DOCSLIB.ORG

Genome-Scale Profiling Reveals a Subset of Genes Regulated by DNA

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Genome-Scale Profiling Reveals a Subset of Genes Regulated by DNA Genes and Immunity (2012) 13, 388–398 & 2012 Macmillan Publishers Limited All rights reserved 1466-4879/12 www.nature.com/gene ORIGINAL ARTICLE Genome-scale profiling reveals a subset of genes regulated by DNA methylation that program somatic T-cell phenotypes in humans D Martino1,2, J Maksimovic1, J-HE Joo1, SL Prescott2,3 and R Saffery1,3 The aim of this study was to investigate the dynamics and relationship between DNA methylation and gene expression during early T-cell development. Mononuclear cells were collected at birth and at 12 months from 60 infants and were either activated with anti-CD3 for 24 h or cultured in media alone, and the CD4 þ T-cell subset purified. DNA and RNA were co-harvested and DNA methylation was measured in 450 000 CpG sites in parallel with expression measurements taken from 25 000 genes. In unstimulated cells, we found that a subset of 1188 differentially methylated loci were associated with a change in expression in 599 genes (adjusted P valueo0.01, b-fold 40.1). These genes were enriched in reprogramming regions of the genome known to control pluripotency. In contrast, over 630 genes were induced following low-level T-cell activation, but this was not associated with any significant change in DNA methylation. We conclude that DNA methylation is dynamic during early T-cell development, and has a role in the consolidation of T-cell-specific gene expression. During the early phase of clonal expansion, DNA methylation is stable and therefore appears to be of limited importance in short-term T-cell responsiveness. Genes and Immunity (2012) 13, 388–398; doi:10.1038/gene.2012.7; published online 12 April 2012 Keywords: T-cell epigenetics; immune epigenetics; DNA methylation; gene expression; T-cell development; reprogramming differentially methylated region INTRODUCTION cytokines, thus remaining ‘poised’ for commitment.5,14 Although Shortly after birth, there are rapid phenotypic and functional these observations have contributed to our understanding of the changes in both innate and adaptive immunity. This critical period mechanisms that govern T-cell plasticity and lineage commitment, of early immune programming is not only important for they have largely been observed under highly polarizing 15–18 establishing normal patterns of immunity, but also represents a experimental conditions, and studied for a restricted period of heightened susceptibility to various immune disorders. number of important cytokine gene loci. Therefore, a more In the adaptive immune compartment, there is a developmental complete picture of the epigenetic processes that govern normal transition as ‘less mature’ T-cells emerging from the thymus T-cell development is warranted, which has only recently been undergo maturation in the periphery.1 This transition becomes possible with the advent of genome-wide technologies. apparent over the first year of life;2 however, the molecular In the current study, we investigated the role of DNA processes that drive this are poorly understood. Understanding methylation and its association with patterns of gene expression these processes is critical, as disruption in these pathways may under two scenarios: (1) during the steady-state development of alter the normal course of T-cell development, and potentially naive CD4 þ T-cells shortly after birth; (2) following the activation program susceptibility may lead to a range of allergic and of T-cells during entry into the cell cycle. Our data provide insights autoimmune diseases.3,4 into the molecular pathways of early T-cell programming, and Epigenetic modifications are likely to mediate early develop- characterize developmental pathways potentially susceptible to mental changes in T-cells, because these modifications are known disruption through early-life environmental exposures. to have a well-defined role in determining both the diversity and plasticity of T-helper cell phenotypes.5–7 Variability in DNA methylation levels and histone modification profiles establishes RESULTS 8–11 active or repressive states of transcription at key cytokine loci. Combined genome-wide DNA methylation and gene expression In differentiated CD4 þ T-cells, these mechanisms are responsible analysis reveals a dynamic genomic program during steady-state for the somatic heritability of differentiated T-cell