bluuebunny/crossref_metadata_2025 huggingface dataset:
Cord blood buffy coat DNA methylation is comparable to whole cord blood methylation
Authors: Dou John , Schmidt Rebecca J. , Benke Kelly S. , Newschaffer Craig , Hertz-Picciotto Irva , Croen Lisa A. , Iosif Ana-Maria , LaSalle Janine M. , Fallin M. Daniele , Bakulski Kelly M.
Published: September 2017

Background

Cord blood DNA methylation is associated with numerous health outcomes and environmental exposures. Whole cord blood DNA reflects all nucleated blood cell types, while centrifuging whole blood separates red blood cells by generating a white blood cell buffy coat. Both sample types are used in DNA methylation studies. Cell types have unique methylation patterns and processing can impact cell distributions, which may influence comparability.

Objectives

To evaluate differences in cell composition and DNA methylation between buffy coat and whole cord blood samples.

Methods

Cord blood DNA methylation was measured with the Infinium EPIC BeadChip (Illumina) in 8 individuals, each contributing buffy coat and whole blood samples. We analyzed principal components (PC) of methylation, performed hierarchical clustering, and computed correlations of mean-centered methylation between pairs. We conducted moderated t-tests on single sites and estimated cell composition.

Results

DNA methylation PCs were associated with individual (PPC1=1.4x10-9;PPC2=2.9x10-5;PPC3=3.8x10-5;PPC4=4.2x10-6;PPC5=9.9x10-13), and not with sample type (PPC1-5>0.7). Samples hierarchically clustered by individual. Pearson correlations of mean-centered methylation between paired individual samples ranged from r=0.66 to r=0.87. No individual site significantly differed between buffy coat and whole cord blood when adjusting for multiple comparisons (5 sites had unadjustedP<10-5). Estimated cell type proportions did not differ by sample type (P=0.86), and estimated cell counts were highly correlated between paired samples (r=0.99).

Conclusions

Differences in methylation and cell composition between buffy coat and whole cord blood are much lower than inter-individual variation, demonstrating that both sample preparation types can be analytically combined and compared.
Mapping DNA damage-dependent genetic interactions in yeast via party mating and barcode fusion genetics
Authors: Díaz-Mejía J. Javier , Celaj Albi , Mellor Joseph C. , Coté Atina , Balint Attila , Ho Brandon , Bansal Pritpal , Shaeri Fatemeh , Gebbia Marinella , Weile Jochen , Verby Marta , Karkhanina Anna , Zhang YiFan , Wong Cassandra , Rich Justin , Prendergast D’Arcy , Gupta Gaurav , Öztürk Sedide , Durocher Daniel , Brown Grant W. , Roth Frederick P.
Published: September 2017

Condition-dependent genetic interactions can reveal functional relationships between genes that are not evident under standard culture conditions. State-of-the-art yeast genetic interaction mapping, which relies on robotic manipulation of arrays of double mutant strains, does not scale readily to multi-condition studies. Here we describe Barcode Fusion Genetics to map Genetic Interactions (BFG-GI), by which double mutant strains generated viaen masse‘party’ mating can also be monitoreden massefor growth and genetic interactions. By using site-specific recombination to fuse two DNA barcodes, each representing a specific gene deletion, BFG-GI enables multiplexed quantitative tracking of double mutants via next-generation sequencing. We applied BFG-GI to a matrix of DNA repair genes under nine different conditions, including methyl methanesulfonate (MMS), 4-nitroquinoline 1-oxide (4NQO), bleomycin, zeocin, and three other DNA-damaging environments. BFG-GI recapitulated known genetic interactions and yielded new condition-dependent genetic interactions. We validated and further explored a subnetwork of condition-dependent genetic interactions involvingMAG1,SLX4, and genes encoding the Shu complex, and inferred that loss of the Shu complex leads to a decrease in the activation or activity of the checkpoint protein kinase Rad53.
Local and global chromatin interactions are altered by large genomic deletions associated with human brain development
Authors: Zhang Xianglong , Zhang Ying , Zhu Xiaowei , Purmann Carolin , Haney Michael S , Ward Thomas , Yao Jie , Weissman Sherman M , Urban Alexander E
Published: September 2017

Background

Large copy number variants (CNVs) in the human genome are strongly associated with common neurodevelopmental, neuropsychiatric disorders such as schizophrenia and autism. Using Hi-C analysis of long-range chromosome interactions, including haplotype-specific Hi-C analysis, and ChIP-Seq analysis of regulatory histone marks, we studied the epigenomic effects of the prominent heterozygous large deletion CNV on chromosome 22q11.2 and also replicated a subset of the findings for the heterozygous large deletion CNV on chromosome 1q21.1.

