plants is modular, and aerial organs originate from stem cells in the center of the shoot apical meristem (SAM) throughout life. Descendants of SAM stem cells in the subepidermal layer (L2) give also rise to male and female gametes (reviewed in 1) and are therefore considered primordial germ cells. In these cells, transmission of somatic mutations including virus and TE insertions must be avoided. Despite their essential role for plant development and intergenerational continuity, no comprehensive molecular analysis of SAM stem cells exists, due to their low number, deep embedding among non-stem cells, and difficult isolation. Here we present a comprehensive analysis of stage-specific gene expression and DNA methylation dynamics in Arabidopsis SAM stem cells. Stem cell expression signatures are mostly defined by development, but we also identified a core set of differentially expressed stemness genes. Surprisingly, vegetative SAM stem cells showed increased expression of transposable elements (TEs) relative to surrounding cells, despite high expression of genes connected to epigenetic silencing. We also find increasing methylation at CHG and a drop in CHH methylation at TEs before stem cells enter the reproductive lineage, indicating an onset of epigenetic reprogramming at an early stage. Transiently elevated TE expression is reminiscent of that in animal primordial germ cells (PGCs) 2 and demonstrates commonality of transposon biology. Our results connect SAM stem cells with germline development and transposon evolution and will allow future experiments to determine the degree of epigenetic heritability between generations.
Plants maximize competition while minimizing competitors belowground: a theoretical analysis of incentives for root competition in space
Authors: Farrior Caroline E.
Published: September 2018

Recent research shows that shared access to belowground resources drives plants to overproliferate fine roots competitively, limiting community-level aboveground biomass. Models of this phenomenon are commonly based on an assumption that belowground resources and fine roots are thoroughly well mixed. In reality, of course, fine roots are spatially structured by individual. Here we investigate how costs of sending roots through horizontal space influence incentives for fine-root overproliferation. We find that these costs restrain overproliferation to the net benefit of community aboveground biomass. And further, the costs eliminate incentives for individuals to grow fine roots beyond their closest neighbors. Plants that interact with the fewest competitors benefit the most in relative fitness from overproliferation of fine roots. Effectively, individual-based optimization of root allocation in space increases the effects of competition while decreasing the number of individual competitors for each individual.

Because an individual’s optimal competitive network consists of only the closest neighbors, we predict the full effects of competition are achieved just shortly after disturbance, making competition belowground an almost inescapable pressure on plants. Together these results have important implications for predicting plant interaction networks, patterns of carbon allocation, and ecosystem carbon storage.
Panton-Valentine leucocidin is the key determinant of Staphylococcus aureus pyomyositis in a bacterial genome-wide association study
Authors: Young Bernadette C , Earle Sarah G , Soeng Sona , Sar Poda , Kumar Varun , Hor Songly , Sar Vuthy , Bousfield Rachel , Sanderson Nicholas D , Barker Leanne , Stoesser Nicole , Emary Katherine RW , Parry Christopher M , Nickerson Emma K , Turner Paul , Bowden Rory , Crook Derrick , Wyllie David , Day Nicholas PJ , Wilson Daniel J , Moore Catrin E
Published: September 2018

Pyomyositis is a severe bacterial infection of skeletal muscle, commonly affecting children in tropical regions and predominantly caused by Staphylococcus aureus. To understand the contribution of bacterial genomic factors to pyomyositis, we conducted a genome-wide association study of S. aureus cultured from 101 children with pyomyositis and 417 children with asymptomatic nasal carriage attending the Angkor Hospital for Children in Cambodia. We found a strong relationship between bacterial genetic variation and pyomyositis, with estimated heritability 63.8% (95% CI 49.2-78.4%). The presence of the Panton-Valentine leucocidin (PVL) locus increased the odds of pyomyositis 130-fold (p =10-17.9). The signal of association mapped both to the PVL-coding sequence and the sequence immediately upstream. Together these regions explained > 99.9% of heritability. Our results establish staphylococcal pyomyositis, like tetanus and diphtheria, as critically dependent on expression of a single toxin and demonstrate the potential for association studies to identify specific bacterial genes promoting severe human disease.
A Data-Driven Evaluation of the Size and Content of Expanded Carrier Screening Panels
Authors: Ben-Shachar Rotem , Ashley Svenson MS , Goldberg James D. , Muzzey Dale
Published: September 2018

Purpose

The American College of Obstetricians and Gynecologists (ACOG) proposed seven criteria for expanded carrier screening (ECS) panel design. To ensure that screening for a condition is sufficiently sensitive to identify carriers and reduce residual risk of non-carriers, one criterion requires a per-condition carrier rate greater than 1-in-100. However, it is unestablished whether this threshold corresponds with a loss in clinical detection. The impact of the proposed panel-design criteria on at-risk couple detection warrants data-driven evaluation.