states, and T-cell development are described in close association with the acquisition of Neonatal CD4 þ T-cells are phenotypically and functionally effector phenotypes, and specialized patterns of cytokine unique compared with later ages. To investigate the epigenetic 11–13 gene expression. In undifferentiated (naive) T-cells, DNA differences between these cell types, we compared DNA methylation marks maintain the plasticity of CD4 þ T-cells, methylation and gene expression in neonatal CD4 þ cells with because these cells express low levels of a broad range of their 12-month counterparts under two conditions. The 1Cancer, Disease and Developmental Epigenetics, Murdoch Children’s Research Institute, Royal Melbourne Hospital, Parkville, Victoria, Australia and 2School of Paediatrics and Child Health, University of Western Australia, Perth, Western Australia, Australia. Correspondence: Dr R Saffery, Cancer, Disease and Developmental Epigenetics, Murdoch Children’s Research Institute, Royal Melbourne Hospital, Flemington Road, Parkville, Victoria 3052, Australia. E-mail: [email protected] 3Equal senior authors. Received 24 January 2012; revised 28 February 2012; accepted 29 February 2012; published online 12 April 2012 Epigenetic programming of CD4 þ T-cell phenotypes during early life D Martino et al 389 Figure 1. Experimental methodology and validation of in vitro protocol. (a) Experimental approach used in this study. (b) Experimental design for methylation comparisons between fresh and 24-h cultured CD4 þ cells. (c) Matrix scatterplot of fresh versus 24-h cord blood and adult blood samples. The figure shows scatterplot comparisons between all samples. MDS, multidimensional scaling plot. experimental strategy is outlined in Figure 1a. Briefly, mono- 1471 probes (31.9%) had no associated gene annotation and were nuclear cells collected from the same infants at birth and located either in intergenic regions or in regions occupied by two 12months were cultured with and without anti-CD3 and IL-2 for or more refseq transcripts. A total of 3224 CpGs (70%) showed 24 h, in a 371 incubator maintained at 5% CO2. After this time, increased methylation between neonatal and 12-month CD4 þ media supernatants were reserved for cytokine analysis and cells, and 1383 CpGs (30%) showed reduced methylation, CD4 þ cells were purified by magnetic bead sorting. DNA and indicative of dynamic changes in DNA methylation during early total RNA were co-harvested for microarray analysis. Studies of development. normal development were conducted in resting (unstimulated) Unsupervised sample clustering based on the 4607 differentially cells, and studies of T-cell activation were conducted in anti-CD3- methylated probes correctly discriminated neonatal from treated cultures. 12-month samples (Supplementary Figure 1). We performed To validate the experimental approach, we first needed to ontology enrichment analysis on the gene-associated probes determine whether T-cells rested in culture for 24 h undergo any and identified terms associated with gene expression, RNA non-physiological changes in DNA methylation that may compli- polymerase II activity and transcription. Alongside these, we also cate data interpretation. To address this, we performed a small observed a host of developmental terms, including cell and tissue pilot experiment detailed in Materials and methods (Figure 1b). morphogenesis, mesenchymal differentiation, skeletal muscle, and We compared freshly thawed T cells with cultured Tcells genome- olfactory and neuronal development (Supplementary Table 1). wide, using exploratory techniques and probe-wise tests for Examples of genes associated with these terms include the differential methylation (adjusted P-valueo0.05 and b-fold myosins (MYO1D, MYOIC), myosin light-chain kinases (MYLK), change40.1). We found no changes in DNA methylation profiles olfactory receptor family members (ORS1E2, OR4D2) and neuronal between fresh and cultured CD4 þ cells, for neonatal or adult peptides (NRP2, NRTN). Epigenetic changes at these loci are likely samples (Figure 1c). This demonstrated that short-term cell culture to reflect the developmental control of gene expression during does not distort the physiological patterns of genomic lineage commitment.19,20 Several immunological terms were also methylation. enriched in the list of differentially methylated genes, and these To gain a broad picture of the extent to which neonatal CD4 þ included antigen processing and presentation, immune response, cells are