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1.  Dietary patterns and changes in body composition in children between 9 and 11 years 
Food & Nutrition Research  2014;58:10.3402/fnr.v58.22769.
Objective
Childhood obesity is rising and dietary intake is a potentially modifiable factor that plays an important role in its development. We aim to investigate the association between dietary patterns, obtained through principal components analysis and gains in fat and lean mass in childhood.
Design
Diet diaries at 10 years of age collected from children taking part in the Avon Longitudinal Study of Parents and Children. Body composition was assessed using dual-energy X-ray absorptiometry at 9 and 11.
Setting
Longitudinal birth cohort.
Subjects
3911 children with complete data.
Results
There was an association between the Health Aware (positive loadings on high-fiber bread, and fruits and vegetables; negative loadings on chips, crisps, processed meat, and soft drinks) pattern score and decreased fat mass gain in girls. After adjusting for confounders, an increase of 1 standard deviation (sd) in this score led to an estimated 1.2% decrease in fat mass gain in valid-reporters and 2.1% in under-reporters. A similar decrease was found only in under-reporting boys. There was also an association between the Packed Lunch (high consumption of white bread, sandwich fillings, and snacks) pattern score and decreased fat mass gain (1.1% per sd) in valid-reporting but not under-reporting girls. The main association with lean mass gain was an increase with Packed Lunch pattern score in valid-reporting boys only.
Conclusions
There is a small association between dietary patterns and change in fat mass in mid-childhood. Differences between under- and valid-reporters emphasize the need to consider valid-reporters separately in such studies.
doi:10.3402/fnr.v58.22769
PMCID: PMC4090365  PMID: 25018688
dietary patterns; principal components analysis; ALSPAC; body composition; valid-reporters
2.  Leveraging cross-link modification events in CLIP-seq for motif discovery 
Nucleic Acids Research  2014;43(1):95-103.
High-throughput protein–RNA interaction data generated by CLIP-seq has provided an unprecedented depth of access to the activities of RNA-binding proteins (RBPs), the key players in co- and post-transcriptional regulation of gene expression. Motif discovery forms part of the necessary follow-up data analysis for CLIP-seq, both to refine the exact locations of RBP binding sites, and to characterize them. The specific properties of RBP binding sites, and the CLIP-seq methods, provide additional information not usually present in the classic motif discovery problem: the binding site structure, and cross-linking induced events in reads. We show that CLIP-seq data contains clear secondary structure signals, as well as technology- and RBP-specific cross-link signals. We introduce Zagros, a motif discovery algorithm specifically designed to leverage this information and explore its impact on the quality of recovered motifs. Our results indicate that using both secondary structure and cross-link modifications can greatly improve motif discovery on CLIP-seq data. Further, the motifs we recover provide insight into the balance between sequence- and structure-specificity struck by RBP binding.
doi:10.1093/nar/gku1288
PMCID: PMC4288180  PMID: 25505146
3.  Preferred Methods of Sexually Transmitted Infection Service Delivery Among an Urban Sample of Underserved Midwestern Men 
Sexually transmitted diseases  2014;41(2):129-132.
Semistructured interviews were completed with a sample of 25 men residing in an urban area of the Midwestern United States to elicit preferred methods of sexually transmitted infection service delivery. Results highlight the influence of stigma, social support, and perceived risk on sexually transmitted infection screening uptake and preferred methods of screening.
doi:10.1097/OLQ.0000000000000082
PMCID: PMC4148079  PMID: 24413494
4.  Using beta-binomial regression for high-precision differential methylation analysis in multifactor whole-genome bisulfite sequencing experiments 
BMC Bioinformatics  2014;15:215.
Background
Whole-genome bisulfite sequencing currently provides the highest-precision view of the epigenome, with quantitative information about populations of cells down to single nucleotide resolution. Several studies have demonstrated the value of this precision: meaningful features that correlate strongly with biological functions can be found associated with only a few CpG sites. Understanding the role of DNA methylation, and more broadly the role of DNA accessibility, requires that methylation differences between populations of cells are identified with extreme precision and in complex experimental designs.
Results
In this work we investigated the use of beta-binomial regression as a general approach for modeling whole-genome bisulfite data to identify differentially methylated sites and genomic intervals.