Methods

Carrier rates and at-risk couple rates were calculated in 56,281 patients who underwent a 176-condition ECS and evaluated for panels satisfying various criteria. Condition-specific clinical detection rate was estimated via simulation.

Results

Different interpretations of the 1-in-100 criterion have variable impact: a compliant panel would include between 3 and 38 conditions, identify 11%-81% fewer at-risk couples, and detect 36%-79% fewer carriers than a 176-condition panel. If the carrier-rate threshold must be exceeded in all ethnicities, ECS panels would lack prevalent conditions like cystic fibrosis. Simulations suggest that clinical detection rate remains >84% for conditions with carrier rates as low as 1-in-1000.

Conclusions

The 1-in-100 criterion limits at-risk couple detection and should be reconsidered.
A mouse tissue atlas of small non-coding RNA
Authors: Isakova Alina , Fehlmann Tobias , Keller Andreas , Quake Stephen R.
Published: September 2018

Small non-coding RNAs (ncRNAs) play a vital role in a broad range of biological processes both in health and disease. A comprehensive quantitative reference of small ncRNA expression would significantly advance our understanding of ncRNA roles in shaping tissue functions. Here, we systematically profiled the levels of five ncRNA classes (miRNA, snoRNA, snRNA, scaRNA and tRNA fragments) across eleven mouse tissues by deep sequencing. Using fourteen biological replicates spanning both sexes, we identified that ~ 30% of small ncRNAs are distributed across the body in a tissue-specific manner with some are also being sexually dimorphic. We found that miRNAs are subject to “arm switching” between healthy tissues and that tRNA fragments are retained within tissues in both a gene- and a tissue-specific manner. Out of eleven profiled tissues we confirmed that brain contains the largest number of unique small ncRNA transcripts, some of which were previously annotated while others are identified for the first time in this study. Furthermore, by combining these findings with single-cell ATAC-seq data, we were able to connect identified brain-specific ncRNA with their cell types of origin. These results yield the most comprehensive characterization of specific and ubiquitous small RNAs in individual murine tissues to date, and we expect that this data will be a resource for the further identification of ncRNAs involved in tissue-function in health and dysfunction in disease.

HIGHLIGHTS

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An atlas of tissue levels of multiple small ncRNA classes generated from 14 biological replicates of both sexes across 11 tissues

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Distinct distribution patterns of miRNA arms and tRNA fragments across tissues suggest the existence of tissue-specific mechanisms of ncRNA cleavage and retention

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miRNA expression is sex specific in healthy tissues

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Small RNA-seq and scATAC-seq data integration produce a detailed map of cell-type specific ncRNA profiles in the mouse brain
Integrated DNA methylation and gene expression profiling across multiple brain regions implicate novel genes in Alzheimer’s disease
Authors: Semick Stephen A. , Bharadwaj Rahul A. , Collado-Torres Leonardo , Tao Ran , Shin Joo Heon , Deep-Soboslay Amy , Weiss James , Weinberger Daniel R. , Hyde Thomas M. , Kleinman Joel E. , Jaffe Andrew E. , Mattay Venkata S.
Published: September 2018

Background

Late-onset Alzheimer’s disease (AD) is a complex age-related neurodegenerative disorder that likely involves epigenetic factors. To better understand the epigenetic state associated with AD represented as variation in DNA methylation (DNAm), we surveyed 420,852 DNAm sites from neurotypical controls (N=49) and late-onset AD patients (N=24) across four brain regions (hippocampus, entorhinal cortex, dorsolateral prefrontal cortex and cerebellum).

Results

We identified 858 sites with robust differential methylation, collectively annotated to 772 possible genes (FDR<5%, within 10kb). These sites were overrepresented in AD genetic risk loci (p=0.00655), and nearby genes were enriched for processes related to cell-adhesion, immunity, and calcium homeostasis (FDR<5%). We analyzed corresponding RNA-seq data to prioritize 130 genes within 10kb of the differentially methylated sites, which were differentially expressed and had expression levels associated with nearby DNAm levels (p<0.05). This validated gene set includes previously reported (e.g. ANK1, DUSP22) and novel genes involved in Alzheimer’s disease, such as ANKRD30B.

