bluuebunny/crossref_metadata_2025 huggingface dataset:
Deficiency of the Transcription Factor NR4A1 Enhances Bacterial Clearance and Prevents Lung Injury During Escherichia Coli Pneumonia
Authors: Cui Ping , Wu Shuijing , Xu Xia , Ye Hui , Hou Jinchao , Liu Xu , Wang Haihong , Fang Xiangming
Published: May 2018

Background:

Bacterial pneumonia is one of the most common diagnoses and a leading cause of death in the intensive care unit. NR4A1 is an early response gene that has been identified as a vital regulator of immune and inflammatory responses. This study aims to explore the role of NR4A1 in Escherichia coli (E. coli) pneumonia.

Methods:

Alveolar macrophages (AMs) were isolated from wild-type (WT) and NR4A1 knock out (Nr4a1

−/−
) mice, and the NR4A1 expression and phagocytic capacity against E. coli were measured in vitro. WT and Nr4a1

−/−
mice were subjected to E. coli or sham pneumonia. Bacterial load, lung injury severity, inflammatory cell infiltration, and cytokines were assessed at 0, 4, and 18 h after surgery. Survival rates within 48 h were evaluated in WT and Nr4a1

−/−
mice. In addition, NR4A1 antagonist (DIM-C-pPhCO2Me) was also used to confirm the role of NR4A1 in vivo and ex vivo.

Results:

NR4A1 was rapidly induced in AMs at 15 min after E. coli stimulation. Compared with untreated WT AMs, NR4A1 deficiency and DIM-C-pPhCO2Me treatment showed an enhanced phagocytic function (47.72 ± 0.74% vs. 62.3 ± 0.9%, P < 0.001; 11.79 ± 1.21% vs. 30.08 ± 0.79%, P < 0.001, respectively) at 30 min after the E. coli challenge in vitro. NR4A1 deficiency significantly improved the survival rate (33.3% in WT vs. 82.4% in Nr4a1

−/−
, P < 0.01), which is comparable with DIM-C-pPhCO2Me pretreatment. The survival advantage of Nr4a1

−/−
mice was associated with decreased bacterial burden and inflammation and alleviated lung damage.

Conclusions:

These data demonstrate that NR4A1 impairs the phagocytic capacity of AMs and disrupts the host defense against invading bacteria, worsening the outcome of E. coli pneumonia in mice.
Effects of Multiday Ethanol Intoxication on Postburn Inflammation, Lung Function, and Alveolar Macrophage Phenotype
Authors: Curtis Brenda J. , Boe Devin M. , Shults Jill A. , Ramirez Luis , Kovacs Elizabeth J.
Published: May 2018

Burn patients who consumed alcohol before injury have worse clinical outcomes, including longer hospital stays, increased ventilator days, and more respiratory infections. Most alcohol consumers are binge drinkers and not chronic alcoholics, and binge drinking patterns fluctuate over the week, with consecutive days of drinking over the weekend followed by relative abstinence during the week. We used a murine model simulating this drinking pattern in the context of burn injury. Mice were given ethanol for 3 days, rested for 4 days, given ethanol for 3 more days, followed by a sham or 15% total body surface area full-thickness burn. We previously demonstrated that mice exposed to the combined insult exhibited respiratory dysfunction and 50% mortality, with those that succumbed to injury dying between 24 and 72 h, thus identifying a therapeutic intervention window. Our goal herein is to characterize inflammatory and respiratory parameters during this critical time frame. We saw that mice exposed to the combined insult had the highest circulating and pulmonary cytokine levels at 24 h, which were normalized by 72 h in survivors. Alveolar macrophage activation was observed at 24 h in burned mice, regardless of intoxication (P < 0.05). However, at 72 h, alveolar macrophages from intoxicated burned mice had elevated CD206, relative to controls (P < 0.05), indicative of an anti-inflammatory phenotype. Taken together, these findings suggest that although lung function and inflammation are normalized by 72 h, the alterations in alveolar macrophage phenotype shed light on a potential mechanism underlying increased infection susceptibility in intoxicated burn patients.
The Sheep and the Goats: Distinguishing transcriptional enhancers in a complex chromatin landscape
Authors: Sonnenschein Anne , Dworkin Ian , Arnosti David N.
Published: May 2018

Predicting regulatory function of non-coding DNA using genomic information remains a major goal in genomics, and an important step in interpreting the cis-regulatory code. Regulatory capacity can be partially inferred from transcription factor occupancy, histone modifications, motif enrichment, and evolutionary conservation. However, combinations of these features in well-studied systems such asDrosophilahave limited predictive accuracy. Here we examine the current limits of computational enhancer prediction by applying machine-learning methods to an extensive set of genomic features, validating predictions with the Fly Enhancer Resource, which characterized the transcriptional activity of approximately fifteen percent of the genome. Supervised machine learning trained on a range of genomic features identify active elements with a high degree of accuracy, but are less successful at distinguishing tissue-specific expression patterns. Consistent with previous observations of their widespread genomic interactions, many transcription factors were associated with enhancers not known to be direct functional targets. Interestingly, no single factor was necessary for enhancer identification, although binding by the ′pioneer′ transcription factor Zelda was the most predictive feature for enhancer activity. Using an increasing number of predictive features improved classification with diminishing returns. Thus, additional single-timepoint ChIP data may have only marginal utility for discerning true regulatory regions. On the other hand, spatially- and temporally-differentiated genomic features may provide more power for this type of computational enhancer identification. Inclusion of new types of information distinct from current chromatin-immunoprecipitation data may enable more precise identification of enhancers, and further insight into the features that distinguish their biological functions.
Pleotropic effects of PPARD accelerate colorectal tumor progression and invasion
Authors: Liu Yi , Deguchi Yasunori , Tian Rui , Wei Daoyan , Chen Weidong , Xu Min , Wu Ling , Liu Fuyao , Gao Shen , Jaoude Jonathan C. , Chrieki Sarah P. , Moussalli Micheline J. , Gagea Mihai , Morris Jeffrey S. , Broaddus Russell , Zuo Xiangsheng , Shureiqi Imad
Published: May 2018