developmentally different from their 12-month counter- leukocyte activation, protein kinase signaling, TGFb signaling and parts, we compared unstimulated cultures. Probe-wise compar- MAPK signaling (Table 1). isons of DNA methylation between neonatal and 12-month CD4 þ In the gene expression data set, we observed 986 probes that cells identified a total of 4607 differentially methylated CpG sites varied significantly between unstimulated neonatal and 12-month (adjusted P valueo0.01 and b-fold change40.10), of which 3136 T cells (adjusted P-valueo0.01, b-fold change42). This consti- sites mapped to 1826 unique genes (Figure 2a). The remaining tuted 287 (29%) upregulated probes and 699 (71%) & 2012
Load more

Recommended publications
  • Genetic Analysis of Retinopathy in Type 1 Diabetes
    Genetic Analysis of Retinopathy in Type 1 Diabetes by Sayed Mohsen Hosseini A thesis submitted in conformity with the requirements for the degree of Doctor of Philosophy Institute of Medical Science University of Toronto © Copyright by S. Mohsen Hosseini 2014 Genetic Analysis of Retinopathy in Type 1 Diabetes Sayed Mohsen Hosseini Doctor of Philosophy Institute of Medical Science University of Toronto 2014 Abstract Diabetic retinopathy (DR) is a leading cause of blindness worldwide. Several lines of evidence suggest a genetic contribution to the risk of DR; however, no genetic variant has shown convincing association with DR in genome-wide association studies (GWAS). To identify common polymorphisms associated with DR, meta-GWAS were performed in three type 1 diabetes cohorts of White subjects: Diabetes Complications and Control Trial (DCCT, n=1304), Wisconsin Epidemiologic Study of Diabetic Retinopathy (WESDR, n=603) and Renin-Angiotensin System Study (RASS, n=239). Severe (SDR) and mild (MDR) retinopathy outcomes were defined based on repeated fundus photographs in each study graded for retinopathy severity on the Early Treatment Diabetic Retinopathy Study (ETDRS) scale. Multivariable models accounted for glycemia (measured by A1C), diabetes duration and other relevant covariates in the association analyses of additive genotypes with SDR and MDR. Fixed-effects meta- analysis was used to combine the results of GWAS performed separately in WESDR, ii RASS and subgroups of DCCT, defined by cohort and treatment group. Top association signals were prioritized for replication, based on previous supporting knowledge from the literature, followed by replication in three independent white T1D studies: Genesis-GeneDiab (n=502), Steno (n=936) and FinnDiane (n=2194).
    [Show full text]
  • Browsing Genes and Genomes with Ensembl
    The Bioinformatics Roadshow Copenhagen, Denmark 16 & 17 June 2011 BROWSING GENES AND GENOMES WITH ENSEMBL EXERCISES AND ANSWERS Note: These exercises are based on Ensembl version 62 (13 April 2011). After in future a new version has gone live, version 62 will still be available at http://e62.ensembl.org. If your answer doesn’t correspond with the given answer, please consult the instructor. ______________________________________________________________ BROWSER ______________________________________________________________ Exercise 1 – Exploring a gene (a) Find the human F9 (Coagulation factor IX) gene. On which chromosome and which strand of the genome is this gene located? How many transcripts (splice variants) have been annotated for it? (b) What is the longest transcript? How long is the protein it encodes? How many exons does it have? Are any of the exons completely or partially untranslated? (c) Have a look at the external references for ENST00000218099. What is the function of F9? (d) Is it possible to monitor expression of ENST00000218099 with the CodeLink microarray? If so, can it also be used to monitor expression of the other two transcripts? (e) In which part (i.e. the N-terminal or C-terminal half) of the protein encoded by ENST00000218099 does its peptidase activity, responsible for the cleavage of factor X to yield its active form factor Xa, reside? (f) How many non-synonymous variants have been discovered for the protein encoded by ENST00000218099? (g) Is there a mouse ortholog predicted for the human F9 gene? (h) On which cytogenetic band and on which contig is the F9 gene located? Is there a BAC clone that contains the complete F9 gene? (i) If you have yourself a gene of interest, explore what information Ensembl displays about it! ______________________________________________________________ Answer (a) Go to the Ensembl homepage (http://www.ensembl.org).
