Publications by authors named "Brett Trost"

50 Publications

Enrichment of loss-of-function and copy number variants in ventricular cardiomyopathy genes in 'lone' atrial fibrillation.

Europace 2021 Mar 3. Epub 2021 Mar 3.

Section of Cardiac Electrophysiology, Division of Cardiology, Department of Medicine, Western University, London, ON, Canada.

Aims: Atrial fibrillation (AF) is a complex heritable disease whose genetic underpinnings remain largely unexplained, though recent work has suggested that the arrhythmia may develop secondary to an underlying atrial cardiomyopathy. We sought to evaluate for enrichment of loss-of-function (LOF) and copy number variants (CNVs) in genes implicated in ventricular cardiomyopathy in 'lone' AF.

Methods And Results: Whole-exome sequencing was performed in 255 early onset 'lone' AF cases, defined as arrhythmia onset prior to 60 years of age in the absence of known clinical risk factors. Subsequent evaluations were restricted to 195 cases of European genetic ancestry, as defined by principal component analysis, and focused on a pre-defined set of 43 genes previously implicated in ventricular cardiomyopathy. Bioinformatic analysis identified 6 LOF variants (3.1%), including 3 within the TTN gene, among cases in comparison with 4 of 503 (0.80%) controls [odds ratio: 3.96; 95% confidence interval (CI): 1.11-14.2; P = 0.033]. Further, two AF cases possessed a novel heterozygous 8521 base pair TTN deletion, confirmed with Sanger sequencing and breakpoint validation, which was absent from 4958 controls (P = 0.0014). Subsequent cascade screening in two families revealed evidence of co-segregation of a LOF variant with 'lone' AF.

Conclusion: 'Lone' AF cases are enriched in rare LOF variants from cardiomyopathy genes, findings primarily driven by TTN, and a novel TTN deletion, providing additional evidence to implicate atrial cardiomyopathy as an AF genetic sub-phenotype. Our results also highlight that AF may develop in the context of these variants in the absence of a discernable ventricular cardiomyopathy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/europace/euaa421DOI Listing
March 2021

Genome Sequencing as a Diagnostic Test in Children With Unexplained Medical Complexity.

JAMA Netw Open 2020 09 1;3(9):e2018109. Epub 2020 Sep 1.

Centre for Genetic Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada.

Importance: Children with medical complexity (CMC) represent a growing population in the pediatric health care system, with high resource use and associated health care costs. A genetic diagnosis can inform prognosis, anticipatory care, management, and reproductive planning. Conventional genetic testing strategies for CMC are often costly, time consuming, and ultimately unsuccessful.

Objective: To evaluate the analytical and clinical validity of genome sequencing as a comprehensive diagnostic genetic test for CMC.

Design, Setting, And Participants: In this cohort study of the prospective use of genome sequencing and comparison with standard-of-care genetic testing, CMC were recruited from May 1, 2017, to November 30, 2018, from a structured complex care program based at a tertiary care pediatric hospital in Toronto, Canada. Recruited CMC had at least 1 chronic condition, technology dependence (child is dependent at least part of each day on mechanical ventilators, and/or child requires prolonged intravenous administration of nutritional substances or drugs, and/or child is expected to have prolonged dependence on other device-based support), multiple subspecialist involvement, and substantial health care use. Review of the care plans for 545 CMC identified 143 suspected of having an undiagnosed genetic condition. Fifty-four families met inclusion criteria and were interested in participating, and 49 completed the study. Probands, similarly affected siblings, and biological parents were eligible for genome sequencing.

Exposures: Genome sequencing was performed using blood-derived DNA from probands and family members using established methods and a bioinformatics pipeline for clinical genome annotation.

Main Outcomes And Measures: The primary study outcome was the diagnostic yield of genome sequencing (proportion of CMC for whom the test result yielded a new diagnosis).

Results: Genome sequencing was performed for 138 individuals from 49 families of CMC (29 male and 20 female probands; mean [SD] age, 7.0 [4.5] years). Genome sequencing detected all genomic variation previously identified by conventional genetic testing. A total of 15 probands (30.6%; 95% CI 19.5%-44.6%) received a new primary molecular genetic diagnosis after genome sequencing. Three individuals had novel diseases and an additional 9 had either ultrarare genetic conditions or rare genetic conditions with atypical features. At least 11 families received diagnostic information that had clinical management implications beyond genetic and reproductive counseling.

Conclusions And Relevance: This study suggests that genome sequencing has high analytical and clinical validity and can result in new diagnoses in CMC even in the setting of extensive prior investigations. This clinical population may be enriched for ultrarare and novel genetic disorders. Genome sequencing is a potentially first-tier genetic test for CMC.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1001/jamanetworkopen.2020.18109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7509619PMC
September 2020

Genome-wide detection of tandem DNA repeats that are expanded in autism.

Nature 2020 10 27;586(7827):80-86. Epub 2020 Jul 27.

Genetics and Genome Biology, The Hospital for Sick Children, Toronto, Ontario, Canada.

Tandem DNA repeats vary in the size and sequence of each unit (motif). When expanded, these tandem DNA repeats have been associated with more than 40 monogenic disorders. Their involvement in disorders with complex genetics is largely unknown, as is the extent of their heterogeneity. Here we investigated the genome-wide characteristics of tandem repeats that had motifs with a length of 2-20 base pairs in 17,231 genomes of families containing individuals with autism spectrum disorder (ASD) and population control individuals. We found extensive polymorphism in the size and sequence of motifs. Many of the tandem repeat loci that we detected correlated with cytogenetic fragile sites. At 2,588 loci, gene-associated expansions of tandem repeats that were rare among population control individuals were significantly more prevalent among individuals with ASD than their siblings without ASD, particularly in exons and near splice junctions, and in genes related to the development of the nervous system and cardiovascular system or muscle. Rare tandem repeat expansions had a prevalence of 23.3% in children with ASD compared with 20.7% in children without ASD, which suggests that tandem repeat expansions make a collective contribution to the risk of ASD of 2.6%. These rare tandem repeat expansions included previously undescribed ASD-linked expansions in DMPK and FXN, which are associated with neuromuscular conditions, and in previously unknown loci such as FGF14 and CACNB1. Rare tandem repeat expansions were associated with lower IQ and adaptive ability. Our results show that tandem DNA repeat expansions contribute strongly to the genetic aetiology and phenotypic complexity of ASD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41586-020-2579-zDOI Listing
October 2020

ExpansionHunter Denovo: a computational method for locating known and novel repeat expansions in short-read sequencing data.

Genome Biol 2020 04 28;21(1):102. Epub 2020 Apr 28.

Illumina Inc., 5200 Illumina Way, San Diego, CA, 92122, USA.