Conclusions
The regression-based analysis can handle medium- and large-scale experiments where it becomes critical to accurately model variation in methylation levels between replicates and account for influence of various experimental factors like cell types or batch effects.
doi:10.1186/1471-2105-15-215
PMCID: PMC4230021  PMID: 24962134
Epigenomics; Differential methylation; Beta-binomial regression
5.  Intergenerational genomic DNA methylation patterns in mouse hybrid strains 
Genome Biology  2014;15(5):R68.
Background
DNA methylation is a contributing factor to both rare and common human diseases, and plays a major role in development and gene silencing. While the variation of DNA methylation among individuals has been partially characterized, the degree to which methylation patterns are preserved across generations is still poorly understood. To determine the extent of methylation differences between two generations of mice we examined DNA methylation patterns in the livers of eight parental and F1 mice from C57BL/6J and DBA/2J mouse strains using bisulfite sequencing.
Results
We find a large proportion of reproducible methylation differences between C57BL/6J and DBA/2J chromosomes in CpGs, which are highly heritable between parent and F1 mice. We also find sex differences in methylation levels in 396 genes, and 11% of these are differentially expressed between females and males. Using a recently developed approach to identify allelically methylated regions independently of genotypic differences, we identify 112 novel putative imprinted genes and microRNAs, and validate imprinting at the RNA level in 10 of these genes.
Conclusions
The majority of DNA methylation differences among individuals are associated with genetic differences, and a much smaller proportion of these epigenetic differences are due to sex, imprinting or stochastic intergenerational effects. Epigenetic differences can be a determining factor in heritable traits and should be considered in association studies for molecular and clinical traits, as we observed that methylation differences in the mouse model are highly heritable and can have functional consequences on molecular traits such as gene expression.
doi:10.1186/gb-2014-15-5-r68
PMCID: PMC4076608  PMID: 24887417
6.  Metastatic Melanoma: Lactate Dehydrogenase Levels and CT Imaging Findings of Tumor Devascularization Allow Accurate Prediction of Survival in Patients Treated with Bevacizumab1 
Radiology  2013;270(2):425-434.
Purpose
To predict survival in patients with metastatic melanoma by evaluating a combination of serum lactate dehydrogenase (LDH) level and initial computed tomographic (CT) findings of tumor devascularization after antiangiogenic therapy.
Materials and Methods
Consent was waived for this institutional review board–approved, retrospective, secondary analysis. Forty-four patients with metastatic melanoma received bevacizumab therapy in a randomized prospective phase II trial. Target lesions on the initial posttherapy CT images were evaluated by using Response Evaluation Criteria in Solid Tumors, the Choi criteria, and Morphology, Attenuation, Size, and Structure (MASS) criteria. Cox proportional hazards models were used to assess the association of baseline clinical variables including serum LDH and imaging findings with progression-free and overall survival. The receiver operating characteristic curve with area under the curve (AUC) was used to evaluate accuracy.
Results
In multivariate analysis, a high baseline serum LDH level was associated with decreased progression-free survival (hazard ratio = 1.29 for each increase of 100 IU/L; P = .002) and overall survival (hazard ratio = 1.44 for each increase of 100 IU/L; P = .001). Evaluation with MASS criteria of the first CT examination after therapy strongly predicted progression-free (P < .001) and overall (P < .001) survival. Baseline serum LDH level was moderately accurate for predicting progression-free survival at 9 months (AUC = 0.793) and overall survival at 18 months (AUC = 0.689). The combination of baseline serum LDH levels and evaluation with MASS criteria at the first CT examination after therapy had significantly higher accuracy for predicting progression-free survival at 9 months (AUC = 0.969) and overall survival at 18 months (AUC = 0.813) than did baseline serum LDH levels alone for prediction of progression-free survival (P = .020).
Conclusion
A combination of baseline serum LDH levels and evaluation with MASS criteria at the first CT examination after bevacizumab therapy had the highest accuracy for predicting survival in patients with metastatic melanoma.
doi:10.1148/radiol.13130776
PMCID: PMC3985552  PMID: 24072776
7.  The Effects of Mary Rose Conservation Treatment on Iron Oxidation Processes and Microbial Communities Contributing to Acid Production in Marine Archaeological Timbers 
PLoS ONE  2014;9(2):e84169.