Colorectal carcinogenesis (CRC) progression requires additional molecular mechanisms to APC mutations/aberrant β-catenin signaling. PPARD is a druggable ligand-activated nuclear receptor that regulates essential genes involved in cell fate. PPARD is upregulated in intestinal epithelial cells (IECs) of human colorectal adenomas and adenocarcinomas. The mechanistic significance of PPARD upregulation in CRC remains unknown. Here we show that targeted PPARD overexpression in IECs of mice strongly augmented β-catenin activation via BMP7/TAK1 signaling, promoted intestinal tumorigenesis in Apcmin mice, and accelerated CRC progression and invasiveness in mice with IEC-targeted ApcΔ580 mutation. Human CRC invasive fronts had higher PPARD expression than their paired adenomas. A PPARD agonist (GW501516) enhanced APCΔ580 mutation-driven CRC, while a PPARD antagonist (GSK3787) suppressed it. Functional proteomics analyses and subsequent validation studies uncovered PPARD upregulation of multiple pro-invasive pathways that drive CRC progression (e.g. PDGFRβ, AKT1, CDK1 and EIF4G1). Our results identify novel mechanisms by which PPARD promotes CRC invasiveness and provide the rational for the development of PPARD antagonists to suppress CRC.
doublesexregulates sexually dimorphic beetle horn formation by integrating spatial and temporal developmental contexts in the Japanese rhinoceros beetleTrypoxylus dichotomus
Authors: Morita Shinichi , Ando Toshiya , Maeno Akiteru , Mizutani Takeshi , Mase Mutsuki , Shigenobu Shuji , Niimi Teruyuki
Published: May 2018

Many scarab beetles have sexually dimorphic exaggerated horns that are an evolutionary novelty. Since the shape, number, size, and location of horns are highly diverged within Scarabaeidae, beetle horns are an attractive model for studying the evolution of sexually dimorphic and novel traits. In beetles including the Japanese rhinoceros beetleTrypoxylus dichotomus, the sex determination genedoublesex(dsx) plays a crucial role in sexually dimorphic horn formation during larval-pupal development. However, knowledge of when and howdsxdrives the gene regulatory network (GRN) for horn formation to form sexually dimorphic horns during development remains elusive. To address this issue, we identified aTrypoxylus-ortholog of the sex determination gene,transformer(tra), that regulates sex-specific splicing of thedsxpre-mRNA, and whose loss of function results in sex transformation. By knocking downtrafunction at multiple developmental timepoints during larval-pupal development, we estimated the onset when the sex-specific GRN for horn formation is driven. In addition, we also revealed thatdsxregulates different aspects of morphogenetic activities during the prepupal and pupal developmental stages to form appropriate morphologies of pupal head and thoracic horn primordia as well as those of adult horns. Based on these findings, we discuss the evolutionary developmental background of sexually dimorphic trait growth in horned beetles.

Author Summary:

Beetle horns are highly enriched in a particular family Scarabaeidae, although the shape, size and number of horns are diversified within the group. In addition, many scarab beetle horns are sexually dimorphic. It has been questioned how a particular group of beetles has originated and diversified evolutionary novel horns. Here we found the exact time when morphological sexual dimorphism of horn primordia appeared, estimated the onset of the developmental program for sexually dimorphic horn formation driven by Doublesex, and revealed that Doublesex regulates different aspects of cell activities of horn primordia depending on the spatiotemporal contexts. Our study provides our understanding regarding regulatory shifts in these mechanisms during the evolution of sexually dimorphic traits in horned beetles.
Attend and Predict: Understanding Gene Regulation by Selective Attention on Chromatin
Authors: Singh Ritambhara , Lanchantin Jack , Sekhon Arshdeep , Qi Yanjun
Published: May 2018

The past decade has seen a revolution in genomic technologies that enabled a flood of genome-wide profiling of chromatin marks. Recent literature tried to understand gene regulation by predicting gene expression from large-scale chromatin measurements. Two fundamental challenges exist for such learning tasks: (1) genome-wide chromatin signals are spatially structured, high-dimensional and highly modular; and (2) the core aim is to understand what the relevant factors are and how they work together. Previous studies either failed to model complex dependencies among input signals or relied on separate feature analysis to explain the decisions. This paper presents an attention-based deep learning approach, AttentiveChrome, that uses a unified architecture to model and to interpret dependencies among chromatin factors for controlling gene regulation. AttentiveChrome uses a hierarchy of multiple Long Short-Term Memory (LSTM) modules to encode the input signals and to model how various chromatin marks cooperate automatically. AttentiveChrome trains two levels of attention jointly with the target prediction, enabling it to attend differentially to relevant marks and to locate important positions per mark. We evaluate the model across 56 different cell types (tasks) in humans. Not only is the proposed architecture more accurate, but its attention scores provide a better interpretation than state-of-the-art feature visualization methods such as saliency maps.1
Systematic Discovery of the Functional Impact of Somatic Genome Alterations in Individual Tumors through Tumor-specific Causal Inference
Authors: Cai Chunhui , Cooper Gregory F. , Lu Kevin N. , Ma Xiaojun , Xu Shuping , Zhao Zhenlong , Chen Xueer , Xue Yifan , Lee Adrian V. , Clark Nathan , Chen Vicky , Lu Songjian , Chen Lujia , Yu Liyue , Hochheiser Harry S , Jiang Xia , Jane Wang Q. , Lu Xinghua
Published: May 2018