Repeat expansions are responsible for over 40 monogenic disorders, and undoubtedly more pathogenic repeat expansions remain to be discovered. Existing methods for detecting repeat expansions in short-read sequencing data require predefined repeat catalogs. Recent discoveries emphasize the need for methods that do not require pre-specified candidate repeats. To address this need, we introduce ExpansionHunter Denovo, an efficient catalog-free method for genome-wide repeat expansion detection. Analysis of real and simulated data shows that our method can identify large expansions of 41 out of 44 pathogenic repeats, including nine recently reported non-reference repeat expansions not discoverable via existing methods.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13059-020-02017-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7187524PMC
April 2020

Associations of clinical and inflammatory biomarker clusters with juvenile idiopathic arthritis categories.

Rheumatology (Oxford) 2020 05;59(5):1066-1075

Department of Pediatrics, University of Saskatchewan, Saskatoon, Canada.

Objective: To identify discrete clusters comprising clinical features and inflammatory biomarkers in children with JIA and to determine cluster alignment with JIA categories.

Methods: A Canadian prospective inception cohort comprising 150 children with JIA was evaluated at baseline (visit 1) and after six months (visit 2). Data included clinical manifestations and inflammation-related biomarkers. Probabilistic principal component analysis identified sets of composite variables, or principal components, from 191 original variables. To discern new clinical-biomarker clusters (clusters), Gaussian mixture models were fit to the data. Newly-defined clusters and JIA categories were compared. Agreement between the two was assessed using Kruskal-Wallis analyses and contingency plots.

Results: Three principal components recovered 35% (three clusters) and 40% (five clusters) of the variance in patient profiles in visits 1 and 2, respectively. None of the clusters aligned precisely with any of the seven JIA categories but rather spanned multiple categories. Results demonstrated that the newly defined clinical-biomarker lustres are more homogeneous than JIA categories.

Conclusion: Applying unsupervised data mining to clinical and inflammatory biomarker data discerns discrete clusters that intersect multiple JIA categories. Results suggest that certain groups of patients within different JIA categories are more aligned pathobiologically than their separate clinical categorizations suggest. Applying data mining analyses to complex datasets can generate insights into JIA pathogenesis and could contribute to biologically based refinements in JIA classification.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/rheumatology/kez382DOI Listing
May 2020

Clinical and associated inflammatory biomarker features predictive of short-term outcomes in non-systemic juvenile idiopathic arthritis.

Rheumatology (Oxford) 2020 09;59(9):2402-2411

Department of PediatricsUniversity of Saskatchewan, Saskatoon, SK, Canada.

Objective: To identify early predictors of disease activity at 18 months in JIA using clinical and biomarker profiling.

Methods: Clinical and biomarker data were collected at JIA diagnosis in a prospective longitudinal inception cohort of 82 children with non-systemic JIA, and their ability to predict an active joint count of 0, a physician global assessment of disease activity of ≤1 cm, and inactive disease by Wallace 2004 criteria 18 months later was assessed. Correlation-based feature selection and ReliefF were used to shortlist predictors and random forest models were trained to predict outcomes.

Results: From the original 112 features, 13 effectively predicted 18-month outcomes. They included age, number of active/effused joints, wrist, ankle and/or knee involvement, ESR, ANA positivity and plasma levels of five inflammatory biomarkers (IL-10, IL-17, IL-12p70, soluble low-density lipoprotein receptor-related protein 1 and vitamin D), at enrolment. The clinical plus biomarker panel predicted active joint count = 0, physician global assessment ≤ 1, and inactive disease after 18 months with 0.79, 0.80 and 0.83 accuracy and 0.84, 0.83, 0.88 area under the curve, respectively. Using clinical features alone resulted in 0.75, 0.72 and 0.80 accuracy, and area under the curve values of 0.81, 0.78 and 0.83, respectively.

Conclusion: A panel of five plasma biomarkers combined with clinical features at the time of diagnosis more accurately predicted short-term disease activity in JIA than clinical characteristics alone. If validated in external cohorts, such a panel may guide more rationally conceived, biologically based, personalized treatment strategies in early JIA.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/rheumatology/kez615DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7449798PMC
September 2020

A large data resource of genomic copy number variation across neurodevelopmental disorders.

NPJ Genom Med 2019 7;4:26. Epub 2019 Oct 7.

1The Centre for Applied Genomics, The Hospital for Sick Children, Toronto, ON Canada.

Copy number variations (CNVs) are implicated across many neurodevelopmental disorders (NDDs) and contribute to their shared genetic etiology. Multiple studies have attempted to identify shared etiology among NDDs, but this is the first genome-wide CNV analysis across autism spectrum disorder (ASD), attention deficit hyperactivity disorder (ADHD), schizophrenia (SCZ), and obsessive-compulsive disorder (OCD) at once. Using microarray (Affymetrix CytoScan HD), we genotyped 2,691 subjects diagnosed with an NDD (204 SCZ, 1,838 ASD, 427 ADHD and 222 OCD) and 1,769 family members, mainly parents. We identified rare CNVs, defined as those found in <0.1% of 10,851 population control samples. We found clinically relevant CNVs (broadly defined) in 284 (10.5%) of total subjects, including 22 (10.8%) among subjects with SCZ, 209 (11.4%) with ASD, 40 (9.4%) with ADHD, and 13 (5.6%) with OCD. Among all NDD subjects, we identified 17 (0.63%) with aneuploidies and 115 (4.3%) with known genomic disorder variants. We searched further for genes impacted by different CNVs in multiple disorders. Examples of NDD-associated genes linked across more than one disorder (listed in order of occurrence frequency) are , , , , , , , , , , and long non-coding RNAs: and . We demonstrated that CNVs impacting the same genes could potentially contribute to the etiology of multiple NDDs. The CNVs identified will serve as a useful resource for both research and diagnostic laboratories for prioritization of variants.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41525-019-0098-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6779875PMC
October 2019

Impact of DNA source on genetic variant detection from human whole-genome sequencing data.

J Med Genet 2019 12 12;56(12):809-817. Epub 2019 Sep 12.

The Centre for Applied Genomics, Hospital for Sick Children, Toronto, Ontario, Canada

Background: Whole blood is currently the most common DNA source for whole-genome sequencing (WGS), but for studies requiring non-invasive collection, self-collection, greater sample stability or additional tissue references, saliva or buccal samples may be preferred. However, the relative quality of sequencing data and accuracy of genetic variant detection from blood-derived, saliva-derived and buccal-derived DNA need to be thoroughly investigated.

Methods: Matched blood, saliva and buccal samples from four unrelated individuals were used to compare sequencing metrics and variant-detection accuracy among these DNA sources.