Conclusions

These results highlight DNAm changes in Alzheimer’s disease that have gene expression correlates, implicating DNAm as an epigenetic mechanism underlying pathological molecular changes associated with AD. Furthermore, our framework illustrates the value of integrating epigenetic and transcriptomic data for understanding complex disease.
Ramp-to-Threshold Dynamics in a Hindbrain Population Controls the Timing of Spontaneous Saccades
Authors: Ramirez Alexandro D. , Aksay Emre R.F.
Published: September 2018

Organisms have the capacity to make decisions based solely on internal drives. However, it is unclear how neural circuits form decisions in the absence of sensory stimuli. Here we provide a comprehensive map of the activity patterns underlying the generation of saccades made in the absence of visual stimuli. We performed calcium imaging in the larval zebrafish to discover a range of responses surrounding spontaneous saccades, from cells that displayed tonic discharge only during fixations to neurons whose activity rose in advance of saccades by multiple seconds. We lesioned cells in these populations and found that ablation of neurons with pre-saccadic rise delayed saccade initiation. We analyzed spontaneous saccade initiation using a ramp-to-threshold model and were able to predict the times of upcoming saccades using pre-saccadic activity. These findings suggest that ramping of neuronal activity to a bound is a critical component of self-initiated saccadic movements.
High-Performance Image-Based Measurements of Biological Forces and Interactions in a Dual Optical Trap
Authors: Killian Jessica L. , Inman James T. , Wang Michelle D.
Published: September 2018

Optical traps enable nanoscale manipulation of individual biomolecules while measuring molecular forces and lengths. This ability relies on the sensitive detection of optically trapped particles, typically accomplished using laser-based interferometric methods. Recently, precise and fast image-based particle tracking techniques have garnered increased interest as a potential alternative to laser-based detection, however successful integration of image-based methods into optical trapping instruments for biophysical applications and force measurements has remained elusive. Here we develop a camera-based detection platform that enables exceptionally accurate and precise measurements of biological forces and interactions in a dual optical trap. In demonstration, we stretch and unzip DNA molecules while measuring the relative distances of trapped particles from their trapping centers with sub-nanometer accuracy and precision, a performance level previously only achieved using photodiodes. We then use the DNA unzipping technique to localize bound proteins with extraordinary sub-base-pair precision, revealing how thermal DNA fluctuations allow an unzipping fork to sense and respond to a bound protein prior to a direct encounter. This work significantly advances the capabilities of image tracking in optical traps, providing a state-of-the-art detection method that is accessible, highly flexible, and broadly compatible with diverse experimental substrates and other nanometric techniques.
The Orphan Kinesin PAKRP2 Achieves Processive Motility Via Noncanonical Stepping
Authors: Gicking Allison M. , Wang Pan , Liu Chun , Mickolajczyk Keith J. , Guo Lijun , Hancock William O. , Qiu Weihong
Published: September 2018

PAKRP2 is an orphan kinesin inArabidopsis thalianathat is thought to transport vesicles along phragmoplast microtubules for cell plate formation. Here, using single-molecule fluorescence microscopy, we show that PAKRP2 exhibits processive plus-end-directed motility on single microtubules as individual homodimers despite having an exceptionally long (32 residues) neck linker. Furthermore, using high-resolution nanoparticle tracking to visualize motor stepping dynamics, we find that PAKRP2 achieves processivity via a noncanonical stepping mechanism that includes small step sizes and frequent lateral steps to adjacent protofilaments. We propose that the small steps sizes are due to a transient intermediate step that involves a prolonged diffusional search of the tethered head due to its long neck linker. Despite this different stepping behavior, ATP is tightly coupled to each 8-nm step. Collectively, this study reveals PAKRP2 as the first orphan kinesin to demonstrate processive motility and broadens our understanding of the diverse kinesin stepping mechanisms.
Ratiometric sensing of Pnt and Yan transcription factor levels confers ultrasensitivity to photoreceptor fate transitions in Drosophila
Authors: Bernasek Sebastian M. , Hur Suzy SJ , Peláez-Restrepo Nicolás , Boisclair Lachance Jean-François , Bakker Rachael , Navarro Heliodoro Tejedor , Sanchez-Luege Nicelio , Amaral Luís A. N. , Bagheri Neda , Rebay Ilaria , Carthew Richard W.
Published: September 2018