The Tudor warship the Mary Rose has reached an important transition point in her conservation. The 19 year long process of spraying with polyethylene glycol (PEG) has been completed (April 29th 2013) and the hull is air drying under tightly controlled conditions. Acidophilic bacteria capable of oxidising iron and sulfur have been previously identified and enriched from unpreserved timbers of the Mary Rose, demonstrating that biological pathways of iron and sulfur oxidization existed potentially in this wood, before preservation with PEG. This study was designed to establish if the recycled PEG spray system was a reservoir of microorganisms capable of iron and sulfur oxidization during preservation of the Mary Rose. Microbial enrichments derived from PEG impregnated biofilm collected from underneath the Mary Rose hull, were examined to better understand the processes of cycling of iron. X-ray absorption spectroscopy was utilised to demonstrate the biological contribution to production of sulfuric acid in the wood. Using molecular microbiological techniques to examine these enrichment cultures, PEG was found to mediate a shift in the microbial community from a co-culture of Stenotrophomonas and Brevunidimonas sp, to a co-culture of Stenotrophomonas and the iron oxidising Alicyclobacillus sp. Evidence is presented that PEG is not an inert substance in relation to the redox cycling of iron. This is the first demonstration that solutions of PEG used in the conservation of the Mary Rose are promoting the oxidation of ferrous iron in acidic solutions, in which spontaneous abiotic oxidation does not occur in water. Critically, these results suggest PEG mediated redox cycling of iron between valence states in solutions of 75% PEG 200 and 50% PEG 2000 (v/v) at pH 3.0, with serious implications for the future use of PEG as a conservation material of iron rich wooden archaeological artefacts.
doi:10.1371/journal.pone.0084169
PMCID: PMC3929279  PMID: 24586230
8.  On the Value of Intra-Motif Dependencies of Human Insulator Protein CTCF 
PLoS ONE  2014;9(1):e85629.
The binding affinity of DNA-binding proteins such as transcription factors is mainly determined by the base composition of the corresponding binding site on the DNA strand. Most proteins do not bind only a single sequence, but rather a set of sequences, which may be modeled by a sequence motif. Algorithms for de novo motif discovery differ in their promoter models, learning approaches, and other aspects, but typically use the statistically simple position weight matrix model for the motif, which assumes statistical independence among all nucleotides. However, there is no clear justification for that assumption, leading to an ongoing debate about the importance of modeling dependencies between nucleotides within binding sites. In the past, modeling statistical dependencies within binding sites has been hampered by the problem of limited data. With the rise of high-throughput technologies such as ChIP-seq, this situation has now changed, making it possible to make use of statistical dependencies effectively. In this work, we investigate the presence of statistical dependencies in binding sites of the human enhancer-blocking insulator protein CTCF by using the recently developed model class of inhomogeneous parsimonious Markov models, which is capable of modeling complex dependencies while avoiding overfitting. These findings lead to a more detailed characterization of the CTCF binding motif, which is only poorly represented by independent nucleotide frequencies at several positions, predominantly at the 3′ end.
doi:10.1371/journal.pone.0085629
PMCID: PMC3899044  PMID: 24465627
9.  Genomic Analyses Reveal Broad Impact of miR-137 on Genes Associated with Malignant Transformation and Neuronal Differentiation in Glioblastoma Cells 
PLoS ONE  2014;9(1):e85591.
miR-137 plays critical roles in the nervous system and tumor development; an increase in its expression is required for neuronal differentiation while its reduction is implicated in gliomagenesis. To evaluate the potential of miR-137 in glioblastoma therapy, we conducted genome-wide target mapping in glioblastoma cells by measuring the level of association between PABP and mRNAs in cells transfected with miR-137 mimics vs. controls via RIPSeq. Impact on mRNA levels was also measured by RNASeq. By combining the results of both experimental approaches, 1468 genes were found to be negatively impacted by miR-137 – among them, 595 (40%) contain miR-137 predicted sites. The most relevant targets include oncogenic proteins and key players in neurogenesis like c-KIT, YBX1, AKT2, CDC42, CDK6 and TGFβ2. Interestingly, we observed that several identified miR-137 targets are also predicted to be regulated by miR-124, miR-128 and miR-7, which are equally implicated in neuronal differentiation and gliomagenesis. We suggest that the concomitant increase of these four miRNAs in neuronal stem cells or their repression in tumor cells could produce a robust regulatory effect with major consequences to neuronal differentiation and tumorigenesis.
doi:10.1371/journal.pone.0085591
PMCID: PMC3899048  PMID: 24465609
10.  Coping with continuous human disturbance in the wild: insights from penguin heart rate response to various stressors 
BMC Ecology  2012;12:10.