    [Show full text]
  • Frequent Expression Loss of Inter-Alpha-Trypsin Inhibitor Heavy Chain (ITIH) Genes in Multiple Human Solid Tumors: a Systematic Expression Analysis
    Hamm, A; Veeck, J; Bektas, N; Wild, P J; Hartmann, A; Heindrichs, U; Kristiansen, G; Werbowetski-Ogilvie, T; Del Maestro, R; Knuechel, R; Dahl, E (2008). Frequent expression loss of Inter-alpha-trypsin inhibitor heavy chain (ITIH) genes in multiple human solid tumors: a systematic expression analysis. BMC Cancer, 8:25. Postprint available at: http://www.zora.uzh.ch University of Zurich Posted at the Zurich Open Repository and Archive, University of Zurich. Zurich Open Repository and Archive http://www.zora.uzh.ch Originally published at: BMC Cancer 2008, 8:25. Winterthurerstr. 190 CH-8057 Zurich http://www.zora.uzh.ch Year: 2008 Frequent expression loss of Inter-alpha-trypsin inhibitor heavy chain (ITIH) genes in multiple human solid tumors: a systematic expression analysis Hamm, A; Veeck, J; Bektas, N; Wild, P J; Hartmann, A; Heindrichs, U; Kristiansen, G; Werbowetski-Ogilvie, T; Del Maestro, R; Knuechel, R; Dahl, E Hamm, A; Veeck, J; Bektas, N; Wild, P J; Hartmann, A; Heindrichs, U; Kristiansen, G; Werbowetski-Ogilvie, T; Del Maestro, R; Knuechel, R; Dahl, E (2008). Frequent expression loss of Inter-alpha-trypsin inhibitor heavy chain (ITIH) genes in multiple human solid tumors: a systematic expression analysis. BMC Cancer, 8:25. Postprint available at: http://www.zora.uzh.ch Posted at the Zurich Open Repository and Archive, University of Zurich. http://www.zora.uzh.ch Originally published at: BMC Cancer 2008, 8:25. Frequent expression loss of Inter-alpha-trypsin inhibitor heavy chain (ITIH) genes in multiple human solid tumors: a systematic expression analysis Abstract BACKGROUND: The inter-alpha-trypsin inhibitors (ITI) are a family of plasma protease inhibitors, assembled from a light chain - bikunin, encoded by AMBP - and five homologous heavy chains (encoded by ITIH1, ITIH2, ITIH3, ITIH4, and ITIH5), contributing to extracellular matrix stability by covalent linkage to hyaluronan.
    [Show full text]
  • A Novel Computational Algorithm for Predicting Immune Cell Types Using Single-Cell RNA Sequencing Data
    A novel computational algorithm for predicting immune cell types using single-cell RNA sequencing data By Shuo Jia A hesis submitted to the Faculty of Graduate Studies of The University of Manitoba n partial fulfillment of the requirements of the degree of MASTER OF SCIENCE Department of Biochemistry and Medical Genetics University of Manitoba Winnipeg, Manitoba, Canada Copyright © 2020 by Shuo Jia Abstract Background: Cells from our immune system detect and kill pathogens to protect our body against many diseases. However, current methods for determining cell types have some major limitations, such as being time-consuming and with low throughput rate, etc. These problems stack up and hinder the deep exploration of cellular heterogeneity. Immune cells that are associated with cancer tissues play a critical role in revealing the stages of tumor development. Identifying the immune composition within tumor microenvironments in a timely manner will be helpful to improve clinical prognosis and therapeutic management for cancer. Single-cell RNA sequencing (scRNA-seq), an RNA sequencing (RNA-seq) technique that focuses on a single cell level, has provided us with the ability to conduct cell type classification. Although unsupervised clustering approaches are the major methods for analyzing scRNA-seq datasets, their results vary among studies with different input parameters and sizes. However, in supervised machine learning methods, information loss and low prediction accuracy are the key limitations. Methods and Results: Genes in the human genome align to chromosomes in a particular order. Hence, we hypothesize incorporating this information into our model will potentially improve the cell type classification performance. In order to utilize gene positional information, we introduce chromosome-based neural network, namely ChrNet, a novel chromosome-specific re-trainable supervised learning method based on a one-dimensional 1 convolutional neural network (1D-CNN).