Cell state transitions are often triggered by large changes in the absolute concentrations of transcription factors and therefore large differences in the stoichiometric ratios between these factors. Whether cells can elicit state transitions using modest changes in the relative ratios of co-expressed factors is unclear. In this study we investigate how cells in theDrosophilaeye resolve cell state transitions by quantifying the expression dynamics of the ETS transcription factors Pnt and Yan. We find that eye progenitor cells maintain a relatively constant ratio of Pnt/Yan protein despite expressing both proteins with pulsatile dynamics. A rapid and sustained two-fold increase in the Pnt/Yan ratio accompanies transitions to photoreceptor fates. Genetic perturbations that modestly disrupt the Pnt/Yan ratio produce fate transition defects consistent with the hypothesis that transitions are normally driven by a two-fold shift in the ratio. A biophysical model based on cooperative Yan-DNA binding coupled with non-cooperative Pnt-DNA binding illustrates how two-fold ratio changes could generate ultrasensitive changes in target gene transcription to drive fate transitions. In this way, coupling cell state transitions to the Pnt/Yan stoichiometric ratio sensitizes the system to modest fold-changes, conferring both robustness and ultrasensitivity to the developmental program.
Miltefosine increases macrophage cholesterol release and inhibits NLRP3-inflammasome assembly and IL-1β release
Authors: Iacano Amanda J , Lewis Harvey , Hazen Jennie E , Andro Heather , Smith Jonathan D , Gulshan Kailash
Published: September 2018

Miltefosine is an FDA approved oral drug for treating cutaneous and visceral leishmaniasis. Leishmania is a flagellated protozoa, which infects and differentiates in macrophages. Here, we studied the effects of Miltefosine on macrophage’s lipid homeostasis, autophagy, and NLRP3 inflammasome assembly/activity. Miltefosine treatment conferred multiple effects on macrophage lipid homeostasis leading to increased cholesterol release from cells, increased lipid-raft disruption, decreased phosphatidylserine (PS) flip from the cell-surface, and redistribution of phosphatidylinositol 4,5-bisphosphate (PIP2) from the plasma membrane to actin rich regions in the cells. Enhanced basal autophagy, lipophagy and mitophagy was observed in cells treated with Miltefosine vs. control. Miltefosine treated cells showed marked increased in phosphorylation of kinases involved in autophagy induction such as; Adenosine monophosphate-activated protein kinase (AMPK) and Unc-51 like autophagy activating kinase (ULK1). The Toll like receptor (TLR) signaling pathway was blunted by Miltefosine treatment, resulting in decreased TLR4 recruitment to cell-surface and ~75% reduction in LPS induced pro-IL-1β mRNA levels. Miltefosine reduced endotoxin-mediated mitochondrial reactive oxygen species and protected the mitochondrial membrane potential. Miltefosine treatment induced mitophagy and dampened NLRP3 inflammasome assembly. Collectively, our data shows that Miltefosine induced ABCA1 mediated cholesterol release, induced AMPK phosphorylation and mitophagy, while dampening NLRP3 inflammasome assembly and IL-1β release.

Significance Statement

Atherosclerosis is driven by cholesterol accumulation and inflammation, and the arterial macrophage is a key cell type in both of these processes. The macrophage characteristics that protect against atherosclerosis include increased cholesterol efflux/reverse cholesterol transport, increased autophagy, and deceased inflammatory cytokine production and signaling. Here, we show that one single orally available compound, Miltefosine, can target multiple macrophage pathways involved in lipid homeostasis and inflammation. Miltefosine activated cholesterol release and autophagy while inhibiting pro IL-1β gene expression and NLRP3 inflammasome assembly. Miltefosine activated AMPK signaling pathway and mitophagy, leading to reduced NLRP3 inflammasome assembly and IL-1β release.
The sleep gene insomniac ubiquitinates targets at postsynaptic densities and is required for retrograde homeostatic signaling
Authors: Kikuma Koto , Li Xiling , Perry Sarah , Li Qiuling , Goel Pragya , Chen Catherine , Kim Daniel , Stavropoulos Nicholas , Dickman Dion
Published: September 2018