Background
A central question for ecologists is the extent to which anthropogenic disturbances (e.g. tourism) might impact wildlife and affect the systems under study. From a research perspective, identifying the effects of human disturbance caused by research-related activities is crucial in order to understand and account for potential biases and derive appropriate conclusions from the data.
Results
Here, we document a case of biological adjustment to chronic human disturbance in a colonial seabird, the king penguin (Aptenodytes patagonicus), breeding on remote and protected islands of the Southern ocean. Using heart rate (HR) as a measure of the stress response, we show that, in a colony with areas exposed to the continuous presence of humans (including scientists) for over 50 years, penguins have adjusted to human disturbance and habituated to certain, but not all, types of stressors. When compared to birds breeding in relatively undisturbed areas, birds in areas of high chronic human disturbance were found to exhibit attenuated HR responses to acute anthropogenic stressors of low-intensity (i.e. sounds or human approaches) to which they had been subjected intensely over the years. However, such attenuation was not apparent for high-intensity stressors (i.e. captures for scientific research) which only a few individuals experience each year.
Conclusions
Habituation to anthropogenic sounds/approaches could be an adaptation to deal with chronic innocuous stressors, and beneficial from a research perspective. Alternately, whether penguins have actually habituated to anthropogenic disturbances over time or whether human presence has driven the directional selection of human-tolerant phenotypes, remains an open question with profound ecological and conservation implications, and emphasizes the need for more knowledge on the effects of human disturbance on long-term studied populations.
doi:10.1186/1472-6785-12-10
PMCID: PMC3543187  PMID: 22784366
Stress; Heart rate; Habituation; Selection; Seabird; Human disturbance; Long-term monitoring
11.  Increased plasma levels of soluble vascular endothelial growth factor (VEGF) receptor 1 (sFlt-1) in women by moderate exercise and increased plasma levels of VEGF in overweight/obese women 
The incidence of breast cancer is increasing worldwide, and this seems to be related to an increase in lifestyle risk factors, including physical inactivity, and overweight/obesity. We previously reported that exercise induced a circulating angiostatic phenotype characterized by increased sFlt-1 and endostatin and decreased unbound-VEGF in men. However, there is no data on women. The present study determines the following: 1) whether moderate exercise increased sFlt-1 and endostatin and decreased unbound-VEGF in the circulation of adult female volunteers; 2) whether overweight/obese women have a higher plasma level of unbound-VEGF than lean women. 72 African American and Caucasian adult women volunteers aged from 18–44 were enrolled into the exercise study. All the participants walked on a treadmill for 30 minutes at a moderate intensity (55–59% heart rate reserve), and oxygen consumption (VO2) was quantified by utilizing a metabolic cart. We had the blood samples before and immediately after exercise from 63 participants. ELISA assays (R&D Systems) showed that plasma levels of sFlt-1 were 67.8±3.7 pg/ml immediately after exercise (30 minutes), significantly higher than basal levels, 54.5±3.3 pg/ml, before exercise (P < 0.01; n=63). There was no significant difference in the % increase of sFlt-1 levels after exercise between African American and Caucasian (P=0.533) or between lean and overweight/obese women (P=0.892). There was no significant difference in plasma levels of unbound VEGF (35.28±5.47 vs. 35.23±4.96 pg/ml; P=0.99) or endostatin (111.12±5.48 vs. 115.45±7.15 ng/ml; P=0.63) before and after exercise. Basal plasma levels of unbound-VEGF in overweight/obese women were 52.26±9.6 pg/ml, significantly higher than basal levels of unbound-VEGF in lean women, 27.34±4.99 pg/ml (P < 0.05). The results support our hypothesis that exercise-induced plasma levels of sFlt-1 could be an important clinical biomarker to explore the mechanisms of exercise training in reducing breast cancer progression and that VEGF is an important biomarker in obesity and obesity-related cancer progression.
doi:10.1097/CEJ.0b013e328353ed81
PMCID: PMC3449013  PMID: 22609636
Exercise; Young adult women; Overweight/obese; sFlt-1; Endostatin; VEGF
12.  Regionally Specific and Genome-Wide Analyses Conclusively Demonstrate the Absence of CpG Methylation in Human Mitochondrial DNA 
Molecular and Cellular Biology  2013;33(14):2683-2690.
