Generalized empirical Bayesian methods for discovery of differential data in high-throughput biology.

MOTIVATION: High-throughput data are now commonplace in biological research. Rapidly changing technologies and application mean that novel methods for detecting differential behaviour that account for a 'large P, small n' setting are required at an increasing rate. The development of such methods is, in general, being done on an ad hoc basis, requiring further development cycles and a lack of standardization between analyses. RESULTS: We present here a generalized method for identifying differential behaviour within high-throughput biological data through empirical Bayesian methods. This approach is based on our baySeq algorithm for identification of differential expression in RNA-seq data based on a negative binomial distribution, and in paired data based on a beta-binomial distribution. Here we show how the same empirical Bayesian approach can be applied to any parametric distribution, removing the need for lengthy development of novel methods for differently distributed data. Comparisons with existing methods developed to address specific problems in high-throughput biological data show that these generic methods can achieve equivalent or better performance. A number of enhancements to the basic algorithm are also presented to increase flexibility and reduce computational costs. AVAILABILITY AND IMPLEMENTATION: The methods are implemented in the R baySeq (v2) package, available on Bioconductor http://www.bioconductor.org/packages/release/bioc/html/baySeq.html. CONTACT: [email protected] SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.This work was supported by European Research Council Advanced Investigator Grant ERC-2013-AdG 340642 – TRIBE.This is the author accepted manuscript. The final version is available from Oxford University Press via http://dx.doi.org/10.1093/bioinformatics/btv56

High-throughput sequencing of cytosine methylation in plant DNA.

: Cytosine methylation is a significant and widespread regulatory factor in plant systems. Methods for the high-throughput sequencing of methylation have allowed a greatly improved characterisation of the methylome. Here we discuss currently available methods for generation and analysis of high-throughput sequencing of methylation data. We also discuss the results previously acquired through sequencing plant methylomes, and highlight remaining challenges in this field.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

Empirical Bayesian analysis of paired high-throughput sequencing data with a beta-binomial distribution.

BACKGROUND: Pairing of samples arises naturally in many genomic experiments; for example, gene expression in tumour and normal tissue from the same patients. Methods for analysing high-throughput sequencing data from such experiments are required to identify differential expression, both within paired samples and between pairs under different experimental conditions. RESULTS: We develop an empirical Bayesian method based on the beta-binomial distribution to model paired data from high-throughput sequencing experiments. We examine the performance of this method on simulated and real data in a variety of scenarios. Our methods are implemented as part of the RbaySeq package (versions 1.11.6 and greater) available from Bioconductor (http://www.bioconductor.org). CONCLUSIONS: We compare our approach to alternatives based on generalised linear modelling approaches and show that our method offers significant gains in performance on simulated data. In testing on real data from oral squamous cell carcinoma patients, we discover greater enrichment of previously identified head and neck squamous cell carcinoma associated gene sets than has previously been achieved through a generalised linear modelling approach, suggesting that similar gains in performance may be found in real data. Our methods thus show real and substantial improvements in analyses of high-throughput sequencing data from paired samples.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

baySeq: empirical Bayesian methods for identifying differential expression in sequence count data.

BACKGROUND: High throughput sequencing has become an important technology for studying expression levels in many types of genomic, and particularly transcriptomic, data. One key way of analysing such data is to look for elements of the data which display particular patterns of differential expression in order to take these forward for further analysis and validation. RESULTS: We propose a framework for defining patterns of differential expression and develop a novel algorithm, baySeq, which uses an empirical Bayes approach to detect these patterns of differential expression within a set of sequencing samples. The method assumes a negative binomial distribution for the data and derives an empirically determined prior distribution from the entire dataset. We examine the performance of the method on real and simulated data. CONCLUSIONS: Our method performs at least as well, and often better, than existing methods for analyses of pairwise differential expression in both real and simulated data. When we compare methods for the analysis of data from experimental designs involving multiple sample groups, our method again shows substantial gains in performance. We believe that this approach thus represents an important step forward for the analysis of count data from sequencing experiments.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