Results: We observed significant differences among DNA sources for sequencing quality metrics such as percentage of reads aligned and mean read depth (p<0.05). Differences were negligible in the accuracy of detecting short insertions and deletions; however, the false positive rate for single nucleotide variation detection was slightly higher in some saliva and buccal samples. The sensitivity of copy number variant (CNV) detection was up to 25% higher in blood samples, depending on CNV size and type, and appeared to be worse in saliva and buccal samples with high bacterial concentration. We also show that methylation-based enrichment for eukaryotic DNA in saliva and buccal samples increased alignment rates but also reduced read-depth uniformity, hampering CNV detection.

Conclusion: For WGS, we recommend using DNA extracted from blood rather than saliva or buccal swabs; if saliva or buccal samples are used, we recommend against using methylation-based eukaryotic DNA enrichment. All data used in this study are available for further open-science investigation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1136/jmedgenet-2019-106281DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6929712PMC
December 2019

Length of Uninterrupted CAG, Independent of Polyglutamine Size, Results in Increased Somatic Instability, Hastening Onset of Huntington Disease.

Am J Hum Genet 2019 06 16;104(6):1116-1126. Epub 2019 May 16.

Centre for Molecular Medicine and Therapeutics, Department of Medical Genetics, University of British Columbia, Vancouver, BC V5Z 4H4, Canada. Electronic address:

Huntington disease (HD) is caused by a CAG repeat expansion in the huntingtin (HTT) gene. Although the length of this repeat is inversely correlated with age of onset (AOO), it does not fully explain the variability in AOO. We assessed the sequence downstream of the CAG repeat in HTT [reference: (CAG)n-CAA-CAG], since variants within this region have been previously described, but no study of AOO has been performed. These analyses identified a variant that results in complete loss of interrupting (LOI) adenine nucleotides in this region [(CAG)n-CAG-CAG]. Analysis of multiple HD pedigrees showed that this LOI variant is associated with dramatically earlier AOO (average of 25 years) despite the same polyglutamine length as in individuals with the interrupting penultimate CAA codon. This LOI allele is particularly frequent in persons with reduced penetrance alleles who manifest with HD and increases the likelihood of presenting clinically with HD with a CAG of 36-39 repeats. Further, we show that the LOI variant is associated with increased somatic repeat instability, highlighting this as a significant driver of this effect. These findings indicate that the number of uninterrupted CAG repeats, which is lengthened by the LOI, is the most significant contributor to AOO of HD and is more significant than polyglutamine length, which is not altered in these individuals. In addition, we identified another variant in this region, where the CAA-CAG sequence is duplicated, which was associated with later AOO. Identification of these cis-acting modifiers have potentially important implications for genetic counselling in HD-affected families.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ajhg.2019.04.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6556907PMC
June 2019

Glutaminase Deficiency Caused by Short Tandem Repeat Expansion in .

N Engl J Med 2019 04;380(15):1433-1441

From Amsterdam University Medical Centers, University of Amsterdam, Departments of Clinical Chemistry, Pediatrics, and Clinical Genetics, Emma Children's Hospital, Amsterdam Gastroenterology and Metabolism (A.B.P.K., R.L., J.K., J. Meijer, L.A.T., M.T., M.W., R.J.A.W., H.R.W., C.D.M.K.), and United for Metabolic Diseases (A.B.P.K., R.J.A.W., H.R.W., C.D.M.K.), Amsterdam, and the Department of Neurology, Brain Center Rudolf Magnus, University Medical Center Utrecht (J.J.F.A.V., J.H.V.), and the Project MinE ALS Sequencing Consortium (J.J.F.A.V., J.H.V.), Utrecht - all in the Netherlands; the Departments of Biochemistry and Molecular Biology and Medical Genetics, Cumming School of Medicine, and Alberta Children's Hospital Research Institute, University of Calgary, Calgary (M.T.-G.), Centre for Molecular Medicine and Therapeutics, BC Children's Hospital Research Institute (P.A.R., M.J.J., M.S.K., J. MacIsaac, W.W.W., C.D.M.K.), the Faculty of Pharmaceutical Sciences (B.I.D., G.E.B.W., C.J.R.), and the Departments of Medical Genetics (C.M., I.-S.R.-B., W.W.W.) and Pediatrics (C.D.M.K.), University of British Columbia, Vancouver, the Zebrafish Centre for Advanced Drug Discovery, St. Michael's Hospital and University of Toronto (K.B.-A., F.K., M.L., Y.W., X.-Y.W.), the Centre for Applied Genomics, Genetics and Genome Biology, the Hospital for Sick Children (C.N., S.W.S., B.T., R.K.C.Y.), and the Department of Molecular Genetics (C.N., S.W.S., R.K.C.Y.), the McLaughlin Centre (S.W.S.), and the Departments of Medicine, Physiology, and Laboratory Medicine and Pathobiology, Institute of Medical Science (X.-Y.W.), University of Toronto, Toronto, and the Division of Medical Genetics, Department of Pediatrics, Children's Hospital Eastern Ontario, University of Ottawa, Ottawa (J.S.W., M.T.G.) - all in Canada; the Departments of Medicine and Physiology, National University of Singapore (M.A.P.), and the Translational Laboratory in Genetic Medicine, Agency for Science, Technology, and Research (M.A.P., B.S., X.X., J.Z.) - both in Singapore; Uppsala University, Department of Chemistry-Biomedical Center, Uppsala, Sweden (D.D.); Illumina, San Diego, CA (E.D., M.A.E.); Gene Structure and Disease Section, Laboratory of Cell and Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD (B.H., D.K., K.U.); and the Department of Clinical Inherited Metabolic Disorders, Birmingham Children's Hospital, Birmingham, United Kingdom (S.S.).

We report an inborn error of metabolism caused by an expansion of a GCA-repeat tract in the 5' untranslated region of the gene encoding glutaminase () that was identified through detailed clinical and biochemical phenotyping, combined with whole-genome sequencing. The expansion was observed in three unrelated patients who presented with an early-onset delay in overall development, progressive ataxia, and elevated levels of glutamine. In addition to ataxia, one patient also showed cerebellar atrophy. The expansion was associated with a relative deficiency of messenger RNA transcribed from the expanded allele, which probably resulted from repeat-mediated chromatin changes upstream of the repeat. Our discovery underscores the importance of careful examination of regions of the genome that are typically excluded from or poorly captured by exome sequencing.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1056/NEJMoa1806627DOI Listing
April 2019

Conservation of kinase-phosphorylation site pairings: Evidence for an evolutionarily dynamic phosphoproteome.

PLoS One 2018 14;13(8):e0202036. Epub 2018 Aug 14.

Vaccine and Infectious Disease Organization-International Vaccine Research Center, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.