We report a tumor-specific causal inference (TCI) framework, which discovers causative somatic genome alterations (SGAs) through inferring causal relationships between SGAs and molecular phenotypes (e.g., transcriptomic, proteomic, or metabolomic changes) within an individual tumor. We applied the TCI algorithm to tumors from The Cancer Genome Atlas (TCGA) and identified those SGAs that causally regulate the differentially expressed genes (DEGs) within each tumor. Overall, TCI identified 634 SGAs that cause cancer-related DEGs in a significant number of tumors, including most of the previously known drivers and many novel candidate cancer drivers. The inferred causal relationships are statistically robust and biologically sensible, and multiple lines of experimental evidence support the predicted functional impact of both well-known and novel candidate drivers. By identifying major candidate drivers and revealing their functional impact in a tumor, TCI shed light on disease mechanisms of each tumor, providing useful information for advancing cancer biology and precision oncology.

Significance statements

Cancer is mainly caused by SGAs. Precision oncology involves identifying and targeting tumor-specific aberrations resulting from causative SGAs. TCI is a novel computational framework for discovering the causative SGAs and their impact on oncogenic processes, thus revealing tumor-specific disease mechanisms. This information can be used to guide precision oncology.
It’s not you, it’s me: Corollary discharge in precerebellar nuclei of sleeping infant rats
Authors: Mukherjee Didhiti , Sokoloff Greta , Blumberg Mark S.
Published: May 2018

In week-old rats, somatosensory input arises predominantly from stimuli in the external environment or from sensory feedback associated with myoclonic twitches during active (REM) sleep. A previous study of neural activity in cerebellar cortex raised the possibility that the brainstem motor structures that produce twitches also send copies of motor commands (or corollary discharge, CD) to the cerebellum. Here, by recording from two precerebellar nuclei—the inferior olive and lateral reticular nucleus—we demonstrate that CD does indeed accompany the production of twitches. Within both structures, the CD signal comprises a surprisingly sharp activity peak within 10 ms of twitch onset. In the inferior olive, this sharp peak is attributable to the opening of slow potassium channels. We conclude that a diversity of neural activity is conveyed to the developing cerebellum preferentially during sleep-related twitching, enabling cerebellar processing of convergent input from CD and reafferent signals.
Modelling structural determinants of ventilation heterogeneity: a perturbative approach
Authors: Whitfield Carl A. , Horsley Alex , Jensen Oliver E.
Published: May 2018

We have developed a computational model of gas mixing and ventilation in the human lung represented as a bifurcating network. We have simulated multiple-breath washout (MBW), a clinical test for measuring ventilation heterogeneity (VH) in patients with obstructive lung conditions. By applying airway constrictions inter-regionally, we have predicted the response of MBW indices to obstructions and found that they detect a narrow range of severe constrictions that reduce airway radius to 10%–30% of healthy values. These results help to explain the success of the MBW test to distinguish obstructive lung conditions from healthy controls. Further, we have used a perturbative approach to account for intra-regional airway heterogeneity that avoids modelling each airway individually. We have found, for random airway heterogeneity, that the variance in MBW indices is greater when indices are already elevated due to constrictions. By quantifying this effect, we have shown that variability in lung structure and mechanical properties alone can lead to clinically significant variability in LCI and Scond, but only in cases simulating obstructive lung conditions. This method is a computationally efficient way to probe the lung’s sensitivity to structural changes, and to quantify uncertainty in predictions due to random variations in lung mechanical and structural properties.
Patterns of tumor progression predict small and tissue-specific tumor-originating niches
Authors: Buder Thomas , Deutsch Andreas , Klink Barbara , Voss-Böhme Anja
Published: May 2018

Cancer development is a multistep process in which cells increase in malignancy through progressive alterations. The early phase of this process is hardly observable which aggravates an understanding of later tumor development. We shed light on this initial phase with a cell-based stochastic model calibrated with epidemiological data from the tissue scale. Our model allows to estimate the number of tumor cells needed for tumor formation in human tissues based on data on the diagnosed ratios of benign and malignant tumors. We find that the minimal number of cells needed for tumor formation is surprisingly small and largely depends on the tissue type. Our results point towards the existence of tumor-originating niches in which the fate of tumor development is early decided. Our estimate for the human colon agrees well with the size of the stem cell niche in colonic crypts. Our estimates might help to identify the tumor-originating cell type, e.g. our analysis suggests for glioblastoma that the tumors originate from a cell type competing in a range of 300 - 1900 cells.