Results

There are local and global gene expression changes as well as pronounced and multilayered effects on chromatin states, chromosome folding and topological domains of the chromatin, that emanate from the large CNV locus. Regulatory histone marks are altered in the deletion flanking regions, and in opposing directions for activating and repressing marks. Histone marks are changed along chromosome 22q and genome wide. Chromosome interaction patterns are weakened within the deletion boundaries and strengthened between the deletion flanking regions. The long-range folding contacts between the telomeric end of chromosome 22q and the distal deletion-flanking region are increased. On the chromosome 22q with deletion the topological domain spanning the CNV boundaries is deleted in its entirety while neighboring domains interact more intensely with each other. Finally, there is a widespread and complex effect on chromosome interactions genome-wide, i.e. involving all other autosomes, with some of the effect directly tied to the deletion region on 22q11.2.

Conclusions

These findings suggest novel principles of how such large genomic deletions can alter nuclear organization and affect genomic molecular activity.
Automated cell cycle and cell size measurements for single-cell gene expression studies
Authors: Guillemin Anissa , Richard Angelique , Gonin-Giraud Sandrine , Gandrillon Olivier
Published: September 2017

Recent rise of single-cell studies revealed the importance of understanding the role of cell-to-cell variability, especially at the transcriptomic level. One of the numerous sources of cell-to-cell variation in gene expression is the heterogeneity in cell proliferation state. How cell cycle and cell size influences gene expression variability at single-cell level is not yet clearly understood. To deconvolute such influences, most of the single-cell studies used dedicated methods that could include some bias. Here, we provide a universal and automatic toxic-free label method, compatible with single-cell high-throughput RT-qPCR. This led to an unbiased gene expression analysis and could be also used for improving single-cell tracking and imaging when combined with cell isolation. As an application for this technique, we showed that cell-to-cell variability in chicken erythroid progenitors was negligibly influenced by cell size nor cell cycle.
Applying the auxin-inducible degradation (AID) system for rapid protein depletion in mammalian cells
Authors: Lambrus Bramwell G. , Moyer Tyler C. , Holland Andrew J.
Published: September 2017

The ability to deplete a protein of interest is critical for dissecting cellular processes. Traditional methods of protein depletion are often slow-acting, which can be problematic when characterizing a cellular process that occurs within a short period of time. Furthermore, these methods are usually not reversible. Recent advances to achieve protein depletion function by inducibly trafficking proteins of interest to an endogenous E3 ubiquitin ligase complex to promote ubiquitination and subsequent degradation by the proteasome. One of these systems, the auxin-inducible degron (AID) system, has been shown to permit rapid and inducible degradation of AID-tagged target proteins in mammalian cells. The AID system can control the abundance of a diverse set of cellular proteins, including those contained within protein complexes, and is active in all phases of the cell cycle. Here we discuss considerations for the successful implementation of the AID system and describe a protocol using CRISPR/Cas9 to achieve bi-allelic insertion of an AID degron in human cells. This method can also be adapted to insert other tags, such as fluorescent proteins, at defined genomic locations.
Neck muscle spindle noise biases reaches in a multi-sensory integration task
Authors: Khoozani Parisa Abedi , Blohm Gunnar
Published: September 2017

Reference frame Transformations (RFTs) are crucial components of sensorimotor transformations in the brain. Stochasticity in RFTs has been suggested to add noise to the transformed signal due to variability in transformation parameter estimates (e.g. angle) as well as the stochastic nature of computations in spiking networks of neurons. Here, we varied the RFT angle together with the associated variability and evaluated the behavioral impact in a reaching task that required variability-dependent visual-proprioceptive multi-sensory integration. Crucially, reaches were performed with the head either straight or rolled 30deg to either shoulder and we also applied neck loads of 0 or 1.8kg (left or right) in a 3×3 design, resulting in different combinations of estimated head roll angle magnitude and variance required in RFTs. A novel 3D stochastic model of multi-sensory integration across reference frames was fitted to the data and captured our main behavioral findings: (1) neck load biased head angle estimation across all head roll orientations resulting in systematic shifts in reach errors; (2) Increased neck muscle tone led to increased reach variability, due to signal-dependent noise; (3) both head roll and neck load created larger angular errors in reaches to visual targets away from the body compared to reaches toward the body. These results show that noise in muscle spindles and stochasticity in general have a tangible effect on RFTs underlying reach planning. Since RFTs are omnipresent in the brain, our results could have implication for processes as diverse as motor control, decision making, posture / balance control, and perception.

New & Noteworthy

We show that increasing neck muscle tone systematically biases reach movements. A novel 3D multisensory integration across reference frames model captures the data well and provides evidence that the brain must have online knowledge of full body geometry together with the associated variability to accurately plan reach movements.
Single-cell analysis of early antiviral gene expression reveals a determinant of stochasticIFNB1expression
Authors: Doğanay Sultan , Lee Maurice Youzong , Baum Alina , Peh Jessie , Hwang Sun-Young , Yoo Joo-Yeon , Hergenrother Paul J. , García-Sastre Adolfo , Myong Sua , Ha Taekjip
Published: September 2017