Extreme Ultraviolet Emission in the Fornax Cluster of Galaxies

We present studies of the Extreme Ultraviolet (EUV) emission in the Fornax cluster of galaxies; a relatively nearby well-studied cluster with X-ray emitting cluster gas and a very large radio source. We examine both the large-scale (~size of the X-ray emitting cluster gas), and the small-scale (<arcmin) emission. We find that this cluster has large-scale diffuse EUV emission. However, at the sensitivity level of the existing EUVE data, this emission is due entirely to the low energy tail of the X-ray emitting gas. We have also examined small-scale structures in raw EUVE images of this cluster. We find that small-scale irregularities are present in all raw Deep Survey images as a result of small-scale detector effects. These effects can be removed by appropriate flat-fielding. After flat-fielding, the Fornax cluster still shows a few significant regions of small-scale EUV enhancement. We find that these are emission from stars and galaxies in the field. We find that at existing levels of sensitivity, there is no excess EUV emission in the cluster on either large or small scales.Comment: 6 pages, 3 eps figures, aastex5, Accepted to ApJ

Getting a Full Dose? Reconsidering Sex Chromosome Dosage Compensation in the Silkworm, Bombyx mori

Dosage compensation—equalizing gene expression levels in response to differences in gene dose or copy number—is classically considered to play a critical role in the evolution of heteromorphic sex chromosomes. As the X and Y diverge through degradation and gene loss on the Y (or the W in female-heterogametic ZW taxa), it is expected that dosage compensation will evolve to correct for sex-specific differences in gene dose. Although this is observed in some organisms, recent genome-wide expression studies in other taxa have revealed striking exceptions. In particular, reports that both birds and the silkworm moth (Bombyx mori) lack dosage compensation have spurred speculation that this is the rule for all female-heterogametic taxa. Here, we revisit the issue of dosage compensation in silkworm by replicating and extending the previous analysis. Contrary to previous reports, our efforts reveal a pattern typically associated with dosage compensated taxa: the global male:female expression ratio does not differ between the Z and autosomes. We believe the previous report of unequal male:female ratios on the Z reflects artifacts of microarray normalization in conjunction with not testing a major assumption that the male:female global expression ratio was unbiased for autosomal loci. However, we also find that the global Z chromosome expression is significantly reduced relative to autosomes, a pattern not expected in dosage compensated taxa. This combination of male:female parity with an overall reduction in expression for sex-linked loci is not consistent with the prevailing evolutionary theory of sex chromosome evolution and dosage compensation

Sex Chromosome Dosage Compensation in Heliconius Butterflies: Global yet Still Incomplete?

The evolution of heterogametic sex chromosomes is often-but not always-accompanied by the evolution of dosage compensating mechanisms that mitigate the impact of sex-specific gene dosage on levels of gene expression. One emerging view of this process is that such mechanisms may only evolve in male-heterogametic (XY) species but not in female-heterogametic (ZW) species, which will consequently exhibit "incomplete" sex chromosome dosage compensation. However, recent results suggest that at least some Lepidoptera (moths and butterflies) may prove to be an exception to this prediction. Studies in bombycoid moths indicate the presence of a chromosome-wide epigenetic mechanism that effectively balances Z chromosome gene expression between the sexes by reducing Z-linked expression in males. In contrast, strong sex chromosome dosage effects without any reduction in male Z-linked expression were previously reported in a pyralid moth, suggesting a lack of any such dosage compensating mechanism. Here we report an analysis of sex chromosome dosage compensation in Heliconius butterflies, sampling multiple individuals for several different adult tissues (head, abdomen, leg, mouth, and antennae). Methodologically, we introduce a novel application of linear mixed-effects models to assess dosage compensation, offering a unified statistical framework that can estimate effects specific to chromosome, to sex, and their interactions (i.e., a dosage effect). Our results show substantially reduced Z-linked expression relative to autosomes in both sexes, as previously observed in bombycoid moths. This observation is consistent with an increasing body of evidence that some lepidopteran species possess an epigenetic dosage compensating mechanism that reduces Z chromosome expression in males to levels comparable with females. However, this mechanism appears to be imperfect in Heliconius, resulting in a modest dosage effect that produces an average 5-20% increase in male expression relative to females on the Z chromosome, depending on the tissue. Thus our results in Heliconius reflect a mixture of previous patterns reported for Lepidoptera. In Heliconius, a moderate pattern of incomplete dosage compensation persists apparently despite the presence of an epigenetic dosage compensating mechanism. The chromosomal distributions of sex-biased genes show an excess of male-biased and a dearth of female-biased genes on the Z chromosome relative to autosomes, consistent with predictions of sexually antagonistic evolution.This research was supported in part by a NSF postdoctoral fellowshiptoJ.R.W. (DBI-0905698). RNA sequencing was funded by the “Capacity and Capability Challenge Program” from The Genome Analysis Centre, Norwich, UK. The computing for this project was performed on the Community Cluster at the Center for Research Computing at the University of Kansas. Luiqi (Aloy) Gu provided valuable comments on the manuscript.This is the final version of the article. It first appeared from Oxford University Press via http://dx.doi.org/10.1093/gbe/evv156