We estimate the number of tumor cells needed for tumor formation in human tissues and propose the existence of small and tissue-specific tumor-originating niches which might help to find tumor-originating cell types, in particular in glioblastoma.
Cellular geometry controls the efficiency of motile sperm aggregates
Authors: Pearce D. J. G. , Hoogerbrugge L. , Hook K. A. , Fisher H. S. , Giomi L.
Published: May 2018

Teams of cooperating sperm have been found across several vertebrate and invertebrate species, ranging from sperm pairs to massive aggregates containing hundreds of cells. Although the biochemical mechanisms involved in sperm-sperm adhesion are unclear, aggregation can enhance the motility of the cells. Here, we report a thorough computational investigation on the role of cellular geometry in the performance of sperm aggregates. The sperm head is modeled as a persistent random walker characterized by a non-trivial three-dimensional shape and equipped with an adhesive region where cell-cell binding occurs. By considering both a simple parametric head shape and a computer reconstruction of a real head shape based on morphometric data, we demonstrate that the geometry of the head and the structure of the adhesive region crucially affect both the stability and motility of the aggregates. Our analysis further suggests that the apical hook, commonly found in the sperm of most muroid rodents, might serve to shield portions of the adhesive region and promote efficient alignment of the velocities of the interacting cells.
Towards measuring growth rates of pathogens during infections by D2O-labeling lipidomics
Authors: Neubauer Cajetan , Sessions Alex L. , Booth Ian R. , Bowen Benjamin P. , Kopf Sebastian H. , Newman Dianne K. , Dalleska Nathan F.
Published: May 2018

RATIONALE

Microbial growth rate is an important physiological parameter that is challenging to measurein situ, partly because microbes grow slowly in many environments. Recently, it has been demonstrated that generation times ofS. aureusin cystic fibrosis (CF) infections can be determined by D2O-labeling of actively synthesized fatty acids. To improve species specificity and allow growth rate monitoring for a greater range of pathogens during the treatment of infections, it is desirable to accurately quantify trace incorporation of deuterium into phospholipids.

METHODS

Lipid extracts of D2O-treatedE. colicultures were measured on LC-ESI-MS instruments equipped with TOF and Orbitrap mass analyzers, and used for comparison with the analysis of fatty acids by isotope-ratio GC-MS. We then develop an approach to enable tracking of lipid labeling, by following the transition from stationary into exponential growth in pure cultures. Lastly, we apply D2O-labeling lipidomics to clinical samples from CF patients with chronic lung infections.

RESULTS

Lipidomics facilitates deuterium quantification in lipids at levels that are useful for many labeling applications (>0.03 at% D). In theE. colicultures, labeling dynamics of phospholipids depend largely on their acyl chains and between phospholipids we notice differences that are not obvious from absolute concentrations alone. For example, cyclopropyl-containing lipids reflect the regulation of cyclopropane fatty acid synthase, which is predominantly expressed at the beginning of stationary phase. The deuterium incorporation into a lipid that is specific forS. aureusin CF sputum, indicates an average generation time of the pathogen on the order of one cell doubling per day.

CONCLUSIONS

This study demonstrates how trace level measurement of stable isotopes in intact lipids can be used to quantify lipid metabolism in pure cultures and provides guidelines that enable growth rate measurements in microbiome samples after incubation with a low percentage of D2O.
A physiological drop in pH decreases mitochondrial respiration, and AMPK and Akt signalling, in L6 myocytes
Authors: Genders Amanda J , Martin Sheree D , McGee Sean L , Bishop David J
Published: May 2018

Exercise stimulates mitochondrial biogenesis and increases mitochondrial respiratory function and content. However, during high-intensity exercise muscle pH can decrease below pH 6.8 with a concomitant increase in lactate concentration. This drop in muscle pH is associated with reduced exercise-induced mitochondrial biogenesis, whilst increased lactate may act as a signaling molecule to affect mitochondrial biogenesis. Therefore, in this study we wished to determine the impact of altering pH and lactate concentration in L6 myotubes on genes and proteins known to be involved in mitochondrial biogenesis. We also examined mitochondrial respiration in response to these perturbations. Differentiated L6 myotubes were exposed to normal (pH 7.5), low (pH 7.0) or high pH (pH 8.0) media with and without 20 mM sodium L-lactate for 1 and 6 h. Low pH and 20 mM Sodium L-Lactate resulted in decreased Akt (Ser473) and AMPK (T172) phosphorylation at 1 h compared to controls, whilst at 6 h the nuclear localisation of HDAC5 was decreased. When the pH was increased both Akt (Ser473) and AMPK (T172) phosphorylation was increased at 1 h. Overall increased lactate decreased the nuclear content of HDAC5 at 6 h. Exposure to both high and low pH media significantly decreased basal mitochondrial respiration, ATP turnover, and maximum mitochondrial respiratory capacity. These data indicate that muscle pH affects several metabolic signalling pathways, including those required for mitochondrial function.
Activity-dependent netrin-1 secretion drives synaptic insertion of GluA1-containing AMPA receptors in the hippocampus
Authors: Glasgow Stephen D. , Labrecque Simon , Beamish Ian V. , Aufmkolk Sarah , Gibon Julien , Han Dong , Harris Stephanie N. , Dufresne Paul , Wiseman Paul W. , Anne McKinney R. , Séguéla Philippe , Koninck Paul De , Ruthazer Edward S. , Kennedy Timothy E.
Published: May 2018