RIG-I-like receptors (RLRs) are cytoplasmic sensors of viral RNA that trigger the signaling cascade that leads to type I interferon (IFN) production. Transcriptional induction of RLRs by IFN is believed to play the role of positive feedback to further amplify viral sensing. We found that RLRs and several other IFN-stimulated genes (ISGs) are induced early in viral infection independent of IFN. Expression of these early ISGs requires IRF3/IRF7 and is highly correlated amongst them. Simultaneous detection of mRNA ofIFNB1, viral replicase, and ISGs revealed distinct populations ofIFNB1expressing and non-expressing cells which are highly correlated with the levels of early ISGs but are uncorrelated with IFN-dependent ISGs and viral gene expression. Individual expression of RLRs madeIFNB1expression more robust and earlier, suggesting a causal relation between levels of RLR and induction of IFN.
CancerDiscover: A configurable pipeline for cancer prediction and biomarker identification using machine learning framework
Authors: Mohammed Akram , Biegert Greyson , Adamec Jiri , Helikar Tomáš
Published: September 2017

Motivation

Use of various high-throughput screening techniques has resulted in an abundance of data, whose complete utility is limited by the tools available for processing and analysis. Machine learning holds great potential for deciphering these data in the context of cancer classification and biomarker identification. However, current machine learning tools require manual processing of raw data from various sequencing platforms, which is both tedious and time-consuming. The current classification tools lack flexibility in choosing the best feature selection algorithms from a range of algorithms and most importantly inability to compare various learning algorithms.

Results

We developed CancerDiscover, an open-source software pipeline that allows users to efficiently and automatically integrate large high-throughput datasets, preprocess, normalize, and selects best performing features from multiple feature selection algorithms. The pipeline lets users apply various learning algorithms and generates multiple classification models and evaluation reports that distinguish cancer from normal samples, as well as different types and subtypes of cancer.

Availability and Implementation

The open source pipeline is freely available for download athttps://github.com/HelikarLab/CancerDiscover.

Contact

elikar2@unl.edu

Supplementary Information

Please refer to the CancerDiscover README (Supplementary File 1) for detailed instructions on installation and operation of the pipeline. For a list of available feature selection methods, see Supplementary File 2.
Hemodynamic forces tune the arrest, adhesion and extravasation of circulating tumor cells
Authors: Follain Gautier , Osmani Naël , Azevedo Sofia , Allio Guillaume , Mercier Luc , Karreman Matthia A. , Solecki Gergely , Garcia-Leon Maria Jesus , Lefebvre Olivier , Fekonja Nina , Hille Claudia , Chabannes Vincent , Dollé Guillaume , Metivet Thibaut , Der Hovsepian François , Prudhomme Christophe , Pichot Angélique , Paul Nicodème , Carapito Raphaël , Bahram Siamak , Ruthensteiner Bernhard , Kemmling André , Siemonsen Susanne , Schneider Tanja , Fiehler Jens , Glatzel Markus , Winkler Frank , Schwab Yannick , Pantel Klaus , Harlepp Sébastien , Goetz Jacky G.
Published: September 2017

Metastatic seeding is driven by cell-intrinsic and environmental cues, yet the contribution of biomechanics is poorly known. We aim to elucidate the impact of blood flow on the arrest and the extravasation of circulating tumor cells (CTCs)in vivo. Using the zebrafish embryo, we show that arrest of CTCs occurs in vessels with favorable flow profiles where flow forces control the adhesion efficacy of CTCs to the endothelium. We biophysically identified the threshold values of flow and adhesion forces allowing successful arrest of CTCs. In addition, flow forces fine-tune tumor cell extravasation by impairing the remodeling properties of the endothelium. Importantly, we also observe endothelial remodeling at arrest sites of CTCs in mouse brain capillaries. Finally, we observed that human supratentorial brain metastases preferably develop in areas with low perfusion. Altogether, these results demonstrate that hemodynamic profiles at metastatic sites regulate key steps of extravasation preceding metastatic outgrowth.
Recovering genomic clusters of secondary metabolites from lakes: a Metagenomics 2.0 approach
Authors: Cuadrat Rafael R. C. , Ionescu Danny , Davila Alberto M. R. , Grossart Hans-Peter
Published: September 2017

Background

Metagenomic approaches became increasingly popular in the past decades due to decreasing costs of DNA sequencing and bioinformatics development. So far, however, the recovery of long genes coding for secondary metabolism still represents a big challenge. Often, the quality of metagenome assemblies is poor, especially in environments with a high microbial diversity where sequence coverage is low and complexity of natural communities high. Recently, new and improved algorithms for binning environmental reads and contigs have been developed to overcome such limitations. Some of these algorithms use a similarity detection approach to classify the obtained reads into taxonomical units and to assemble draft genomes. This approach, however, is quite limited since it can classify exclusively sequences similar to those available (and well classified) in the databases.

In this work, we used draft genomes from Lake Stechlin, north-eastern Germany, recovered by MetaBat, an efficient binning tool that integrates empirical probabilistic distances of genome abundance, and tetranucleotide frequency for accurate metagenome binning. These genomes were screened for secondary metabolism genes, such as polyketide synthases (PKS) and non-ribosomal peptide synthases (NRPS), using the Anti-SMASH and NAPDOS workflows.

Results

With this approach we were able to identify 243 secondary metabolite clusters from 121 genomes recovered from the lake samples. A total of 18 NRPS, 19 PKS and 3 hybrid PKS/NRPS clusters were found. In addition, it was possible to predict the partial structure of several secondary metabolite clusters allowing for taxonomical classifications and phylogenetic inferences.