The nervous system confronts challenges during development and experience that can destabilize information processing. To adapt to these perturbations, synapses homeostatically adjust synaptic strength, a process referred to as homeostatic synaptic plasticity. At the Drosophila neuromuscular junction, inhibition of postsynaptic glutamate receptors activates retrograde signaling that precisely increases presynaptic neurotransmitter release to restore baseline synaptic strength. However, the nature of the underlying postsynaptic induction process remains enigmatic. Here, we designed a forward genetic screen to identify factors necessary in the postsynaptic compartment to generate retrograde homeostatic signaling. This approach identified insomniac (inc), a gene that encodes a putative adaptor for the Cullin-3 ubiquitin ligase complex and is essential for normal sleep regulation. Intriguingly, we find that Inc rapidly traffics to postsynaptic densities and is required for increased ubiquitination following acute receptor inhibition. Our study suggests that Inc-dependent ubiquitination, compartmentalized at postsynaptic densities, gates retrograde signaling and provides an intriguing molecular link between the control of sleep behavior and homeostatic plasticity at synapses.
Mechanism of IRSp53 inhibition by 14-3-3
Authors: Kast David J. , Dominguez Roberto
Published: September 2018

Filopodia are precursors of dendritic spines and polarized cell migration. The I-BAR-domain protein IRSp53 is an essential regulator of filopodia dynamics that couples Rho-GTPase signaling to cytoskeleton and membrane remodeling, playing essential roles in neuronal development and cell motility. Here, we describe a mechanism whereby phosphorylation-dependent inhibition of IRSp53 by 14-3-3 counters membrane binding and activation by Cdc42 or downstream cytoskeletal effectors. Phosphoproteomics, quantitative binding studies and crystal structures show that 14-3-3 binds to two pairs of phosphorylation sites in IRSp53. Using bicistronic expression we obtained a heterodimer of IRSp53 in which only one subunit is phosphorylated, and show that each subunit of the IRSp53 dimer independently binds a 14-3-3 dimer. A FRET-sensor assay developed using natively phosphorylated and 14-3-3-binding competent IRSp53 purified from mammalian cells reveals opposite conformational changes in IRSp53 upon binding of activatory (Cdc42, Eps8) vs. inhibitory (14-3-3) inputs.
Spatial and temporal inhibition of FGFR2b ligands reveals continuous requirements and novel targets in mouse inner ear morphogenesis
Authors: Urness Lisa D. , Wang Xiaofen , Doan Huy , Shumway Nathan , Noyes C. Albert , Gutierrez-Magana Edgar , Lu Ree , Mansour Suzanne L.
Published: September 2018

Morphogenesis of the inner ear epithelium requires coordinated deployment of several signaling pathways and disruptions cause abnormalities of hearing and/or balance. The FGFR2b ligands, FGF3 and FGF10, are expressed throughout otic development and are required individually for normal morphogenesis, but their prior and redundant roles in otic placode induction complicates investigation of subsequent combinatorial functions in morphogenesis. To interrogate these roles and identify new effectors of FGF3 and FGF10 signaling at the earliest stages of otic morphogenesis, we used conditional gene ablation after otic placode induction and temporal inhibition of signaling with a secreted, dominant-negative FGFR2b ectodomain. We show that both ligands are required continuously after otocyst formation for maintenance of the otic ganglion and patterning and proliferation of the epithelium, leading to normal morphogenesis of both the cochlear and vestibular domains. Furthermore, the first genomewide identification of proximal targets of FGFR2b signaling in the early otocyst reveals novel candidate genes for inner ear development and function.
Structural Basis for Antiarrhythmic Drug Interactions with the Human Cardiac Sodium Channel
Authors: Nguyen Phuong T. , DeMarco Kevin R. , Vorobyov Igor , Clancy Colleen E. , Yarov-Yarovoy Vladimir
Published: September 2018

The human voltage-gated sodium channel, hNav1.5, is responsible for the rapid upstroke of the cardiac action potential and is target for antiarrhythmic therapy. Despite the clinical relevance of hNav1.5 targeting drugs, structure-based molecular mechanisms of promising or problematic drugs have not been investigated at atomic scale to inform drug design. Here, we used Rosetta structural modeling and docking as well as molecular dynamics simulations to study the interactions of antiarrhythmic and local anesthetic drugs with hNav1.5. These calculations revealed several key drug binding sites formed within the pore lumen that can simultaneously accommodate up to two drug molecules. Molecular dynamics simulations identified a hydrophilic access pathway through the intracellular gate and a hydrophobic access pathway through a fenestration between domains III and IV. Our results advance the understanding of molecular mechanisms of antiarrhythmic and local anesthetic drug interactions with hNav1.5 and will be useful for rational design of novel therapeutics.
Effect of Thiobacillus and superabsorbent on essential oil components in Thyme species
Authors: Pouramini Pouneh , Fotokian Mohammad Hossein , Dehghan Hossein , Hensel Goetz
Published: September 2018