Dynamic trafficking of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid glutamate receptors (AMPARs) to synapses is critical for activity-dependent synaptic plasticity underlying learning and memory, however the identity of key molecular effectors remains elusive. Here, we demonstrate that membrane depolarization and N-methyl-D-aspartate receptor (NMDAR) activation triggers secretion of the chemotropic guidance cue netrin-1 from dendrites. Using selective genetic deletion, we show that netrin-1 expression by excitatory neurons is required for NMDAR-dependent long-term potentiation (LTP) in the adult hippocampus. Further, we demonstrate that application of exogenous netrin-1 is sufficient to trigger the potentiation of excitatory glutamatergic transmission at hippocampal Schaffer collateral synapses via Ca2+-dependent recruitment of GluA1-containing AMPARs, promoting the maturation of immature or nascent synapses. These findings identify a central role for activity-dependent release of netrin-1 as a critical effector of synaptic plasticity in the adult hippocampus.
AgriSeqDB: an online RNA-Seq database for functional studies in agriculturally relevant plant species
Authors: Robinson Andrew J. , Tamiru Muluneh , Salby Rachel , Bolitho Clayton , Williams Andrew , Huggard Simon , Fisch Eva , Unsworth Kathryn , Whelan James , Lewsey Mathew G.
Published: May 2018

Background

The genome-wide expression profile of genes in different tissues/cell types and developmental stages is a vital component of many functional genomic studies. Transcriptome data obtained by RNA-sequencing (RNA-Seq) is often deposited in public databases that are made available via data portals. Data visualization is one of the first steps in assessment and hypothesis generation. However, these databases do not typically include visualization tools and establishing one is not trivial for users who are not computational experts. This, as well as the various formats in which data is commonly deposited, makes the processes of data access, sharing and utility more difficult. Our goal was to provide a simple and user-friendly repository that meets these needs for datasets from major agricultural crops.

Description

AgriSeqDB (https://expression.latrobe.edu.au/agriseqdb), is a database for viewing, analysing and interpreting developmental and tissue/cell-specific transcriptome data from several species, including major agricultural crops such as wheat, rice, maize, barley and tomato. The disparate manner in which public transcriptome data is often warehoused and the challenge of visualizing raw data are both major hurdles to data reuse. The popular eFP browser does an excellent job of presenting transcriptome data in an easily interpretable view, but previous implementation has been mostly on a case-by-case basis. Here we present an integrated visualisation database of transcriptome datasets from six species that did not previously have public-facing visualisations. We combine the eFP browser, for gene-by-gene investigation, with the Degust browser, which enables visualisation of all transcripts across multiple samples. The two visualisation interfaces launch from the same point, enabling users to easily switch between analysis modes. The tools allow users, even those without bioinformatics expertise, to mine into datasets and understand the behaviour of transcripts of interest across samples and time. We have also incorporated an additional graphic download option to simplify incorporation into presentations or publications.

Conclusion

Powered by eFP and Degust browsers, AgriSeqDB is a quick and easy-to-use platform for data analysis and visualization in five crops and Arabidopsis. Furthermore, it provides a tool that makes it easy for researchers to share their datasets, promoting research collaborations and dataset reuse.
Hhex regulates the specification and growth of the hepatopancreatic ductal system
Authors: Villasenor Alethia , Gauvrit Sébastien , Collins Michelle M. , Parajes Silvia , Maischein Hans-Martin , Stainier Didier
Published: May 2018

Significant efforts have advanced our understanding of foregut-derived organ development; however, little is known about the molecular mechanisms that underlie the formation of the hepatopancreatic ductal (HPD) system. Here, we report a role for the homeodomain transcription factor Hhex in directing HPD progenitor specification in zebrafish. Loss of Hhex function results in impaired HPD system formation. We found that Hhex specifies a distinct population of HPD progenitors that gives rise to the cystic duct, common bile duct, and extra-pancreatic duct. Sincehhexis not uniquely expressed in the HPD region but is also expressed in endothelial cells and the yolk syncytial layer (YSL), we tested the role of blood vessels as well as the YSL in HPD formation. We found that blood vessels are required for HPD patterning, but not for HPD progenitor specification. In addition, we found that Hhex is required in both the endoderm and the YSL for HPD development. Our results shed light on the mechanisms necessary to direct endodermal progenitors towards the HPD fate and also advance our understanding of HPD system formation.
C1 CAGE detects transcription start sites and enhancer activity at single-cell resolution
Authors: Kouno Tsukasa , Moody Jonathan , Kwon Andrew , Shibayama Youtaro , Kato Sachi , Huang Yi , Böttcher Michael , Motakis Efthymios , Mendez Mickaёl , Severin Jessica , Luginbühl Joachim , Abugessaisa Imad , Hasegawa Akira , Takizawa Satoshi , Arakawa Takahiro , Furuno Masaaki , Ramalingam Naveen , West Jay , Suzuki Harukazu , Kasukawa Takeya , Lassmann Timo , Hon Chung-Chau , Arner Erik , Carninci Piero , Plessy Charles , Shin Jay W
Published: May 2018