Conclusions

Our approach revealed a great potential to recover and study secondary metabolites genes from any aquatic ecosystem.
Timescales of influenza A/H3N2 antibody dynamics
Authors: Kucharski Adam J. , Lessler Justin , Cummings Derek A.T. , Riley Steven
Published: September 2017

Human immunity influences the evolution and impact of novel influenza strains. Because individuals are infected with multiple influenza strains during their lifetime and each virus can generate a cross-reactive antibody response, it is challenging to quantify the processes that shape observed immune responses, or to reliably detect recent infection from serological samples. Using a Bayesian model of antibody dynamics at multiple timescales, we explain complex cross-reactive antibody landscapes by inferring participants’ histories of infection with serological data from cross-sectional and longitudinal studies of influenza A/H3N2 in southern China and Vietnam. We show an individual’s influenza antibody profile can be explained by a short-lived, broadly cross-reactive response that decays within a year to leave a smaller long-term response acting against a narrower range of strains. We also demonstrate that accounting for dynamic immune responses can provide a more accurate alternative to traditional definitions seroconversion for the estimation of infection attack rates. Our work provides a general model for explaining mechanisms of influenza immunity acting at multiple timescales based on contemporary serological data, and suggests a two-armed immune response to influenza infection consistent with competitive dynamics between B cell populations. This approach to analysing multiple timescales for antigenic responses could also be applied to other multi-strain pathogens such as dengue and related flaviviruses.
Hsf1 and Hsp70 constitute a two-component feedback loop that regulates the yeast heat shock response
Authors: Krakowiak Joanna , Zheng Xu , Patel Nikit , Anandhakumar Jayamani , Valerius Kendra , Gross David S. , Khalil Ahmad S. , Pincus David
Published: September 2017

Models for regulation of the eukaryotic heat shock response typically invoke a negative feedback loop consisting of the transcriptional activator Hsf1 and a molecular chaperone encoded by an Hsf1 target gene. Previously, we identified Hsp70 as the chaperone responsible for Hsf1 repression inSaccharomyces cerevisiaeand constructed a mathematical model based on Hsp70-mediated negative feedback that recapitulated the dynamic activity of Hsf1 during heat shock. The model was based on two assumptions: dissociation of Hsp70 activates Hsf1, and transcriptional induction of Hsp70 deactivates Hsf1. Here we validated these assumptions. First, we severed the feedback loop by uncoupling Hsp70 expression from Hsf1 regulation. As predicted by the model, Hsf1 was unable to efficiently deactivate in the absence of Hsp70 transcriptional induction. Next we mapped a discrete Hsp70 binding site on Hsf1 to a motif in the C-terminal activation domain known as conserved element 2 (CE2). Removal of CE2 resulted in increased Hsf1 activity under non-heat shock conditions and delayed deactivation kinetics. In addition, we uncovered a role for the N-terminal domain of Hsf1 in negatively regulating DNA binding. These results reveal the quantitative control mechanisms underlying the feedback loop charged with maintaining cytosolic proteostasis.
Fitting mathematical models of biochemical pathways to steady state perturbation response data without simulating perturbation experiments
Authors: Santra Tapesh
Published: September 2017

A common experimental approach for studying signal transduction networks (STNs) is to measure the steady state concentrations of their components following perturbations to individual components. Such data is frequently used to reconstruct topological models of STNs, but, are rarely used for calibrating kinetic models of these networks. This is because, existing calibration algorithms operate by assigning different sets of values to the parameters of the kinetic models, and for each set of values simulating all perturbations performed in the biochemical experiments. This process is highly computation intensive and may be infeasible when molecular level information of the perturbation experiments is unavailable. Here, I propose an algorithm which can calibrate ordinary differential equation (ODE) based kinetic models of STNs using steady-state perturbation responses (SSPRs) without simulating perturbation experiments. The proposed algorithm uses modular response analysis (MRA) to calculate the scaled Jacobian matrix of the ODE model of an STN using SSPR data. The model parameters are then calibrated to fit the scaled Jacobian matrix calculated in the above step. This procedure does not require simulating the perturbation experiments. Therefore, it is significantly less computation intensive than existing algorithms and can be implemented without molecular level knowledge of the mechanism of perturbations. It is also parallelizable, i.e. can explore multiple sets of parameter values simultaneously, and therefore is scalable. The capabilities and shortcomings of the proposed algorithm are demonstrated using both simulated and real perturbation responses of Mitogen Activated Protein Kinase (MAPK) STN.