Although CpG methylation clearly distributes genome-wide in vertebrate nuclear DNA, the state of methylation in the vertebrate mitochondrial genome has been unclear. Several recent reports using immunoprecipitation, mass spectrometry, and enzyme-linked immunosorbent assay methods concluded that human mitochondrial DNA (mtDNA) has much more than the 2 to 5% CpG methylation previously estimated. However, these methods do not provide information as to the sites or frequency of methylation at each CpG site. Here, we have used the more definitive bisulfite genomic sequencing method to examine CpG methylation in HCT116 human cells and primary human cells to independently answer these two questions. We found no evidence of CpG methylation at a biologically significant level in these regions of the human mitochondrial genome. Furthermore, unbiased next-generation sequencing of sodium bisulfite treated total DNA from HCT116 cells and analysis of genome-wide sodium bisulfite sequencing data sets from several other DNA sources confirmed this absence of CpG methylation in mtDNA. Based on our findings using regionally specific and genome-wide approaches with multiple human cell sources, we can definitively conclude that CpG methylation is absent in mtDNA. It is highly unlikely that CpG methylation plays any role in direct control of mitochondrial function.
doi:10.1128/MCB.00220-13
PMCID: PMC3700126  PMID: 23671186
13.  A profile of transcriptomic changes in the rd10 mouse model of retinitis pigmentosa 
Molecular Vision  2014;20:1612-1628.
Purpose
Retinitis pigmentosa (RP) is a photoreceptor disease that affects approximately 100,000 people in the United States. Treatment options are limited, and the prognosis for most patients is progressive vision loss. Unfortunately, understanding of the molecular underpinnings of RP initiation and progression is still limited. However, the development of animal models of RP, coupled with high-throughput sequencing, has provided an opportunity to study the underlying cellular and molecular changes in this disease.
Methods
Using RNA-Seq, we present the first retinal transcriptome analysis of the rd10 murine model of retinal degeneration.
Results
Our data confirm the loss of rod-specific transcripts and the increased relative expression of Müller-specific transcripts, emphasizing the important role of reactive gliosis and innate immune activation in RP. Moreover, we report substantial changes in relative isoform usage among neuronal differentiation and morphogenesis genes, including a marked shift to shorter transcripts.
Conclusions
Our analyses implicate remodeling of the inner retina and possible Müller cell dedifferentiation.
PMCID: PMC4235044  PMID: 25489233
14.  A Reference Methylome Database and Analysis Pipeline to Facilitate Integrative and Comparative Epigenomics 
PLoS ONE  2013;8(12):e81148.
DNA methylation is implicated in a surprising diversity of regulatory, evolutionary processes and diseases in eukaryotes. The introduction of whole-genome bisulfite sequencing has enabled the study of DNA methylation at a single-base resolution, revealing many new aspects of DNA methylation and highlighting the usefulness of methylome data in understanding a variety of genomic phenomena. As the number of publicly available whole-genome bisulfite sequencing studies reaches into the hundreds, reliable and convenient tools for comparing and analyzing methylomes become increasingly important. We present MethPipe, a pipeline for both low and high-level methylome analysis, and MethBase, an accompanying database of annotated methylomes from the public domain. Together these resources enable researchers to extract interesting features from methylomes and compare them with those identified in public methylomes in our database.
doi:10.1371/journal.pone.0081148
PMCID: PMC3855694  PMID: 24324667
15.  scAAV-Mediated Gene Transfer of Interleukin 1-Receptor Antagonist to Synovium and Articular Cartilage in Large Mammalian Joints 
Gene therapy  2012;20(6):670-677.
With the long-term goal of developing a gene-based treatment for osteoarthritis (OA), we performed studies to evaluate the equine joint as a model for AAV-mediated gene transfer to large, weight-bearing human joints. A self-complementary AAV2 vector containing the coding regions for human interleukin-1 receptor antagonist (hIL-1Ra) or green fluorescent protein (GFP) was packaged in AAV capsid serotypes 1, 2, 5, 8 and 9. Following infection of human and equine synovial fibroblasts in culture, we found that both were only receptive to transduction with AAV1, 2 and 5. For these serotypes, however, transgene expression from the equine cells was consistently at least 10-fold higher. Analyses of AAV surface receptor molecules and intracellular trafficking of vector genomes implicate enhanced viral uptake by the equine cells. Following delivery of 1 × 1011 vector genomes of serotypes 2, 5 and 8 into the forelimb joints of the horse, all three enabled hIL-1Ra expression at biologically relevant levels and effectively transduced the same cell types, primarily synovial fibroblasts and, to a lesser degree, chondrocytes in articular cartilage. These results provide optimism that AAV vectors can be effectively adapted for gene delivery to large human joints affected by OA.
doi:10.1038/gt.2012.81
PMCID: PMC3577988  PMID: 23151520
Osteoarthritis; Self-complementary Adeno-Associated Virus; Interleukin-1 Receptor Antagonist; Synovium; Cartilage; Equine
16.  Site identification in high-throughput RNA–protein interaction data 
Bioinformatics  2012;28(23):3013-3020.