Single-cell transcriptomic profiling is a powerful tool to explore cellular heterogeneity. However, most of these methods focus on the 3’-end of polyadenylated transcripts and provide only a partial view of the transcriptome. We introduce C1 CAGE, a method for the detection of transcript 5’-ends with an original sample multiplexing strategy in the C1™ microfluidic system. We first quantified the performance of C1 CAGE and found it as accurate and sensitive as other methods in C1 system. We then used it to profile promoter and enhancer activities in the cellular response to TGF-β of lung cancer cells and discovered subpopulations of cells differing in their response. We also describe enhancer RNA dynamics revealing transcriptional bursts in subsets of cells with transcripts arising from either strand within a single-cell in a mutually exclusive manner, which was validated using single molecule fluorescence in-situ hybridization.
Ultrasensitive capture of human herpes simplex virus genomes directly from clinical samples reveals extraordinarily limited evolution in cell culture
Authors: Greninger Alexander L. , Roychoudhury Pavitra , Xie Hong , Casto Amanda , Cent Anne , Pepper Gregory , Koelle David M. , Huang Meei-Li , Wald Anna , Johnston Christine , Jerome Keith R.
Published: May 2018

Herpes simplex viruses (HSV) are difficult to sequence due to their large DNA genome, high GC content, and the presence of repeats. To date, most HSV genomes have been recovered from culture isolates, raising concern that these genomes may not accurately represent circulating clinical strains. We report the development and validation of a DNA oligonucleotide hybridization panel to recover near complete HSV genomes at abundances up to 50,000-fold lower than previously reported. Using copy number information on herpesvirus and host DNA background via quantitative PCR, we developed a protocol for pooling for cost-effective recovery of more than 50 HSV-1 or HSV-2 genomes per MiSeq run. We demonstrate the ability to recover >99% of the HSV genome at >100X coverage in 72 hours at viral loads that allow whole genome recovery from latently-infected ganglia. We also report a new computational pipeline for rapid HSV genome assembly and annotation. Using the above tools and a series of 17 HSV-1-positive clinical swabs sent to our laboratory for viral isolation, we show limited evolution of HSV-1 during viral isolation in human fibroblast cells compared to the original clinical samples. Our data indicate that previous studies using low passage clinical isolates of herpes simplex viruses are reflective of the viral sequences present in the lesion and thus can be used in phylogenetic analyses. We also detect superinfection within a single sample with unrelated HSV-1 strains recovered from separate oral lesions in an immunosuppressed patient during a 2.5-week period, illustrating the power of direct-from-specimen sequencing of HSV.

Importance

Herpes simplex viruses affect more than 4 billion people across the globe, constituting a large burden of disease. Understanding global diversity of herpes simplex viruses is important for diagnostics and therapeutics as well as cure research and tracking transmission among humans. To date, most HSV genomics has been performed on culture isolates and DNA swabs with high quantities of virus. We describe the development of wet-lab and computational tools that enable the accurate sequencing of near-complete genomes of HSV-1 and HSV-2 directly from clinical specimens at abundances >50,000-fold lower than previously sequenced and at significantly reduced cost. We use these tools to profile circulating HSV-1 strains in the community and illustrate limited changes to the viral genome during the viral isolation process. These techniques enable cost-effective, rapid sequencing of HSV-1 and HSV-2 genomes that will help enable improved detection, surveillance, and control of this human pathogen.
Development and Validation of a Method for Human Papillomavirus Genotyping Based on Molecular Beacon Probes
Authors: Lyu Jiangfeng , Yu Yuefeng , Ren Xuyi
Published: May 2018

We describe a new assay system for the genotyping of human papillomavirus (HPV) based on linear-after-the-exponential-PCR and melting curve analysis. This system can detect and identify the 23 most common HPV strains (types 6, 11, 16, 18, 31, 33, 35, 39,42, 45, 51, 52, 53, 56, 58, 59, 66, 68, 70, 73, 81, 82, and 83) in two sealed reaction tubes within 2 h. The sensitivity and specificity of this new system was validated using cloned HPV DNA and clinical samples. The detection limit was 5–500 copies/reaction depending on the genotype, and no cross-reactivity was observed with any other low-risk HPV or pathogen that was commonly found in the female genital tract. When compared with the HPV GenoArray test kit, the testing of 1104 clinical samples produced a good overall agreement between the two methods of 98.37% (95% CI: 97.44%–98.97%) and a kappa value of 0.954. Thus, this new HPV genotyping assay system represents a simple, rapid, universally applicable, sensitive, and highly specific detection methodology that should be useful for cervical lesion screening and, thus, is potentially of great value in future clinical applications.
Genotype by environment interactions for the shade avoidance syndrome in the plantAntirrhinum majus, the snapdragon
Authors: Mousset Mathilde , Marin Sara , Archambeau Juliette , Blot Christel , Bonhomme Vincent , Garaud Laura , Pujol Benoit
Published: May 2018

A classical example of phenotypic plasticity in plants is the set of trait changes in response to shade,i.e.the shade avoidance syndrome. There is widespread evidence that plants in low light conditions often avoid shade by growing taller or by increasing their photosynthetic efficiency. This plastic response is expected to have evolved in response to selection in several species, yet there is limited evidence for its genetic variation within populations, which is required for any evolutionary response to selection. In this study, we investigated the shade avoidance syndrome in snapdragon plants (Antirrhinum majus) by using a common garden approach on four natural populations from the Mediterranean region. Our results showed that, in the four populations, individual plants reacted strongly to the presence of shade by growing longer shoots, longer internodes, and increasing their specific leaf area. Our results also revealed genetic variation for the plastic response within these populations, as well as few genetic constraints to its evolution. Our findings imply that the plastic response to shade has the potential to evolve in response to selection in natural populations ofA. majus.
Oral health-related quality of life in elderly women participating in a coexistence group in southern Brazil: Oral health and quality of life in elderly women
Authors: Dallasta Thaís Cauduro , Medina Vanessa Bischoff , Dallepiane Loiva Beatriz
Published: May 2018