Availability

All source codes and data needed to replicate the results in this manuscript are available fromhttps://github.com/SBIUCD/MRA_SMC_ABC1
Metabolite profile and mitochondrial energetics characterize poor early recovery of muscle mass following hind limb unloading in old mice
Authors: Zhang Xiaolei , Trevino Michelle B. , Wang Miao , Gardell Stephen J. , Ayala Julio E. , Han Xianlin , Kelly Daniel P. , Goodpaster Bret H. , Vega Rick B. , Coen Paul M.
Published: September 2017

The progression of age-related sarcopenia can be accelerated by impaired recovery of muscle mass following periods of disuse due to illness or immobilization. The molecular underpinnings of poor recovery of aging muscle following disuse remain largely unknown. However, recent evidence suggests that mitochondrial energetics may play an important role. Here, we report that 22-24 month old mice with low muscle mass and insulin resistance display poor early recovery of muscle mass following 10 days of hind limb unloading. We took an unbiased approach to identify changes in energy metabolism gene expression and metabolite pools and show for the first time that persistent mitochondrial dysfunction, dysregulated fatty acid β-oxidation and elevated H2O2emission underlie poor early recovery of muscle mass following a period of disuse in old mice. Importantly, this is linked to more severe whole-body insulin resistance. The findings suggest that muscle fuel metabolism and mitochondrial energetics should be a focus for mining therapeutic targets to improve recovery of muscle mass following periods of disuse in older animals.
Validation of RT-qPCR approaches to monitorPseudomonas syringaegene expression during infection and exposure to pattern-triggered immunity
Authors: Smith Amy , Lovelace Amelia H. , Kvitko Brian H.
Published: September 2017

Pseudomonas syringaepv. tomato DC3000 (DC3000) is an important model plant pathogen, with a fully annotated genome and multiple compatible plant hosts. Very few studies have examined the regulation of DC3000 gene expressionin vivo. We developed a RT-qPCR assay to monitor transcriptional changes in DC3000 inoculated intoArabidopsis thalianaleaves during disease and exposure to pattern-triggered immunity (PTI). In our approach, bacterial RNA concentrations in total tissue RNA are standardized usingP.syringae-specific16S ribosomal RNA primers. We validated multiple stable reference genes for normalization in calculating the relative expression of genes of interest. We used empirically derived rates of amplification efficiency to calculate relative expression of key marker genes for virulence-associated regulation. We demonstrated that exposure to PTI alters DC3000 expression of Type III secretion system, coronatine synthesis genes and flagellar marker genes.
Adeno-associated virus mediated gene delivery: Implications for scalablein vitroandin vivocardiac optogenetic models
Authors: Ambrosi Christina M. , Sadananda Gouri , Klimas Aleksandra , Entcheva Emilia
Published: September 2017

Aims

Adeno-associated viruses (AAVs) provide advantages in long-term, cardiac-specific gene expression. However, AAV serotype specificity data is lacking in cardiac models relevant to optogenetics. We aimed to identify the optimal AAV serotype (1, 6, or 9) in pursuit of scalable rodent and human models for cardiac optogenetics and elucidate the mechanism of virus uptake.

Methods

In vitrosyncytia of primary neonatal rat ventricular cardiomyocytes (NRVMs) and human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) were infected with AAVs 1, 6, and 9 containing the transgene for eGFP or channelrhodopsin-2 (ChR2) fused to mCherry.In vivoadult rats were intravenously injected with AAV1 and 9 containing ChR2-mCherry.

Results

Transgene expression profiles of rat and human cellsin vitrorevealed that AAV1 and 6 significantly outperformed AAV9. In contrast, systemic delivery of AAV9 in adult rat hearts yielded significantly higher levels of ChR2-mCherry expression and optogenetic responsiveness. We tracked the mechanism of virus uptake to purported receptor-mediators for AAV 1/6 (cell surface sialic acid) and AAV9(37/67kDa laminin receptor, LamR).In vitrodesialylation of NRVMs and hiPSC-CMs with neuraminidase significantly decreased AAV1,6-mediated gene expression, but interestingly, desialylation of hiPSC-CMs increased AAV9-mediated expression. In fact, only very high viral doses of AAV9-ChR2-mCherry, combined with neuraminidase treatment yielded consistent optogenetic responsiveness in hiPSC-CMs. Differences between thein vitroandin vivoperformance of AAV9 could be correlated to robust LamR expression in the adult and neonatal rat hearts, but no expressionin vitroin cultured cells. The dynamic nature of LamR expression and its dependence on environmental factors was further corroborated in intact adult human ventricular tissue slices.

Conclusion

The combined transgene expression and cell surface receptor data may explain the preferential efficiency of AAV1/6in vitroand AAV9in vivofor cardiac delivery and mechanistic knowledge of their action can help guide cardiac optogenetic efforts.
Transcriptomes of major renal collecting-duct cell types in mouse identified by single-cell RNA-Seq
Authors: Chen Lihe , Lee Jae Wook , Chou Chung-Lin , Nair Anilkumar , Battistone Maria Agustina , Paunescu Teodor , Mekulova Maria , Breton Sylvie , Verlander Jill W. , Wall Susan , Brown Dennis , Burg Maurice B. , Knepper Mark A.
Published: September 2017