Motivation: Post-transcriptional and co-transcriptional regulation is a crucial link between genotype and phenotype. The central players are the RNA-binding proteins, and experimental technologies [such as cross-linking with immunoprecipitation- (CLIP-) and RIP-seq] for probing their activities have advanced rapidly over the course of the past decade. Statistically robust, flexible computational methods for binding site identification from high-throughput immunoprecipitation assays are largely lacking however.
Results: We introduce a method for site identification which provides four key advantages over previous methods: (i) it can be applied on all variations of CLIP and RIP-seq technologies, (ii) it accurately models the underlying read-count distributions, (iii) it allows external covariates, such as transcript abundance (which we demonstrate is highly correlated with read count) to inform the site identification process and (iv) it allows for direct comparison of site usage across cell types or conditions.
Availability and implementation: We have implemented our method in a software tool called Piranha. Source code and binaries, licensed under the GNU General Public License (version 3) are freely available for download from http://smithlab.usc.edu.
Contact: andrewds@usc.edu
Supplementary information: Supplementary data available at Bioinformatics online.
doi:10.1093/bioinformatics/bts569
PMCID: PMC3509493  PMID: 23024010
17.  Predicting the molecular complexity of sequencing libraries 
Nature methods  2013;10(4):325-327.
Predicting the molecular complexity of a genomic sequencing library has emerged as a critical but difficult problem in modern applications of genome sequencing. Available methods to determine either how deeply to sequence, or predict the benefits of additional sequencing, are almost completely lacking. We introduce an empirical Bayesian method to implicitly model any source of bias and accurately characterize the molecular complexity of a DNA sample or library in almost any sequencing application.
doi:10.1038/nmeth.2375
PMCID: PMC3612374  PMID: 23435259
18.  MLML: consistent simultaneous estimates of DNA methylation and hydroxymethylation 
Bioinformatics  2013;29(20):2645-2646.
Motivation: The two major epigenetic modifications of cytosines, 5-methylcytosine (5-mC) and 5-hydroxymethylcytosine (5-hmC), coexist with each other in a range of mammalian cell populations. Increasing evidence points to important roles of 5-hmC in demethylation of 5-mC and epigenomic regulation in development. Recently developed experimental methods allow direct single-base profiling of either 5-hmC or 5-mC. Meaningful analyses seem to require combining these experiments with bisulfite sequencing, but doing so naively produces inconsistent estimates of 5-mC or 5-hmC levels.
Results: We present a method to jointly model read counts from bisulfite sequencing, oxidative bisulfite sequencing and Tet-Assisted Bisulfite sequencing, providing simultaneous estimates of 5-hmC and 5-mC levels that are consistent across experiment types.
Availability: http://smithlab.usc.edu/software/mlml
Contact: andrewds@usc.edu
Supplementary information: Supplementary material is available at Bioinformatics online.
doi:10.1093/bioinformatics/btt459
PMCID: PMC3789553  PMID: 23969133
19.  A Geometric Interpretation for Local Alignment-Free Sequence Comparison 
Journal of Computational Biology  2013;20(7):471-485.
Abstract
Local alignment-free sequence comparison arises in the context of identifying similar segments of sequences that may not be alignable in the traditional sense. We propose a randomized approximation algorithm that is both accurate and efficient. We show that under D2 and its important variant \documentclass{aastex}\usepackage{amsbsy}\usepackage{amsfonts}\usepackage{amssymb}\usepackage{bm}\usepackage{mathrsfs}\usepackage{pifont}\usepackage{stmaryrd}\usepackage{textcomp}\usepackage{portland, xspace}\usepackage{amsmath, amsxtra}\pagestyle{empty}\DeclareMathSizes{10}{9}{7}{6}\begin{document} $$D_2^*$$ \end{document} as the similarity measure, local alignment-free comparison between a pair of sequences can be formulated as the problem of finding the maximum bichromatic dot product between two sets of points in high dimensions. We introduce a geometric framework that reduces this problem to that of finding the bichromatic closest pair (BCP), allowing the properties of the underlying metric to be leveraged. Local alignment-free sequence comparison can be solved by making a quadratic number of alignment-free substring comparisons. We show both theoretically and through empirical results on simulated data that our approximation algorithm requires a subquadratic number of such comparisons and trades only a small amount of accuracy to achieve this efficiency. Therefore, our algorithm can extend the current usage of alignment-free–based methods and can also be regarded as a substitute for local alignment algorithms in many biological studies.