The objective of this work was to evaluate the association between quality of life with the oral health in elderly women participating in a coexistence group in Southern Brazil. Study of the descriptive type, analytical, cross-sectional approach, with women aged 60 years or more, participants of a coexistence group in a city in Southern Brazil. Data collection used the instrument Oral Health Impact Profile (OHIP-14). The study had the participation of 64 elderly women aged between 60 and 88 years old with a mean of 69.8 ± 7.31 years. The areas that presented the highest values were “Physical Pain”, “Psychological Distress” and “Physical Disability”. The highest averages of the total scores of the OHIP-14 occurred in individuals with lower family income and low education, who showed signs of depression, changes in taste, difficulty to feel the taste of certain foods and malnutrition. Low education, change of taste and malnutrition by the arm circumference were associated with poor quality of life arising from oral disorders.
Dynamics of supercoiled DNA with complex knots: large-scale rearrangements and persistent multi-strand interlocking
Authors: Coronel Lucia , Suma Antonio , Micheletti Cristian
Published: May 2018

Knots and supercoiling are both introduced in bacterial plasmids by catalytic processes involving DNA strand passages. While the effects on plasmid organization has been extensively studied for knotting and supercoiling taken separately, much less is known about their concurrent action. Here, we use molecular dynamics simulations and oxDNA, an accurate mesoscopic DNA model, to study the kinetic and metric changes introduced by complex (five-crossing) knots and supercoiling in 2kbp-long DNA rings. We find several unexpected results. First, the conformational ensemble is dominated by two distinct states, differing in branchedness and knot size. Secondly, fluctuations between these states are as fast as the metric relaxation of unknotted rings. In spite of this, certain boundaries of knotted and plectonemically-wound regions can persist over much longer timescales. These pinned regions involve multiple strands that are interlocked by the cooperative action of topological and supercoiling constraints. Their long-lived character may be relevant for the simplifying action of topoisomerases.
Highly Multiplexed Genome Engineering Using CRISPR/Cas9 gRNA Arrays
Authors: Kurata Morito , Wolf Natalie K. , Lahr Walker S. , Weg Madison T. , Lee Samantha , Hui Kai , Shiraiwa Masano , Webber Beau R. , Moriarity Branden
Published: May 2018

The CRISPR/Cas9 system is an RNA guided nuclease system that evolved as a mechanism of adaptive immunity in bacteria. This system has been adopted for numerous genome engineering applications in research and recently, therapeutics. The CRISPR/Cas9 system has been largely implemented by delivery of Cas9 as protein, RNA, or plasmid along with a chimeric crRNA-tracrRNA guide RNA (gRNA) under the expression of a pol III promoter, such as U6. Using this approach, multiplex genome engineering has been achieved by delivering several U6-gRNA plasmids targeting multiple loci. However, this approach is limiting due to the efficiently of delivering multiple plasmids to a single cell at one time. To augment the capability and accessibility of multiplexed genome engineering, we developed an efficient golden gate based method to assemble gRNAs linked by optimalCsy4ribonuclease sequences to deliver up to 10 gRNAs as a single gRNA array transcript. Here we report the optimal expression of our guide RNA array under a strong pol II promoter. This system can be implemented alongside the myriad of CRISPR applications, allowing users to model complex biological processes requiring numerous gRNAs.
A retinal circuit generating a dynamic predictive code for orientated features
Authors: Johnston Jamie , Seibel Sofie-Helene , Darnet Léa Simone Adele , Renninger Sabine , Orger Michael , Lagnado Leon
Published: May 2018

Sensory systems must reduce the transmission of redundant information to function efficiently. One strategy is to continuously adjust the sensitivity of neurons to suppress responses to common features of the input while enhancing responses to new ones. Here we image both the excitatory synaptic inputs and outputs of retinal ganglion cells to understand how such dynamic predictive coding is implemented in the analysis of spatial patterns. Synapses of bipolar cells become tuned to orientation through presynaptic inhibition generating lateral antagonism in the orientation domain. Individual ganglion cells receive excitatory synapses tuned to different orientations but feedforward inhibition generates a high-pass filter that only transmits the initial activation of these inputs, thereby removing redundancy. These results demonstrate how a dynamic predictive code can be implemented by circuit motifs common to many parts of the brain.
Immunobiochemical reconstruction of influenza lung infection - Melanoma skin cancer interactions
Authors: Nikolaev Evgeni V. , Zloza Andrew , Sontag Eduardo D.
Published: May 2018