Kinase-mediated protein phosphorylation is a central mechanism for regulation of cellular responses and phenotypes. While considerable information is available regarding the evolutionary relationships within the kinase family, as well as the evolutionary conservation of phosphorylation sites, each aspect of this partnership is typically considered in isolation, despite their clear functional relationship. Here, to offer a more holistic perspective on the evolution of protein phosphorylation, the conservation of protein phosphorylation sites is considered in the context of the conservation of the corresponding modifying kinases. Specifically, conservation of defined kinase-phosphorylation site pairings (KPSPs), as well as of each of the component parts (the kinase and the phosphorylation site), were examined across a range of species. As expected, greater evolutionary distance between species was generally associated with lower probability of KPSP conservation, and only a small fraction of KPSPs were maintained across all species, with the vast majority of KPSP losses due to the absence of the phosphorylation site. This supports a model in which a relatively stable kinome promotes the emergence of functional substrates from an evolutionarily malleable phosphoproteome.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0202036PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6091962PMC
January 2019

The Personal Genome Project Canada: findings from whole genome sequences of the inaugural 56 participants.

CMAJ 2018 02;190(5):E126-E136

The Centre for Applied Genomics (Reuter, Walker, Thiruvahindrapuram, Whitney, Yuen, Trost, Paton, Pereira, Herbrick, Wintle, Merico, Howe, MacDonald, Lu, Nalpathamkalam, Sung, Wang, Patel, Pellecchia, J. Wei, Strug, Bell, Kellam, Mahtani, Hosseini, Fiume, Marshall, Buchanan, Scherer); Divisions of Clinical Pharmacology and Toxicology (I. Cohn), or Clinical, and Metabolic Genetics (Sondheimer, Weksberg, Shuman, Bowdin, Meyn, Monfared), The Hospital for Sick Children; Departments of Paediatrics (Sondheimer, R. Cohn) and Molecular Genetics (Yuen, Weksberg, Shuman, R. Cohn, Ellis, Meyn), University of Toronto; Deep Genomics Inc. (Merico); Department of Psychiatry (Bassett), University Health Network and Centre for Addiction and Mental Health, University of Toronto; Li Ka Shing Knowledge Institute (Bombard), St. Michael's Hospital; Institute of Health Policy, Management and Evaluation (Bombard), University of Toronto; Centre for Genetic Medicine (Stavropoulos, Bowdin, Ray, Monfared); Molecular Genetics Laboratory (Stavropoulos, Ray, Marshall), Division of Genome Diagnostics, Paediatric Laboratory Medicine; Developmental and Stem Cell Biology (Hildebrandt, W. Wei, Romm, Pasceri, Ellis); Ted Rogers Cardiac Genome Clinic (Hosseini); Cytogenetics Laboratory (Joseph-George), Division of Genome Diagnostics, Paediatric Laboratory Medicine, The Hospital for Sick Children; Departments of Biochemistry and Laboratory Medicine, and Pathobiology (Keeley), University of Toronto; DNAstack (Cook, Fiume); McLaughlin Centre (Lee, Scherer), University of Toronto; Medcan Health Management Inc. (Davies, Hazell); Dalla Lana School of Public Health (Szego), Department of Family and Community Medicine, and The Joint Centre for Bioethics, University of Toronto; Centre for Clinical Ethics (Szego), St. Joseph's Health Centre, Toronto, Ont.

Background: The Personal Genome Project Canada is a comprehensive public data resource that integrates whole genome sequencing data and health information. We describe genomic variation identified in the initial recruitment cohort of 56 volunteers.

Methods: Volunteers were screened for eligibility and provided informed consent for open data sharing. Using blood DNA, we performed whole genome sequencing and identified all possible classes of DNA variants. A genetic counsellor explained the implication of the results to each participant.

Results: Whole genome sequencing of the first 56 participants identified 207 662 805 sequence variants and 27 494 copy number variations. We analyzed a prioritized disease-associated data set ( = 1606 variants) according to standardized guidelines, and interpreted 19 variants in 14 participants (25%) as having obvious health implications. Six of these variants (e.g., in or mosaic loss of an X chromosome) were pathogenic or likely pathogenic. Seven were risk factors for cancer, cardiovascular or neurobehavioural conditions. Four other variants - associated with cancer, cardiac or neurodegenerative phenotypes - remained of uncertain significance because of discrepancies among databases. We also identified a large structural chromosome aberration and a likely pathogenic mitochondrial variant. There were 172 recessive disease alleles (e.g., 5 individuals carried mutations for cystic fibrosis). Pharmacogenomics analyses revealed another 3.9 potentially relevant genotypes per individual.

Interpretation: Our analyses identified a spectrum of genetic variants with potential health impact in 25% of participants. When also considering recessive alleles and variants with potential pharmacologic relevance, all 56 participants had medically relevant findings. Although access is mostly limited to research, whole genome sequencing can provide specific and novel information with the potential of major impact for health care.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1503/cmaj.171151DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5798982PMC
February 2018

Transcriptome analysis of response to Plasmodiophora brassicae infection in the Arabidopsis shoot and root.

BMC Genomics 2018 01 5;19(1):23. Epub 2018 Jan 5.

Department of Biology, University of Saskatchewan, Saskatoon, S7N 5E2, Canada.

Background: Clubroot is an important disease caused by the obligate parasite Plasmodiophora brassicae that infects the Brassicaceae. As a soil-borne pathogen, P. brassicae induces the generation of abnormal tissue in the root, resulting in the formation of galls. Root infection negatively affects the uptake of water and nutrients in host plants, severely reducing their growth and productivity. Many studies have emphasized the molecular and physiological effects of the clubroot disease on root tissues. The aim of the present study is to better understand the effect of P. brassicae on the transcriptome of both shoot and root tissues of Arabidopsis thaliana.

Results: Transcriptome profiling using RNA-seq was performed on both shoot and root tissues at 17, 20 and 24 days post inoculation (dpi) of A. thaliana, a model plant host for P. brassicae. The number of differentially expressed genes (DEGs) between infected and uninfected samples was larger in shoot than in root. In both shoot and root, more genes were differentially regulated at 24 dpi than the two earlier time points. Genes that were highly regulated in response to infection in both shoot and root primarily were involved in the metabolism of cell wall compounds, lipids, and shikimate pathway metabolites. Among hormone-related pathways, several jasmonic acid biosynthesis genes were upregulated in both shoot and root tissue. Genes encoding enzymes involved in cell wall modification, biosynthesis of sucrose and starch, and several classes of transcription factors were generally differently regulated in shoot and root.