doi:10.1089/cmb.2012.0280
PMCID: PMC3704055  PMID: 23829649
algorithms; alignment; dynamic programming; metagenomics
20.  DNA methylation dynamics during intestinal stem cell differentiation reveals enhancers driving gene expression in the villus 
Genome Biology  2013;14(5):R50.
Background
DNA methylation is of pivotal importance during development. Previous genome-wide studies identified numerous differentially methylated regions upon differentiation of stem cells, many of them associated with transcriptional start sites.
Results
We present the first genome-wide, single-base-resolution view into DNA methylation dynamics during differentiation of a mammalian epithelial stem cell: the mouse small intestinal Lgr5+ stem cell. Very little change was observed at transcriptional start sites and our data suggest that differentiation-related genes are already primed for expression in the stem cell. Genome-wide, only 50 differentially methylated regions were identified. Almost all of these loci represent enhancers driving gene expression in the differentiated part of the small intestine. Finally, we show that binding of the transcription factor Tcf4 correlates with hypo-methylation and demonstrate that Tcf4 is one of the factors contributing to formation of differentially methylated regions.
Conclusions
Our results reveal limited DNA methylation dynamics during small intestine stem cell differentiation and an impact of transcription factor binding on shaping the DNA methylation landscape during differentiation of stem cells in vivo.
doi:10.1186/gb-2013-14-5-r50
PMCID: PMC4053812  PMID: 23714178
Adult stem cells; Differentiation; DNA Methylation; Methylome; Enhancer; Tcf4
21.  Transcription Start Site Evolution in Drosophila 
Molecular Biology and Evolution  2013;30(8):1966-1974.
Transcription start site (TSS) evolution remains largely undescribed in Drosophila, likely due to limited annotations in non-melanogaster species. In this study, we introduce a concise new method that selectively sequences from the 5′-end of mRNA and used it to identify TSS in four Drosophila species, including Drosophila melanogaster, D. simulans, D. sechellia, and D. pseudoobscura. For verification, we compared our results in D. melanogaster with known annotations, published 5′-rapid amplification of cDNA ends data, and with RNAseq from the same mRNA pool. Then, we paired 2,849 D. melanogaster TSS with its closest equivalent TSS in each species (likely to be its true ortholog) using the available multiple sequence alignments. Most of the D. melanogaster TSSs were successfully paired with an ortholog in each species (83%, 86%, and 55% for D. simulans, D. sechellia, and D. pseudoobscura, respectively). On the basis of the number and distribution of reads mapped at each TSS, we also estimated promoter-specific expression (PSE) and TSS peak shape, respectively. Among paired TSS orthologs, the location and promoter activity were largely conserved. TSS location appears important as PSE, and TSS peak shape was more frequently divergent among TSS that had moved. Unpaired TSS were surprisingly common in D. pseudoobscura. An increased mutation rate upstream of TSS might explain this pattern. We found an enrichment of ribosomal protein genes among diverged TSS, suggesting that TSS evolution is not uniform across the genome.