Our recent experimental results that combine a mouse model of influenza A virus (IAV) infection (A/H1N1/PR8) and a highly aggressive model of infection-unrelated cancer, B16-F10 skin melanoma, showed that acute influenza infection of the lung promotes distal melanoma growth in the dermis of the flank and leads to decreased host survival. Here, we proceed to ground the experimental observations in a mechanistic immunobiochemical model that incorporates the T cell receptor signaling pathway, various transcription factors, and a gene regulatory network (GRN). A core component of our model is a biochemical motif, which we call a Triple Incoherent Feed-Forward Loop (TIFFL), and which reflects known interactions between IRF4, Blimp-1, and Bcl-6. The different activity levels of the TIFFL components, as a function of the cognate antigen levels and the given inflammation context, manifest themselves in phenotypically distinct outcomes. Specifically, both the TIFFL reconstruction and quantitative estimates obtained from the model allowed us to formulate a hypothesis that it is the loss of the fundamental TIFFL-induced adaptation of the expression of PD-1 receptors on anti-melanoma CD8+ T cells that constitutes the essence of the previously unrecognized immunologic factor that promotes the experimentally observed distal tumor growth in the presence of acute non-ocogenic infection. We therefore hope that this work can further highlight the importance of adaptive mechanisms by which immune functions contribute to the balance between self and non-self immune tolerance, adaptive resistance, and the strength of TCR-induced activation, thus contributing to the understanding of a broader complexity of fundamental interactions between pathogens and tumors.
The effect of nutrient storage on courtship behavior and copulation frequency in the fruit fly,Drosophila melanogaster
Authors: Bougleux Gomes Hannah Ananda , DiAngelo Justin R. , Santangelo Nicholas
Published: May 2018

Nutrient storage and metabolism effects on reproductive behavior are well studied in higher vertebrates like mammals, but are less understood in simpler systems.Drosophila melanogasteris well suited to study the ramifications of diet and metabolic energy storage on reproductive behaviors as they are commonly used to explore energy mobilization pathways. We tested, for the first time, courtship of the naturally occurringadipose (adp60)mutant which over-accumulates triglycerides and glycogen on a normal diet. We also fed wild type (WT) flies either a normal diet, high fat diet or food deprived them before measuring courtship, copulations, and glycogen and triglyceride levels.Adiposemutants decreased both courtship and copulation frequency, yet showed the highest glycogen and triglyceride levels. We suggest theadp60physique and/or an altered ability to utilize mobilize energy explains these effects. Food deprived WT flies had the lowest glycogen and triglycerides but exhibited shortened courtship latencies with increased courtship behaviors. This may be due to a decreased lifespan of food deprived flies leading to a greater reproductive drive. However, high fat fed flies copulated more frequently and had the highest triglycerides among WT groups, yet equal glycogen levels to the normal fed WT group. Thus, a high fat diet either increases male attractivity or male courtship persistence. Taken together, available diet and nutrient storage affects male fly reproductive behavior in a unique manner, which may be explained by their natural history, and provides a paradigm for understanding energetics based on reproductive potential.
Behavioral effect of chemogenetic inhibition is directly related to receptor transduction levels in rhesus monkeys
Authors: Upright Nicholas A. , Brookshire Stephen W. , Schnebelen Wendy , Damatac Christienne G. , Hof Patrick R. , Browning Philip G. F. , Croxson Paula L. , Rudebeck Peter H. , Baxter Mark G.
Published: May 2018

We used inhibitory DREADDs (Designer Receptors Exclusively Activated by Designer Drugs) to reversibly disrupt dorsolateral prefrontal cortex (dlPFC) function in male macaque monkeys. Monkeys were tested on a spatial delayed response task to assess working memory function after intramuscular injection of either clozapine-N-oxide (CNO) or vehicle. CNO injections given before DREADD transduction were without effect on behavior. rAAV5/hsyn-hM4Di-mCherry was injected bilaterally into the dlPFC of five male rhesus monkeys, to produce neuronal expression of the inhibitory (Gi-coupled) DREADD receptor. We quantified the percentage of DREADD- transduced cells using stereological analysis of mCherry-immunolabeled cells. We found a greater number of immunolabeled neurons in monkeys that displayed CNO-induced behavioral impairment after DREADD transduction compared to monkeys that showed no behavioral effect after CNO. Even in monkeys that showed reliable effects of CNO on behavior after DREADD transduction, the number of prefrontal neurons transduced with DREADD receptor was on the order of 3% of total prefrontal neurons counted. This level of histological analysis facilitates our understanding of behavioral effects, or lack thereof, after DREADD vector injection in monkeys. It also implies that a functional silencing of a relatively small fraction of dlPFC neurons, albeit in a widely distributed area, is sufficient to disrupt spatial working memory.

Significance Statement

Cognitive domains such as working memory and executive function are mediated by the dorsolateral prefrontal cortex (dlPFC). Impairments in these domains are common in neurodegenerative diseases as well as normal aging. The present study sought to measure deficits in a spatial delayed response task following activation of viral-vector transduced inhibitory DREADD (Designer Receptor Exclusively Activated by Designer Drug) receptors in rhesus macaques and compare this to the level of transduction in dlPFC using stereology. We found a significant relationship between the extent of DREADD transduction and the magnitude of behavioral deficit following administration of the DREADD actuator compound clozapine-N- oxide (CNO). These results demonstrate it will be critical to validate transduction to ensure DREADDs remain a powerful tool for neuronal disruption.
LaueG software for displaying and processing neutron Laue images
Authors: Piltz Ross O.
Published: May 2018

An overview is presented of an integrated software package for the processing of Laue data from reactor and other continuous neutron sources. LaueG appears to the user as a graphical user interface with interactive graphic modes that depend on the task being performed: simple image display, cell indexing and refinement, peak integration, wavelength normalization and intensity corrections, or identification of statistical outliers. Other capabilities under development, though still usable, include satellite spots and modulated structures, twinned crystal analysis, and ab initio indexing of unit cells. The emphasis is on accurate intensities for structure refinement using an integrated graphical approach.