Conclusions: These results highlight the similarities and differences in the transcriptomic response of above- and below-ground tissues of the model host Arabidopsis following P. brassicae infection. The main transcriptomic changes in root metabolism during clubroot disease progression were identified. An overview of DEGs in the shoot underlined the physiological changes in above-ground tissues following pathogen establishment and disease progression. This study provides insights into host tissue-specific molecular responses to clubroot development and may have applications in the development of clubroot markers for more effective breeding strategies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s12864-017-4426-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5756429PMC
January 2018

A Comprehensive Workflow for Read Depth-Based Identification of Copy-Number Variation from Whole-Genome Sequence Data.

Am J Hum Genet 2018 01;102(1):142-155

The Centre for Applied Genomics, Genetics and Genome Biology, The Hospital for Sick Children, Toronto, ON M5G 0A4, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada; McLaughlin Centre, University of Toronto, Toronto, ON M5G 0A4, Canada. Electronic address:

A remaining hurdle to whole-genome sequencing (WGS) becoming a first-tier genetic test has been accurate detection of copy-number variations (CNVs). Here, we used several datasets to empirically develop a detailed workflow for identifying germline CNVs >1 kb from short-read WGS data using read depth-based algorithms. Our workflow is comprehensive in that it addresses all stages of the CNV-detection process, including DNA library preparation, sequencing, quality control, reference mapping, and computational CNV identification. We used our workflow to detect rare, genic CNVs in individuals with autism spectrum disorder (ASD), and 120/120 such CNVs tested using orthogonal methods were successfully confirmed. We also identified 71 putative genic de novo CNVs in this cohort, which had a confirmation rate of 70%; the remainder were incorrectly identified as de novo due to false positives in the proband (7%) or parental false negatives (23%). In individuals with an ASD diagnosis in which both microarray and WGS experiments were performed, our workflow detected all clinically relevant CNVs identified by microarrays, as well as additional potentially pathogenic CNVs < 20 kb. Thus, CNVs of clinical relevance can be discovered from WGS with a detection rate exceeding microarrays, positioning WGS as a single assay for genetic variation detection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ajhg.2017.12.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5777982PMC
January 2018

Technological advances for interrogating the human kinome.

Biochem Soc Trans 2017 02;45(1):65-77

Department of Biochemistry, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5E5, Canada

There is increasing appreciation among researchers and clinicians of the value of investigating biology and pathobiology at the level of cellular kinase (kinome) activity. Kinome analysis provides valuable opportunity to gain insights into complex biology (including disease pathology), identify biomarkers of critical phenotypes (including disease prognosis and evaluation of therapeutic efficacy), and identify targets for therapeutic intervention through kinase inhibitors. The growing interest in kinome analysis has fueled efforts to develop and optimize technologies that enable characterization of phosphorylation-mediated signaling events in a cost-effective, high-throughput manner. In this review, we highlight recent advances to the central technologies currently available for kinome profiling and offer our perspectives on the key challenges remaining to be addressed.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1042/BST20160163DOI Listing
February 2017

Genome and Transcriptome Assembly of the Canadian Beaver ().

G3 (Bethesda) 2017 02 9;7(2):755-773. Epub 2017 Feb 9.

The Centre for Applied Genomics, The Hospital for Sick Children, Toronto, Ontario M5G 0A4, Canada

The Canadian beaver () is the largest indigenous rodent in North America. We report a draft annotated assembly of the beaver genome, the first for a large rodent and the first mammalian genome assembled directly from uncorrected and moderate coverage (< 30 ×) long reads generated by single-molecule sequencing. The genome size is 2.7 Gb estimated by k-mer analysis. We assembled the beaver genome using the new Canu assembler optimized for noisy reads. The resulting assembly was refined using Pilon supported by short reads (80 ×) and checked for accuracy by congruency against an independent short read assembly. We scaffolded the assembly using the exon-gene models derived from 9805 full-length open reading frames (FL-ORFs) constructed from the beaver leukocyte and muscle transcriptomes. The final assembly comprised 22,515 contigs with an N50 of 278,680 bp and an N50-scaffold of 317,558 bp. Maximum contig and scaffold lengths were 3.3 and 4.2 Mb, respectively, with a combined scaffold length representing 92% of the estimated genome size. The completeness and accuracy of the scaffold assembly was demonstrated by the precise exon placement for 91.1% of the 9805 assembled FL-ORFs and 83.1% of the BUSCO (Benchmarking Universal Single-Copy Orthologs) gene set used to assess the quality of genome assemblies. Well-represented were genes involved in dentition and enamel deposition, defining characteristics of rodents with which the beaver is well-endowed. The study provides insights for genome assembly and an important genomics resource for Castoridae and rodent evolutionary biology.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1534/g3.116.038208DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5295618PMC
February 2017

DAPPLE 2: a Tool for the Homology-Based Prediction of Post-Translational Modification Sites.

J Proteome Res 2016 08 13;15(8):2760-7. Epub 2016 Jul 13.

Vaccine and Infectious Disease Organization, ‡Department of Computer Science, and §Department of Biochemistry, University of Saskatchewan , Saskatoon, SK S7N 5A2, Canada.

The post-translational modification of proteins is critical for regulating their function. Although many post-translational modification sites have been experimentally determined, particularly in certain model organisms, experimental knowledge of these sites is severely lacking for many species. Thus, it is important to be able to predict sites of post-translational modification in such species. Previously, we described DAPPLE, a tool that facilitates the homology-based prediction of one particular post-translational modification, phosphorylation, in an organism of interest using known phosphorylation sites from other organisms. Here, we describe DAPPLE 2, which expands and improves upon DAPPLE in three major ways. First, it predicts sites for many post-translational modifications (20 different types) using data from several sources (15 online databases). Second, it has the ability to make predictions approximately 2-7 times faster than DAPPLE depending on the database size and the organism of interest. Third, it simplifies and accelerates the process of selecting predicted sites of interest by categorizing them based on gene ontology terms, keywords, and signaling pathways. We show that DAPPLE 2 can successfully predict known human post-translational modification sites using, as input, known sites from species that are either closely (e.g., mouse) or distantly (e.g., yeast) related to humans. DAPPLE 2 can be accessed at http://saphire.usask.ca/saphire/dapple2 .
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.jproteome.6b00304DOI Listing
August 2016

Computational Analysis of the Predicted Evolutionary Conservation of Human Phosphorylation Sites.

PLoS One 2016 5;11(4):e0152809. Epub 2016 Apr 5.

Vaccine and Infectious Disease Organization, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.