Prior RNA sequencing (RNA-Seq) studies have identified complete transcriptomes for most renal epithelial cell types. The exceptions are the cell types that make up the renal collecting duct, namely intercalated cells (ICs) and principal cells (PCs), which account for only a small fraction of the kidney mass, but play critical physiological roles in the regulation of blood pressure, extracellular fluid volume and extracellular fluid composition. To enrich these cell types, we used fluorescence-activated cell sorting (FACS) that employed well established lectin cell surface markers for PCs and type B ICs, as well as a newly identified cell surface marker for type A ICs, viz. c-Kit. Single-cell RNA-Seq using the 1C- and PC-enriched populations as input enabled identification of complete transcriptomes of A-ICs, B-ICs and PCs. The data were used to create a freely-accessible online gene-expression database for collecting duct cells. This database allowed identification of genes that are selectively expressed in each cell type including cell-surface receptors, transcription factors, transporters and secreted proteins. The analysis also identified a small fraction of hybrid cells expressing both aquapor¡n-2 and either anion exchanger 1 or pendrin transcripts. In many cases, mRNAs for receptors and their ligands were identified in different cells (e.g.Notch2chiefly in PCs vsJag1chiefly in ICs) suggesting signaling crosstalk among the three cell types. The identified patterns of gene expression among the three types of collecting duct cells provide a foundation for understanding physiological regulation and pathophysiology in the renal collecting duct.

SIGNIFICANCE STATEMENT

A long-term goal in mammalian biology is to identify the genes expressed in every cell type of the body. In kidney, the expressed genes (“transcriptome”) of all epithelial cell types have already been identified with the exception of the cells that make up the renal collecting duct, responsible for regulation of blood pressure and body fluid composition. Here, a technique called "single-cell RNA-Seq" was used in mouse to identify transcriptomes for the major collecting-duct cell types: type A intercalated cells, type B intercalated cells and principal cells. The information was used to create a publicly-accessible online resource. The data allowed identification of genes that are selectively expressed in each cell type, informative for cell-level understanding of physiology and pathophysiology.
SEMI-PARAMETRIC COVARIATE-MODULATED LOCAL FALSE DISCOVERY RATE FOR GENOME-WIDE ASSOCIATION STUDIES
Authors: Zablocki Rong W. , Levine Richard A. , Schork Andrew J. , Xu Shujing , Wang Yunpeng , Fan Chun C. , Thompson Wesley K.
Published: September 2017

While genome-wide association studies (GWAS) have discovered thousands of risk loci for heritable disorders, so far even very large meta-analyses have recovered only a fraction of the heritability of most complex traits. Recent work utilizing variance components models has demonstrated that a larger fraction of the heritability of complex phenotypes is captured by the additive effects of SNPs than is evident only in loci surpassing genome-wide significance thresholds, typically set at a Bonferroni-inspiredp≤ 5 x 10-8. Procedures that control false discovery rate can be more powerful, yet these are still under-powered to detect the majority of non-null effects from GWAS. The current work proposes a novel Bayesian semi-parametric two-group mixture model and develops a Markov Chain Monte Carlo (MCMC) algorithm for a covariate-modulated local false discovery rate (cmfdr). The probability of being non-null depends on a set of covariates via a logistic function, and the non-null distribution is approximated as a linear combination of B-spline densities, where the weight of each B-spline density depends on a multinomial function of the covariates. The proposed methods were motivated by work on a large meta-analysis of schizophrenia GWAS performed by the Psychiatric Genetics Consortium (PGC). We show that the new cmfdr model fits the PGC schizophrenia GWAS test statistics well, performing better than our previously proposed parametric gamma model for estimating the non-null density and substantially improving power over usual fdr. Using loci declared significant at cmfdr ≤ 0.20, we perform follow-up pathway analyses using the Kyoto Encyclopedia of Genes and Genomes (KEGG)homo sapienspathways database. We demonstrate that the increased yield from the cmfdr model results in an improved ability to test for pathways associated with schizophrenia compared to using those SNPs selected according to usual fdr.
Synchronous Induction of Meiosis in the Fission Yeast Schizosaccharomyces pombe
Authors: Yamashita Akira , Sakuno Takeshi , Watanabe Yoshinori , Yamamoto Masayuki
Published: September 2017

In fission yeast Schizosaccharomyces pombe, initiation of meiosis is repressed by Pat1 kinase. This protocol describes how ectopic inactivation of the temperature-sensitive Pat1-114 kinase in G1-arrested h−/h− diploid cells carrying mat1-Pc induces a highly synchronized commitment to and execution of meiosis. Haploid or diploid pat1-114 mutants without mat1-Pc can also be used for convenience, although less synchrony may be attained compared with induction using true diploids. An essentially identical protocol can be used for induction via inhibition of genetically sensitized Pat1 kinase by ATP analogs.
Using Phage Display to Create Recombinant Antibodies
Authors: Dasch James R. , Dasch Amy L.
Published: September 2017