Optimal nutrition along with non-stress conditions has a significant impact on the quantity and quality of essential oil in medicinal plants. The objective of this research was to examine the possibility of improving the quantity and quality of essential oil in thyme through nutrition of the seedlings using Thiobacillus bio-fertilizer and treatment by Superabsorbent. For this purpose, seedlings of two thyme species (Thymus vulgaris and Thymus daenensis) were sown in pots and exposed to different levels of Thiobacillus and superabsorbent. Results of Gas Chromatography Mass Spectrometry (GC/MS) revealed that the main compounds detected for T. vulgaris were thymol (31.5%), p-cymene (23.4%), γ-terpinene (13.9%), linalool (38.3%) and carvacrol (2.7%) while the main compounds of T. daenensis were thymol (51.2%), o-cymene (12.9%), γ-terpinene (4.5%), linalool (1.7%) and borneol (3.1%). Furthermore, the application of Thiobacillus had a significant effect on α-pinene content (p = 0.05). Moreover, the interaction between superabsorbent and Thiobacillus significantly changed the percentage of thymol, borneol, and caryophyllene. In conclusion, the mean of essence components in T. vulgaris was more than T. daenensis with the exception of thymol and caryophyllene.
Predicting gene expression in the human malaria parasite Plasmodium falciparum
Authors: Read David F. , Lu Yang Y. , Cook Kate , Roch Karine Le , Noble William Stafford
Published: September 2018

Empirical evidence suggests that the malaria parasite Plasmodium falciparum employs a broad range of mechanisms to regulate gene transcription throughout the organism’s complex life cycle. To better understand this regulatory machinery, we assembled a rich collection of genomic and epigenomic data sets, including information about transcription factor (TF) binding motifs, patterns of covalent histone modifications, nucleosome occupancy, GC content, and global 3D genome architecture. We used these data to train machine learning models to discriminate between high-expression and low-expression genes, focusing on three distinct stages of the red blood cell phase of the Plasmodium life cycle. Our results highlight the importance of histone modifications and 3D chromatin architecture and suggest a relatively small role for TF binding in Plasmodium transcriptional regulation.
Model colibactins exhibit human cell genotoxicity in the absence of host bacteria
Authors: Shine Emilee E. , Xue Mengzhao , Patel Jaymin R. , Healy Alan R. , Surovtseva Yulia V. , Herzon Seth B. , Crawford Jason M.
Published: September 2018

Colibactins are genotoxic secondary metabolites produced in select Enterobacteriaceae, which induce downstream DNA double-strand breaks (DSBs) in human cell lines and are thought to promote the formation of colorectal tumors. Although key structural and functional features of colibactins have been elucidated, the full molecular mechanisms regulating these phenotypes remain unknown. Here, we demonstrate that free model colibactins induce DSBs in human cell cultures and do not require delivery by host bacteria. Through domain-targeted editing, we demonstrate that a subset of native colibactins generated from observed module skipping in the nonribosomal peptide synthetase-polyketide synthase (NRPS-PKS) biosynthetic assembly line share DNA alkylation phenotypes with the model colibactins in vitro. However, module skipping eliminates the strong DNA interstrand cross-links formed by the wildtype pathway in cell culture. This product diversification during the modular NRPS-PKS biosynthesis produces a family of metabolites with varying observed mechanisms of action – DNA alkylation versus crosslinking – in cell culture. The presence of membranes separating human cells from model colibactins attenuated genotoxicity, suggesting that membrane diffusion limits colibactin activity and could account for the observed bacteria-human cell-to-cell contact phenotype. Additionally, extracellular supplementation of the colibactin resistance protein ClbS was able to intercept colibactins in an E. coli-human cell transient infection model. Our studies demonstrate that free model colibactins recapitulate cellular phenotypes associated with moduleskipped products in the native colibactin pathway and define specific protein domains that are required for efficient DNA interstrand crosslinking in the native pathway.
Satellite gravity – enhancements from new satellites and new altimeter technology
Authors: Green C.M. , Fletcher K.M.U. , Cheyney S. , Dawson G.J. , Campbell S.J.
Published: September 2018