Protein kinase-mediated phosphorylation is among the most important post-translational modifications. However, few phosphorylation sites have been experimentally identified for most species, making it difficult to determine the degree to which phosphorylation sites are conserved. The goal of this study was to use computational methods to characterize the conservation of human phosphorylation sites in a wide variety of eukaryotes. Using experimentally-determined human sites as input, homologous phosphorylation sites were predicted in all 432 eukaryotes for which complete proteomes were available. For each pair of species, we calculated phosphorylation site conservation as the number of phosphorylation sites found in both species divided by the number found in at least one of the two species. A clustering of the species based on this conservation measure was concordant with phylogenies based on traditional genomic measures. For a subset of the 432 species, phosphorylation site conservation was compared to conservation of both protein kinases and proteins in general. Protein kinases exhibited the highest degree of conservation, while general proteins were less conserved and phosphorylation sites were least conserved. Although preliminary, these data tentatively suggest that variation in phosphorylation sites may play a larger role in explaining phenotypic differences among organisms than differences in the complements of protein kinases or general proteins.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0152809PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4821552PMC
August 2016

Investigation of the cause of geographic disparities in IDEXX ELISA sensitivity in serum samples from Mycobacterium bovis-infected cattle.

Sci Rep 2016 Mar 7;6:22763. Epub 2016 Mar 7.

Vaccine and Infectious Disease Organization, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5E3, Canada.

Accurately identifying Mycobacterium bovis-infected cattle is critical for bovine tuberculosis prevention and control. One method for identifying infected cattle is an ELISA developed by IDEXX laboratories, which detects antibodies to two M. bovis proteins, MPB70 and MPB83. The assay's sensitivity varies by geographic region, with sensitivities of 77%, 45%, and 9% in bovine serum samples from the United Kingdom (n = 126), the United States (n = 146), and Mexico (n = 128), respectively. We hypothesized that geographically-biased sequence variation in mpb70 and mpb83, or in the genes that regulate their expression (sigK and rskA), may explain these differing sensitivities. This hypothesis was tested by comparing the sequences of these four genes in 455 M. bovis strains isolated from cattle in the aforementioned countries. For each gene, a single, common sequence was identified in most genomes of the M. bovis strains collected in all three countries. Twelve of the 455 strains were isolated from infected cattle for which the IDEXX ELISA was also performed. Five of the seven ELISA-positive genomes and three of the five ELISA-negative genomes contained the most common sequence of all four genes. Thus, sequence variation in mpb70, mpb83, sigK, and rskA does not explain the geographic disparities in IDEXX ELISA sensitivity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/srep22763DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4780098PMC
March 2016

Rapamycin reduces fibroblast proliferation without causing quiescence and induces STAT5A/B-mediated cytokine production.

Nucleus 2015 ;6(6):490-506

a Department of Food and Bioproduct Sciences ; University of Saskatchewan ; Saskatoon , Canada.

Rapamycin is a well-known inhibitor of the Target of Rapamycin (TOR) signaling cascade; however, the impact of this drug on global genome function and organization in normal primary cells is poorly understood. To explore this impact, we treated primary human foreskin fibroblasts with rapamycin and observed a decrease in cell proliferation without causing cell death. Upon rapamycin treatment chromosomes 18 and 10 were repositioned to a location similar to that of fibroblasts induced into quiescence by serum reduction. Although similar changes in positioning occurred, comparative transcriptome analyses demonstrated significant divergence in gene expression patterns between rapamycin-treated and quiescence-induced fibroblasts. Rapamycin treatment induced the upregulation of cytokine genes, including those from the Interleukin (IL)-6 signaling network, such as IL-8 and the Leukemia Inhibitory Factor (LIF), while quiescent fibroblasts demonstrated up-regulation of genes involved in the complement and coagulation cascade. In addition, genes significantly up-regulated by rapamycin treatment demonstrated increased promoter occupancy of the transcription factor Signal Transducer and Activator of Transcription 5A/B (STAT5A/B). In summary, we demonstrated that the treatment of fibroblasts with rapamycin decreased proliferation, caused chromosome territory repositioning and induced STAT5A/B-mediated changes in gene expression enriched for cytokines.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/19491034.2015.1128610DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4915505PMC
November 2016

Concordance between RNA-sequencing data and DNA microarray data in transcriptome analysis of proliferative and quiescent fibroblasts.

R Soc Open Sci 2015 Sep 30;2(9):150402. Epub 2015 Sep 30.

Department of Life Sciences , Brunel University , Uxbridge UB8 3PH, UK ; Department of Food and Bioproduct Sciences , University of Saskatchewan , Saskatoon Canada S7N 5A8.

DNA microarrays and RNA sequencing (RNA-seq) are major technologies for performing high-throughput analysis of transcript abundance. Recently, concerns have been raised regarding the concordance of data derived from the two techniques. Using cDNA libraries derived from normal human foreskin fibroblasts, we measured changes in transcript abundance as cells transitioned from proliferative growth to quiescence using both DNA microarrays and RNA-seq. The internal reproducibility of the RNA-seq data was greater than that of the microarray data. Correlations between the RNA-seq data and the individual microarrays were low, but correlations between the RNA-seq values and the geometric mean of the microarray values were moderate. The two technologies had good agreement when considering probes with the largest (both positive and negative) fold change (FC) values. An independent technique, quantitative reverse-transcription PCR (qRT-PCR), was used to measure the FC of 76 genes between proliferative and quiescent samples, and a higher correlation was observed between the qRT-PCR data and the RNA-seq data than between the qRT-PCR data and the microarray data.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1098/rsos.150402DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4593695PMC
September 2015

Induction of tissue- and stressor-specific kinomic responses in chickens exposed to hot and cold stresses.

Poult Sci 2015 Jun 1;94(6):1333-45. Epub 2015 Apr 1.

Department of Food and Bioproduct Science, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5A8, Canada.

Defining cellular responses at the level of global cellular kinase (kinome) activity is a powerful approach to deciphering complex biology and identifying biomarkers. Here we report on the development of a chicken-specific peptide array and its application to characterizing kinome responses within the breast (pectoralis major) and thigh (iliotibialis) muscles of poultry subject to temperature stress to mimic conditions experienced by birds during commercial transport. Breast and thigh muscles exhibited unique kinome profiles, highlighting the distinct nature of these tissues. Against these distinct backgrounds, tissue- and temperature-specific kinome responses were observed. In breast, both cold and hot stresses activated calcium-dependent metabolic adaptations. Also within breast, but specific to cold stress, was the activation of ErbB signaling as well as dynamic patterns of phosphorylation of AMPK, a key regulatory enzyme of metabolism. In thigh, cold stress induced responses suggestive of the occurrence of tissue damage, including activation of innate immune signaling pathways and tissue repair pathways (TGF-β). In contrast, heat stress in thigh activated pathways associated with protein and fat metabolism through adipocytokine and mammalian target of rapamycin (mTOR) signaling. Defining the responses of these tissues to these stresses through conventional markers of pH, glycolytic potential, and meat quality offered a similar conclusion of the tissue- and stressor-specific responses, validating the kinome results. Collectively, the results of this study highlight the unique cellular responses of breast and thigh tissues to heat and cold stresses and may offer insight into the unique susceptibilities, as well as functional consequences, of these tissues to thermal stress.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3382/ps/pev046DOI Listing
June 2015

EpIC: a rational pipeline for epitope immunogenicity characterization.