A variety of phage display technologies have been developed since the approach was first described for antibodies. The most widely used approaches incorporate antibody sequences into the minor coat protein pIII of the nonlytic filamentous phage fd or M13. Libraries of variable gene sequences, encoding either scFv or Fab fragments, are made by incorporating sequences into phagemid vectors. The phagemid is packaged into phage particles with the assistance of a helper phage to produce the antibody display phage. This protocol describes a method for creating a phagemid library. The multiple cloning site (MCS) of the pBluescript KS(–) phagemid vector is replaced by digestion with the restriction enzyme BssHII, followed by the insertion of four overlapping oligonucleotides to create a new MCS within the vector. Next, the 3′ portion of gene III (from M13mp18) is amplified and combined with an antibody sequence using overlap extension PCR. This product is inserted into the phagemid vector to create pPDS. Two helper plasmids are also created from the modified pBluescript vector: pLINK provides the linker between the heavy and light chains, and pFABC provides the CH1 domain of the heavy chain. An antibody cDNA library is constructed from the RNA of interest and ligated into pPDS. The phagemid library is electroporated into Escherichia coli cells along with the VCS-M13 helper phage.
Genome-Wide RNA Interference: Functional Genomics in the Postgenomics Era
Authors: Politi Katerina , Wajapeyee Narendra
Published: September 2017

This introduction briefly describes the types of RNAi libraries (both shRNA-based and double-stranded siRNA-based) that are available for understanding diverse biological questions and then discusses recent advances in RNAi screening methodologies in mouse, rat, humans, Drosophila, and worms.
Dynamics of environmental consciousness and green purchase behaviour: an empirical study
Authors: Mishal Aditi , Dubey Rameshwar , Gupta Omprakash K. , Luo Zongwei
Published: September 2017

Purpose

The purpose of this paper is to investigate the relationships between environmental consciousness (ECO), green purchase attitude (GPA), green purchase intention (GPI), perceived customer effectiveness (PCE), green behaviour (GRB) and green purchase behaviour (GPB). Based on the statistical analyses, this paper offers some further research directions to advance the extant literature.

Design/methodology/approach

The theoretical model is firmly grounded in extant literature. To test the study hypotheses, the authors have developed a survey instrument following a two-stage process. The constructs were first operationalized by the authors and then pre-tested by experts. Dillman’s (2007) guidelines were then followed to gather data. Finally, the theoretical model was tested using multivariate statistical tools.

Findings

Results indicate that ECO has an influence on GPA and PCE; GPA has an influence on PCE and GRB; GPI has an influence on PCE; and GRB has an influence on GPB. Environmental benefit still ranks at the sixth position among eight product-selection criteria, as is evident from qualitative in-depth interviews indicating a primarily rationalistic and not an altruistic purchase approach. The gap in translation of ECO into GB and GPB can be attributed to costliness, non-availability with less variety, lack of brand reputation of green products and budget constraints for customers.

Research limitations/implications

The study faces the limitation of generalizability of the results because it was carried out in a particular state in India; it may not be the perception of the country as a whole. The bias owing to social desirability, selective memory and telescoping with the use of self-reported data could also be a limitation for the current empirical study.

Originality/value

This study aimed to extend pro-environmental behaviour studies beyond developed countries and to empirically validate the models built on the theory of ECO leading to GPB, especially for India, a rising market. A novel approach to empirically discuss the situational and market factors will provide a much-needed thrust for research on these lines.
Promoting educational opportunity and achievement through 1:1 iPads
Authors: Thieman Gayle Y. , Cevallos Tatiana
Published: September 2017

Purpose

The purpose of this paper is to expand the growing body of research on the educational impact of 1:1 mobile devices, investigating the iPad’s potential to reduce the disparity of access to high-quality instructional technology and achievement for low income, racially, and linguistically diverse students.

Design/methodology/approach

This three-year, mixed-method study investigated the degree to which a 1:1 iPad initiative reduced the disparity of technology access and instructional use and improved student learning and attendance. The research design included survey data on student technology skills, experiences and use and teacher focus groups to confirm and contextualize the survey data.

Findings

Across all three years, ninth graders and students who were white, female, eligible for free/reduced lunch and those with stable enrollment were more likely to have an individual iPad (THP). Having a THP was associated with greater satisfaction, ease and frequency of technology use, higher attendance and GPA.

Research limitations/implications

Inequitable distribution, inconsistent administrative support, and uneven faculty support for iPads limit generalizability.

Practical implications

This research identifies barriers to successful technology integration and impact on student achievement including inequitable access, inconsistent administrative and teacher support, and limited professional development.

Originality/value

This research focuses on an issue of digital equity that has not yet been studied in depth with 1:1 iPads and investigates students’ technology access, experiences and use to support student achievement in a high poverty, highly diverse high school.
Innovation risk in digital business models: the German energy sector
Authors: Dellermann Dominik , Fliaster Alexander , Kolloch Michael
Published: September 2017

Purpose

Past research demonstrated that novel IT-based business models generate tremendous returns for innovators. However, the risks associated with these innovations remain under-explored. This paper aims to address this critical gap analyzing risks and offering important insights particularly for practitioners.

Design/methodology/approach

The authors adopted an exploratory multiple-case study research design. It draws on 22 semi-structured interviews with managers from leading energy utilities, as well as leading providers of virtual power plants technology within the German energy industry.