doi:10.1093/molbev/mst085
PMCID: PMC3708499  PMID: 23649539
promoter; transcription start site; gene expression; CAGE; TSS; Drosophila
22.  Lebrikizumab in the personalized management of asthma 
There is a need for improved therapies for severe asthma. Lebrikizumab, a humanized monoclonal antibody that binds to interleukin (IL)-13, is under development for the treatment of poorly controlled asthma. This article reviews the potential role of IL-13 in the pathogenesis of asthma, the efficacy and safety of lebrikizumab in humans, and progress in patient selection for lebrikizumab therapy. IL-13 is a T-helper (Th2) cell-derived cytokine implicated in inflammatory responses in asthma, including serum immunoglobulin-E synthesis, mucus hypersecretion, and subepithelial fibrosis. Blocking the pro-inflammatory effects of IL-13 with lebrikizumab has the potential to improve asthma control. Published data on the efficacy and safety of lebrikizumab in the treatment of asthma are relatively limited. The late asthmatic response after inhaled allergen challenge is reduced by almost 50%, following treatment with lebrikizumab. In a Phase II study performed in 219 adults with poorly controlled asthma despite inhaled corticosteroids (MILLY trial), lebrikizumab produced an improvement in prebronchodilator forced expiratory volume in 1 second of 5.5% compared with placebo at 12 weeks, but had no effects on other efficacy end points. Adverse effects were similar to placebo, except that musculoskeletal side effects occurred slightly more often with lebrikizumab. Stratifying patients into a high Th2 phenotype using serum periostin, which is upregulated in lung epithelial cells by IL-13, may identify individuals responsive to blockade of IL-13. In the MILLY trial, lebrikizumab treatment was associated with greater improvement in lung function in patients with elevated serum periostin levels compared with those with low periostin levels. Two large Phase III randomized controlled trials in patients with uncontrolled asthma are underway to establish the safety and efficacy of lebrikizumab when administered over a 52-week period. These studies will also help to determine whether identifying patients with a Th2 high inflammatory phenotype using serum periostin allows a personalized approach to the treatment of asthma.
doi:10.2147/BTT.S28666
PMCID: PMC3459551  PMID: 23055690
asthma; periostin; interleukin-13; phenotypes; exhaled nitric oxide; lebrikizumab
23.  Directional DNA Methylation Changes and Complex Intermediate States Accompany Lineage Specificity in the Adult Hematopoietic Compartment 
Molecular cell  2011;44(1):17-28.
SUMMARY
DNA methylation has been implicated as an epigenetic component of mechanisms that stabilize cell-fate decisions. Here, we have characterized the methylomes of human female hematopoietic stem/progenitor cells (HSPCs) and mature cells from the myeloid and lymphoid lineages. Hypomethylated regions (HMRs) associated with lineage-specific genes were often methylated in the opposing lineage. In HSPCs, these sites tended to show intermediate, complex patterns that resolve to uniformity upon differentiation, by increased or decreased methylation. Promoter HMRs shared across diverse cell types typically display a constitutive core that expands and contracts in a lineage-specific manner to fine-tune the expression of associated genes. Many newly identified intergenic HMRs, both constitutive and lineage specific, were enriched for factor binding sites with an implied role in genome organization and regulation of gene expression, respectively. Overall, our studies represent an important reference data set and provide insights into directional changes in DNA methylation as cells adopt terminal fates.
doi:10.1016/j.molcel.2011.08.026
PMCID: PMC3412369  PMID: 21924933
24.  Before It Gets Started: Regulating Translation at the 5′ UTR 
Translation regulation plays important roles in both normal physiological conditions and diseases states. This regulation requires cis-regulatory elements located mostly in 5′ and 3′ UTRs and trans-regulatory factors (e.g., RNA binding proteins (RBPs)) which recognize specific RNA features and interact with the translation machinery to modulate its activity. In this paper, we discuss important aspects of 5′ UTR-mediated regulation by providing an overview of the characteristics and the function of the main elements present in this region, like uORF (upstream open reading frame), secondary structures, and RBPs binding motifs and different mechanisms of translation regulation and the impact they have on gene expression and human health when deregulated.
doi:10.1155/2012/475731
PMCID: PMC3368165  PMID: 22693426
25.  Sperm methylation profiles reveal features of epigenetic inheritance and evolution in primates 
Cell  2011;146(6):1029-1041.
Summary
During germ cell and preimplantation development, mammalian cells undergo nearly complete reprogramming of DNA methylation patterns. We profiled the methylomes of human and chimp sperm as a basis for comparison to methylation patterns of ES cells. While the majority of promoters escape methylation in both ES cells and sperm, the corresponding hypomethylated regions show substantial structural differences. Repeat elements are heavily methylated in both germ and somatic cells; however, retrotransposons from several subfamilies evade methylation more effectively during male germ cell development, while other subfamilies show the opposite trend. Comparing methylomes of human and chimp sperm revealed a subset of differentially methylated promoters and strikingly divergent methylation in retrotransposon subfamilies, with an evolutionary impact that is apparent in the underlying genomic sequence. Thus, the features that determine DNA methylation patterns differ between male germ cells and somatic cells, and elements of these features have diverged between humans and chimpanzees.
doi:10.1016/j.cell.2011.08.016
PMCID: PMC3205962  PMID: 21925323

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