Bioinformatics 2015 Jul 6;31(14):2388-90. Epub 2015 Mar 6.

Department of Biochemistry, University of Saskatchewan, Saskatoon, SK, S7N 5E5 Canada, Vaccine and Infectious Disease Organization-International Vaccine Centre, University of Saskatchewan, Saskatoon, SK, S7N 5E3 Canada and.

Unlabelled: Efforts to develop peptide-based vaccines, in particular those requiring site-specific targeting of self-proteins, rely on the ability to optimize the immunogenicity of the peptide epitopes. Currently, screening of candidate vaccines is typically performed through low-throughput, high-cost animal trials. To improve on this we present the program EpIC, which enables high-throughput prediction of peptide immunogenicity based on the endogenous occurrence of B-cell epitopes within native protein sequences. This information informs rational selection of immunogenicity-optimized epitopes for peptide vaccines.

Availability And Implementation: EpIC is available as a web server at http://saphire.usask.ca/saphire/epic.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/bioinformatics/btv136DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4495294PMC
July 2015

Characterization of the host response to pichinde virus infection in the Syrian golden hamster by species-specific kinome analysis.

Mol Cell Proteomics 2015 Mar 8;14(3):646-57. Epub 2015 Jan 8.

§§Integrated Research Facility, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Frederick, Maryland, USA;

The Syrian golden hamster has been increasingly used to study viral hemorrhagic fever (VHF) pathogenesis and countermeasure efficacy. As VHFs are a global health concern, well-characterized animal models are essential for both the development of therapeutics and vaccines as well as for increasing our understanding of the molecular events that underlie viral pathogenesis. However, the paucity of reagents or platforms that are available for studying hamsters at a molecular level limits the ability to extract biological information from this important animal model. As such, there is a need to develop platforms/technologies for characterizing host responses of hamsters at a molecular level. To this end, we developed hamster-specific kinome peptide arrays to characterize the molecular host response of the Syrian golden hamster. After validating the functionality of the arrays using immune agonists of defined signaling mechanisms (lipopolysaccharide (LPS) and tumor necrosis factor (TNF)-α), we characterized the host response in a hamster model of VHF based on Pichinde virus (PICV(1)) infection by performing temporal kinome analysis of lung tissue. Our analysis revealed key roles for vascular endothelial growth factor (VEGF), interleukin (IL) responses, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling, and Toll-like receptor (TLR) signaling in the response to PICV infection. These findings were validated through phosphorylation-specific Western blot analysis. Overall, we have demonstrated that hamster-specific kinome arrays are a robust tool for characterizing the species-specific molecular host response in a VHF model. Further, our results provide key insights into the hamster host response to PICV infection and will inform future studies with high-consequence VHF pathogens.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/mcp.M114.045443DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4349984PMC
March 2015

Case study: using sequence homology to identify putative phosphorylation sites in an evolutionarily distant species (honeybee).

Brief Bioinform 2015 Sep 6;16(5):820-9. Epub 2014 Nov 6.

The majority of scientific resources are devoted to studying a relatively small number of model species, meaning that the ability to translate knowledge across species is of considerable importance. Obtaining species-specific knowledge enables targeted investigations of the biology and pathobiology of a particular species, and facilitates comparative analyses. Phosphorylation is the most widespread posttranslational modification in eukaryotes, and although many phosphorylation sites have been experimentally identified for some species, little or no data are available for others. Using the honeybee as a test organism, this case study illustrates the process of using protein sequence homology to identify putative phosphorylation sites in a species of interest using experimentally determined sites from other species. A number of issues associated with this process are examined and discussed. Several databases of experimentally determined phosphorylation sites exist; however, it can be difficult for the nonspecialist to ascertain how their contents compare. Thus, this case study assesses the content and comparability of several phosphorylation site databases. Additional issues examined include the efficacy of homology-based phosphorylation site prediction, the impact of the level of evolutionary relatedness between species in making these predictions, the ability to translate knowledge of phosphorylation sites across large evolutionary distances and the criteria that should be used in selecting probable phosphorylation sites in the species of interest. Although focusing on phosphorylation, the issues discussed here also apply to the homology-based cross-species prediction of other posttranslational modifications, as well as to sequence motifs in general.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/bib/bbu040DOI Listing
September 2015

A better sequence-read simulator program for metagenomics.

BMC Bioinformatics 2014 10;15 Suppl 9:S14. Epub 2014 Sep 10.

Background: There are many programs available for generating simulated whole-genome shotgun sequence reads. The data generated by many of these programs follow predefined models, which limits their use to the authors' original intentions. For example, many models assume that read lengths follow a uniform or normal distribution. Other programs generate models from actual sequencing data, but are limited to reads from single-genome studies. To our knowledge, there are no programs that allow a user to generate simulated data following non-parametric read-length distributions and quality profiles based on empirically-derived information from metagenomics sequencing data.

Results: We present BEAR (Better Emulation for Artificial Reads), a program that uses a machine-learning approach to generate reads with lengths and quality values that closely match empirically-derived distributions. BEAR can emulate reads from various sequencing platforms, including Illumina, 454, and Ion Torrent. BEAR requires minimal user input, as it automatically determines appropriate parameter settings from user-supplied data. BEAR also uses a unique method for deriving run-specific error rates, and extracts useful statistics from the metagenomic data itself, such as quality-error models. Many existing simulators are specific to a particular sequencing technology; however, BEAR is not restricted in this way. Because of its flexibility, BEAR is particularly useful for emulating the behaviour of technologies like Ion Torrent, for which no dedicated sequencing simulators are currently available. BEAR is also the first metagenomic sequencing simulator program that automates the process of generating abundances, which can be an arduous task.

Conclusions: BEAR is useful for evaluating data processing tools in genomics. It has many advantages over existing comparable software, such as generating more realistic reads and being independent of sequencing technology, and has features particularly useful for metagenomics work.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/1471-2105-15-S9-S14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4168713PMC
December 2014

Equivalent input produces different output in the UniFrac significance test.

BMC Bioinformatics 2014 Aug 13;15:278. Epub 2014 Aug 13.

Department of Computer Science, University of Saskatchewan, 176 Thorvaldson Bldg, 110 Science Place, Saskatoon, Canada.

Background: UniFrac is a well-known tool for comparing microbial communities and assessing statistically significant differences between communities. In this paper we identify a discrepancy in the UniFrac methodology that causes semantically equivalent inputs to produce different outputs in tests of statistical significance.