This paper reviews the impacts of new satellite altimeter data sets and new technology on the production of satellite gravity. It considers the contribution of the increased data volume, the application of new altimeter acquisition technology and the potential for future developments. Satellite altimeter derived gravity has provided gravity maps of the world's seas since the 1980s, but, from 1995 to 2010, virtually all improvements were in the processing as there were no new satellite data with closely spaced tracks. In recent years, new data from CryoSat‐2 (launched in 2010) and the geodetic mission of Jason‐1 (2012–2013) have provided a wealth of additional coverage and new technology allows further improvements. The synthetic aperture radar mode of CryoSat‐2 uses a scanning approach to limit the size of the altimeter sea surface footprint in the along‐track direction. Tests indicate that this allows reliable data to be acquired closer to coastlines. The synthetic aperture radar interferometric mode of CryoSat‐2 uses two altimeters to locate sea‐surface reflection points laterally away from the satellite track. In a study to generate gravity for freshwater lakes, this mode is found to be valuable in extending the available satellite coverage. The AltiKa altimeter uses higher frequency radar to provide less noisy sea‐surface signals and its new orbit mode gives potential for further improvements in satellite gravity. Future developments include the potential for swath mapping to provide further gravity improvements.
Legitimation as justification: Foregrounding public philosophies in explanations of gradual ideational change
Authors: CARSTENSEN MARTIN B. , HANSEN MAGNUS PAULSEN
Published: September 2018

In accounts of institutional change, discursive institutionalists point to the role of economic and political ideas in upending institutional stability and providing the raw material for the establishment of a new institutional setup. This approach has typically entailed a conceptualisation of ideas as coherent and monolithic and actors as almost automatically following the precepts of the ideas they hold and support. Recent theorising stresses how ideas are in fact composite and heterogeneous, and actors pragmatic and strategic in how they employ ideas in political struggles. However, this change of focus has, until recently, not included how foundational ideas of a polity, often referred to as ‘public philosophies’, are theorised to impact on institution‐building. Drawing on French Pragmatic Sociology, and taking as a starting point recent efforts within discursive institutionalism to conceptualise the dynamic nature of public philosophies, this article seeks to foreground moral justification in accounts of ideational and institutional change. It suggests that public philosophies are reflexively used by actors in continual processes of normative justification that may produce significant policy shifts over time. The empirical relevance of the argument is demonstrated through an analysis of gradual ideational and institutional change in French labour market policy, specifically the development from the state‐guaranteed minimum income scheme of 1988 to the neoliberal make‐work‐pay logic of the 2009 scheme,Revenu de solidarité active. The analysis shows that public and moral justifications have underpinned and gradually shaped these radical changes.
Participatory innovations and maxi‐publics: The influence of participation possibilities on perceived legitimacy at the local level in Finland
Authors: JÄSKE MAIJA
Published: September 2018

Participatory innovations (PIs) have been introduced as one possible cure to democratic malaises. Empirical research on these mechanisms for citizen participation has, however, focused on their effects on individuals and policy outcomes, leaving aside their consequences for the wider public. This article fills part of the gap by examining the effect of PIs on perceived legitimacy. The article acknowledges that citizens value not only outcomes but also the inclusiveness of decision‐making processes, and defines procedural fairness and outcome satisfaction as the key evaluative criteria behind perceived legitimacy. Both total number and type of PIs are considered as possible factors shaping legitimacy evaluations. By analysing data from 9,022 citizens in 30 Finnish municipalities, the article reveals that introducing PIs is not a simple fix for legitimacy of local governments. The type of participation matters, with discursive participation generating the strongest effects on procedural fairness. However, attention should also be paid to citizens’ awareness of participation possibilities.
Conditionally autoregressive models improve occupancy analyses of autocorrelated data: An example with environmental DNA
Authors: Chen Wentao , Ficetola Gentile Francesco
Published: September 2018