Towards annotating the plant epigenome: the Arabidopsis thaliana small RNA locus map.

Based on 98 public and internal small RNA high throughput sequencing libraries, we mapped small RNAs to the genome of the model organism Arabidopsis thaliana and defined loci based on their expression using an empirical Bayesian approach. The resulting loci were subsequently classified based on their genetic and epigenetic context as well as their expression properties. We present the results of this classification, which broadly conforms to previously reported divisions between transcriptional and post-transcriptional gene silencing small RNAs, and to PolIV and PolV dependencies. However, we are able to demonstrate the existence of further subdivisions in the small RNA population of functional significance. Moreover, we present a framework for similar analyses of small RNA populations in all species

Sex Chromosome Dosage Compensation in Heliconius Butterflies: Global yet Still Incomplete?

The evolution of heterogametic sex chromosomes is often—but not always—accompanied by the evolution of dosage compensating mechanisms that mitigate the impact of sex-specific gene dosage on levels of gene expression. One emerging view of this process is that such mechanisms may only evolve in male-heterogametic (XY) species but not in female-heterogametic (ZW) species, which will consequently exhibit “incomplete” sex chromosome dosage compensation. However, recent results suggest that at least some Lepidoptera (moths and butterflies) may prove to be an exception to this prediction. Studies in bombycoid moths indicate the presence of a chromosome-wide epigenetic mechanism that effectively balances Z chromosome gene expression between the sexes by reducing Z-linked expression in males. In contrast, strong sex chromosome dosage effects without any reduction in male Z-linked expression were previously reported in a pyralid moth, suggesting a lack of any such dosage compensating mechanism. Here we report an analysis of sex chromosome dosage compensation in Heliconius butterflies, sampling multiple individuals for several different adult tissues (head, abdomen, leg, mouth, and antennae). Methodologically, we introduce a novel application of linear mixed-effects models to assess dosage compensation, offering a unified statistical framework that can estimate effects specific to chromosome, to sex, and their interactions (i.e., a dosage effect). Our results show substantially reduced Z-linked expression relative to autosomes in both sexes, as previously observed in bombycoid moths. This observation is consistent with an increasing body of evidence that some lepidopteran species possess an epigenetic dosage compensating mechanism that reduces Z chromosome expression in males to levels comparable with females. However, this mechanism appears to be imperfect in Heliconius, resulting in a modest dosage effect that produces an average 5–20% increase in male expression relative to females on the Z chromosome, depending on the tissue. Thus our results in Heliconius reflect a mixture of previous patterns reported for Lepidoptera. In Heliconius, a moderate pattern of incomplete dosage compensation persists apparently despite the presence of an epigenetic dosage compensating mechanism. The chromosomal distributions of sex-biased genes show an excess of male-biased and a dearth of female-biased genes on the Z chromosome relative to autosomes, consistent with predictions of sexually antagonistic evolution