Results: The phylogenetic trees that are input into UniFrac may or may not contain abundance counts. An isomorphic transform can be defined that will convert trees between these two formats without altering the semantic meaning of the trees. UniFrac produces different outputs for these equivalent forms of the same input tree. This is illustrated using metagenomics data from a lake sediment study.

Conclusions: Results from the UniFrac tool can vary greatly for the same input depending on the arbitrary choice of input format. Practitioners should be aware of this issue and use the tool with caution to ensure consistency and validity in their analyses. We provide a script to transform inputs between equivalent formats to help researchers achieve this consistency.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/1471-2105-15-278DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4141948PMC
August 2014

Ebola virus modulates transforming growth factor β signaling and cellular markers of mesenchyme-like transition in hepatocytes.

J Virol 2014 Sep 18;88(17):9877-92. Epub 2014 Jun 18.

Emerging Viral Pathogens Section, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA Integrated Research Facility, Division of Clinical Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Frederick, Maryland, USA.

Unlabelled: Ebola virus (EBOV) causes a severe hemorrhagic disease in humans and nonhuman primates, with a median case fatality rate of 78.4%. Although EBOV is considered a public health concern, there is a relative paucity of information regarding the modulation of the functional host response during infection. We employed temporal kinome analysis to investigate the relative early, intermediate, and late host kinome responses to EBOV infection in human hepatocytes. Pathway overrepresentation analysis and functional network analysis of kinome data revealed that transforming growth factor (TGF-β)-mediated signaling responses were temporally modulated in response to EBOV infection. Upregulation of TGF-β signaling in the kinome data sets correlated with the upregulation of TGF-β secretion from EBOV-infected cells. Kinase inhibitors targeting TGF-β signaling, or additional cell receptors and downstream signaling pathway intermediates identified from our kinome analysis, also inhibited EBOV replication. Further, the inhibition of select cell signaling intermediates identified from our kinome analysis provided partial protection in a lethal model of EBOV infection. To gain perspective on the cellular consequence of TGF-β signaling modulation during EBOV infection, we assessed cellular markers associated with upregulation of TGF-β signaling. We observed upregulation of matrix metalloproteinase 9, N-cadherin, and fibronectin expression with concomitant reductions in the expression of E-cadherin and claudin-1, responses that are standard characteristics of an epithelium-to-mesenchyme-like transition. Additionally, we identified phosphorylation events downstream of TGF-β that may contribute to this process. From these observations, we propose a model for a broader role of TGF-β-mediated signaling responses in the pathogenesis of Ebola virus disease.

Importance: Ebola virus (EBOV), formerly Zaire ebolavirus, causes a severe hemorrhagic disease in humans and nonhuman primates and is the most lethal Ebola virus species, with case fatality rates of up to 90%. Although EBOV is considered a worldwide concern, many questions remain regarding EBOV molecular pathogenesis. As it is appreciated that many cellular processes are regulated through kinase-mediated phosphorylation events, we employed temporal kinome analysis to investigate the functional responses of human hepatocytes to EBOV infection. Administration of kinase inhibitors targeting signaling pathway intermediates identified in our kinome analysis inhibited viral replication in vitro and reduced EBOV pathogenesis in vivo. Further analysis of our data also demonstrated that EBOV infection modulated TGF-β-mediated signaling responses and promoted "mesenchyme-like" phenotypic changes. Taken together, these results demonstrated that EBOV infection specifically modulates TGF-β-mediated signaling responses in epithelial cells and may have broader implications in EBOV pathogenesis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.01410-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136307PMC
September 2014

Identification of developmentally-specific kinotypes and mechanisms of Varroa mite resistance through whole-organism, kinome analysis of honeybee.

Front Genet 2014 21;5:139. Epub 2014 May 21.

Vaccine and Infectious Disease Organization, University of Saskatchewan Saskatoon, SK, Canada ; Department of Biochemistry, University of Saskatchewan Saskatoon, SK, Canada.

Recent investigations associate Varroa destructor (Mesostigmata: Varroidae) parasitism and its associated pathogens and agricultural pesticides with negative effects on colony health, resulting in sporadic global declines in domestic honeybee (Apis mellifera) populations. These events have motivated efforts to develop research tools that can offer insight into the causes of declining bee health as well as identify biomarkers to guide breeding programs. Here we report the development of a bee-specific peptide array for characterizing global cellular kinase activity in whole bee extracts. The arrays reveal distinct, developmentally-specific signaling profiles between bees with differential susceptibility to infestation by Varroa mites. Gene ontology analysis of the differentially phosphorylated peptides indicates that the differential susceptibility to Varroa mite infestation does not reflect compromised immunity; rather, there is evidence for mite-mediated immune suppression within the susceptible phenotype that may reduce the ability of these bees to counter secondary viral infections. This hypothesis is supported by the demonstration of more diverse viral infections in mite-infested, susceptible adult bees. The bee-specific peptide arrays are an effective tool for understanding the molecular basis of this complex phenotype as well as for the discovery and utilization of phosphorylation biomarkers for breeding programs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fgene.2014.00139DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4033134PMC
June 2014

A Comparison of the Chicken and Turkey Proteomes and Phosphoproteomes in the Development of Poultry-Specific Immuno-Metabolism Kinome Peptide Arrays.

Front Vet Sci 2014 13;1:22. Epub 2014 Nov 13.

United States Department of Agriculture, Southern Plains Agricultural Research Center (SPARC), Agricultural Research Service , College Station, TX , USA.

The use of species-specific peptide arrays for the study of animal kinomes has a proven track record of success. This technique has been used in a variety of species for the study of host-pathogen interactions and metabolism. Species-specific peptide arrays have been designed previously for use with chicken but a turkey array has never been attempted. In addition, arrays designed around individual cellular functions have been designed and utilized, but cross-function immuno-metabolic arrays have not been considered previously. Antecedent to designing separate chicken and turkey immuno-metabolic kinome peptide arrays, we show that while the chicken and turkey genomes are quite similar, the two species are much more distinct at the proteome and phosphoproteome levels. Despite a genome identity of approximately 90%, we observe that only 83% of chicken and turkey orthologous proteins display sequence matches between the two species. Further, less than 70% of kinase recognition target sequences are exact matches between chicken and turkey. Thus, our analysis shows that, at the proteome and kinome level, these two species must be considered separately in the design of novel peptide arrays. Our ultimate array design covers numerous immune and metabolic processes including innate and adaptive immunity, inflammatory responses, carbohydrate, protein, and fat metabolism, and response to hormones. We have shown the proteomic and phosphoproteomic diversity of chicken and turkey and have designed a valuable research tool for the study of immuno-metabolism within these two species.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fvets.2014.00022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4668846PMC
December 2015