Findings

The research reveals that main risks in new digital business models in the energy sector are associated with three forms of interdependence between innovation actors: the regulatory, the technological and the collaborative. To deal with these interdependencies, the authors propose an original multi-step risk management framework. This framework considers the outreach as a critical dimension for risk assessment and offers a new risk response matrix to draw individual and collective mitigation activities for specific types of risks.

Practical implications

This paper offers a framework for the management of interdependence risks that are fundamental for business model innovations based on IT. Thus, it is applicable in companies both inside the energy sector and beyond.

Originality/value

This paper analyzes an important digital business model innovation that has not yet been explored in management literature – the virtual power plant (VPP). It is based on original and current empirical work and proposes a novel risk management framework for business organizations.
Chained to a rock
Authors: Hirsch Peter Buell
Published: September 2017

Purpose

The purpose of this paper is to examine the extraordinary reputational challenge for brands in social media in an era of heightened political and cultural polarization. In a time when the tension between liberal and conservative consumers has grown significantly, brands are being threatened with boycotts from both the left and right. In this paper, the authors identify some core approaches for brands facing this dilemma.

Design/methodology/approach

The authors’ approach is to review the experience of various brands which have been the victims of consumer boycotts motivated by political considerations. The authors examined these events from the perspective of the severity/duration of their impact on the brand, how the brands chose to respond and how their response was perceived to suggest the approaches that seem to be most effective in mitigating brand damage.

Findings

The authors have found that the critical factors in mitigating brand damage are giving clear guidelines to employees about customer relations, understanding the composition of the customer base more deeply through the lens of politics and culture, developing a comprehensive risk management approach and creating a consistent point of view on handling political threats and boycotts to ensure consistency.

Research limitations/implications

The authors’ evaluation is by definition subjective and the insights gained have not been tested empirically.

Practical implications

While the potential political threats to a specific brand are reasonably predictable, consumer perceptions are influenced by many factors only partially within the brand’s control.

Social implications

Much as companies gained expertise in managing reputation crises throughout the 1980s and 1990s, the authors believe that brands adopting their approach to politically driven boycotts will gradually learn how to manage them and the threats will become a routine part of a brand’s relationship with its stakeholders.

Originality/value

While a great deal has been written about the nature and growth of politically driven brand threats, the authors believe this paper is an original contribution to how to manage them.
Integrated learning in schools: an Icelandic case study
Authors: Pálsdóttir Kolbrún
Published: September 2017

Purpose

The purpose of this paper is to explore the integration of school and leisure activities and to inform professional practice by exploring the opportunities and challenges that arise when school-day teachers and leisure-care personnel set out to build a collaborative network.

Design/methodology/approach

This paper presents a multiple case study of five Reykjavik elementary schools that participated in a project called “The Day of the Child.” Data were gathered using semi-structured interviews and focus groups with school leaders, leisure-time center coordinators, teachers, and leisure-care personnel, as well as using field notes and documentary analysis.

Findings

The participants in the study reported that increased integration supported the well-being of children by offering leisure activities within the school day. Emphasis on leisure reinforced children’s informal and social learning. Challenges identified were organizational barriers, such as professional boundaries between leisure and school, unclear roles of leisure-care personnel and lack of active collaboration. Drawing on the findings, the author explores possibilities for enhanced professional practice within schools.

Practical implications

Leisure and informal learning are marginalized in the educational discourse due to the global push in education toward outcomes-based education and standardized testing. This study underlines the need for leisure pedagogy in elementary schools.

Originality/value

This paper provides an understanding of the importance of informal learning within the formal school culture. The findings reflect the experiences and insights of those working with children and focuses on a much neglected part of education, informal learning.
The brand meaning co-creation process on Facebook
Authors: Rosenthal Benjamin , Brito Eliane Pereira Zamith
Published: September 2017

Purpose

The purpose of this paper is to propose a framework for brand meaning co-creation between brands and fans on Facebook.

Design/methodology/approach

A case study methodology was applied. Brand posts in the form of text, pictures, videos and fan comments of three brand pages – Nike Run, Mizuno and Jack Daniel’s – were collected and analyzed in a netnographic manner. Seven influential fans of the Nike Run brand page, who were identified in the data, and one marketing manager of each brand were interviewed.

Findings

This paper shows how brand meanings are orchestrated by brand managers and co-created through a process in which the brand leads the dialog through several types of brand actions. It also shows how fans engage in this dialog through multiple forms of reactions. A brand page’s content should be curated by its manager based on the role of the content on fans’ lives and their potential reactions and not merely on the meanings that the brand desires to communicate.

Research limitations/implications

This paper proposes a conceptual framework for understanding brand meaning co-creation at the micro-level of brand-fans daily interactions. Nevertheless, this study analyzed only three brand pages in two product categories – alcoholic beverages and running. Therefore, the authors do not claim that one can extrapolate from their findings.

Practical implications

The brand meaning co-creation process that is identified here provides a useful frame of reference for brand managers who seek to understand how they can best influence fans to co-create brand meanings in directions that benefit their companies.

Originality/value

This paper evolves with the co-creation of brand meanings literature by proposing a framework of brand meaning co-creation on Facebook. This framework can help brand managers to fine tune their content strategy in social media.