Site occupancy‐detection models (SODMs) are statistical models widely used for biodiversity surveys where imperfect detection of species occurs. For instance, SODMs are increasingly used to analyse environmental DNA (eDNA) data, taking into account the occurrence of both false‐positive and false‐negative errors. However, species occurrence data are often characterized by spatial and temporal autocorrelation, which might challenge the use of standard SODMs. Here we reviewed the literature of eDNA biodiversity surveys and found that most of studies do not take into account spatial or temporal autocorrelation. We then demonstrated how the analysis of data with spatial or temporal autocorrelation can be improved by using a conditionally autoregressive SODM, and show its application to environmental DNA data. We tested the autoregressive model on both simulated and real data sets, including chronosequences with different degrees of autocorrelation, and a spatial data set on a virtual landscape. Analyses of simulated data showed that autoregressive SODMs perform better than traditional SODMs in the estimation of key parameters such as true‐/false‐positive rates and show a better discrimination capacity (e.g., higher true skill statistics). The usefulness of autoregressive SODMs was particularly high in data sets with strong autocorrelation. When applied to real eDNA data sets (eDNA from lake sediment cores and freshwater), autoregressive SODM provided more precise estimation of true‐/false‐positive rates, resulting in more reasonable inference of occupancy states. Our results suggest that analyses of occurrence data, such as many applications of eDNA, can be largely improved by applying conditionally autoregressive specifications to SODMs.
Minding the Gap: A Qualitative Study of Provider Experience to Optimize Care for Critically Ill Children in General Emergency Departments
Authors: Query Lindsey A. , Olson Krisjon R. , Meyer Michael T. , Drendel Amy L.
Published: September 2018

Background

Pediatric emergency care provision in the United States is uneven. Institutional barriers to readiness in the general emergency department (GED) are known, but little is understood about the frontline providers. Our objective was to explore the lived experiences of emergency medicine (EM) providers caring for acutely ill children in the GED and identify opportunities to optimize their pediatric practice.

Methods

This grounded theory study used theoretical sampling with snowball recruitment to enroll EM physicians and advanced practice providers from 25 Wisconsin GEDs. Participants completed one‐on‐one, semistructured interviews. Audio recordings were transcribed and coded by a multi‐investigator team drawing on theory produced from comparative analysis.

Results

We reached theoretical saturation with 18 participants. The data suggested that providers felt competent managing routine pediatric care, but critically ill children outstripped their resources and expertise. They recognized environmental constraints on the care they could safely provide, which were intensified by unanticipated knowledge gaps and lack of awareness regarding pediatric practice guidelines. A fragmented medical network to support their pediatric practice was identified as a challenge to their care provision at critical junctures. Due to lack of guidance and feedback, providers internalized their experience with critically ill children with uncertainty, which limited learning and practice change. They benefited from meaningful relationships with pediatricians and pediatric subspecialists, targeted education, timely consults, and looped feedback about care provided and patient outcomes.

Conclusions

General ED providers struggled with critically ill children because they could not anticipate their pediatric‐specific knowledge gaps and only realized them at critical junctures. EM providers were isolated and frustrated when seeking help; without guidance and feedback they internalized their experience with uncertainty and were left underprepared for subsequent encounters. The data suggested the need for provider‐focused interventions to address gaps in pediatric‐specific continuing medical education, just‐in‐time assistance, and knowledge transfer.
Decidual leucocytes infiltrating human spiral arterioles are rich source of matrix metalloproteinases and degrade extracellular matrix in vitro and in situ
Authors: Choudhury Ruhul H. , Dunk Caroline E. , Lye Stephen J. , Harris Lynda K. , Aplin John D. , Jones Rebecca L.
Published: September 2018

Problem

During pregnancy, the decidual spiral arterioles (SpAs) that supply maternal blood to the placenta undergo a series of changes to optimise the transfer of nutrients and oxygen to the developing foetus. Recent studies have shown that initiation of SpA transformation coincides with decidual leucocyte infiltration. Leucocytes are known to be a source of matrix metalloproteinases (MMPs); however, the complete profile of MMPs expressed by decidual NK cells (dNK) and macrophages has not been characterised. We hypothesised that leucocyte‐derived MMPs contribute to SpA remodelling.

Methods

Decidual NK cells and macrophages were isolated from first trimester decidua and their MMP repertoire profiled by qRT‐PCR (n = 10; 5‐11 weeks). Dual immunofluorescence was used to localise MMP expression in situ (n = 3; 5‐12 weeks). Gelatin zymography was carried out to assess whether leucocyte‐derived MMPs can degrade ECM. In situ zymography and immunofluorescence identified MMP activity in tissue‐resident dNK and macrophages.

Results

Decidual NK cells cells and macrophages expressed MMP2, ‐7, ‐9, ‐11, ‐16, ‐19 and tissue inhibitors of metalloproteinase‐1, ‐2, and ‐3. Both cell types degraded gelatin using MMP2 and MMP9 and broke down collagen in an in vitro model of the SpA. Extravillous trophoblasts (EVTs) expressed a similar repertoire of MMPs.

Conclusion

We suggest that matrix remodelling in SpA is initiated by infiltrating leucocytes, while EVTs become involved at later stages.