    [Show full text]
  • Transcriptomic Analysis of Native Versus Cultured Human and Mouse Dorsal Root Ganglia Focused on Pharmacological Targets Short
    bioRxiv preprint doi: https://doi.org/10.1101/766865; this version posted September 12, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-ND 4.0 International license. Transcriptomic analysis of native versus cultured human and mouse dorsal root ganglia focused on pharmacological targets Short title: Comparative transcriptomics of acutely dissected versus cultured DRGs Andi Wangzhou1, Lisa A. McIlvried2, Candler Paige1, Paulino Barragan-Iglesias1, Carolyn A. Guzman1, Gregory Dussor1, Pradipta R. Ray1,#, Robert W. Gereau IV2, # and Theodore J. Price1, # 1The University of Texas at Dallas, School of Behavioral and Brain Sciences and Center for Advanced Pain Studies, 800 W Campbell Rd. Richardson, TX, 75080, USA 2Washington University Pain Center and Department of Anesthesiology, Washington University School of Medicine # corresponding authors [email protected], [email protected] and [email protected] Funding: NIH grants T32DA007261 (LM); NS065926 and NS102161 (TJP); NS106953 and NS042595 (RWG). The authors declare no conflicts of interest Author Contributions Conceived of the Project: PRR, RWG IV and TJP Performed Experiments: AW, LAM, CP, PB-I Supervised Experiments: GD, RWG IV, TJP Analyzed Data: AW, LAM, CP, CAG, PRR Supervised Bioinformatics Analysis: PRR Drew Figures: AW, PRR Wrote and Edited Manuscript: AW, LAM, CP, GD, PRR, RWG IV, TJP All authors approved the final version of the manuscript. 1 bioRxiv preprint doi: https://doi.org/10.1101/766865; this version posted September 12, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.
    [Show full text]
  • 4-6 Weeks Old Female C57BL/6 Mice Obtained from Jackson Labs Were Used for Cell Isolation
    Methods Mice: 4-6 weeks old female C57BL/6 mice obtained from Jackson labs were used for cell isolation. Female Foxp3-IRES-GFP reporter mice (1), backcrossed to B6/C57 background for 10 generations, were used for the isolation of naïve CD4 and naïve CD8 cells for the RNAseq experiments. The mice were housed in pathogen-free animal facility in the La Jolla Institute for Allergy and Immunology and were used according to protocols approved by the Institutional Animal Care and use Committee. Preparation of cells: Subsets of thymocytes were isolated by cell sorting as previously described (2), after cell surface staining using CD4 (GK1.5), CD8 (53-6.7), CD3ε (145- 2C11), CD24 (M1/69) (all from Biolegend). DP cells: CD4+CD8 int/hi; CD4 SP cells: CD4CD3 hi, CD24 int/lo; CD8 SP cells: CD8 int/hi CD4 CD3 hi, CD24 int/lo (Fig S2). Peripheral subsets were isolated after pooling spleen and lymph nodes. T cells were enriched by negative isolation using Dynabeads (Dynabeads untouched mouse T cells, 11413D, Invitrogen). After surface staining for CD4 (GK1.5), CD8 (53-6.7), CD62L (MEL-14), CD25 (PC61) and CD44 (IM7), naïve CD4+CD62L hiCD25-CD44lo and naïve CD8+CD62L hiCD25-CD44lo were obtained by sorting (BD FACS Aria). Additionally, for the RNAseq experiments, CD4 and CD8 naïve cells were isolated by sorting T cells from the Foxp3- IRES-GFP mice: CD4+CD62LhiCD25–CD44lo GFP(FOXP3)– and CD8+CD62LhiCD25– CD44lo GFP(FOXP3)– (antibodies were from Biolegend). In some cases, naïve CD4 cells were cultured in vitro under Th1 or Th2 polarizing conditions (3, 4).
    [Show full text]
  • Prox1regulates the Subtype-Specific Development of Caudal Ganglionic
    The Journal of Neuroscience, September 16, 2015 • 35(37):12869–12889 • 12869 Development/Plasticity/Repair Prox1 Regulates the Subtype-Specific Development of Caudal Ganglionic Eminence-Derived GABAergic Cortical Interneurons X Goichi Miyoshi,1 Allison Young,1 Timothy Petros,1 Theofanis Karayannis,1 Melissa McKenzie Chang,1 Alfonso Lavado,2 Tomohiko Iwano,3 Miho Nakajima,4 Hiroki Taniguchi,5 Z. Josh Huang,5 XNathaniel Heintz,4 Guillermo Oliver,2 Fumio Matsuzaki,3 Robert P. Machold,1 and Gord Fishell1 1Department of Neuroscience and Physiology, NYU Neuroscience Institute, Smilow Research Center, New York University School of Medicine, New York, New York 10016, 2Department of Genetics & Tumor Cell Biology, St. Jude Children’s Research Hospital, Memphis, Tennessee 38105, 3Laboratory for Cell Asymmetry, RIKEN Center for Developmental Biology, Kobe 650-0047, Japan, 4Laboratory of Molecular Biology, Howard Hughes Medical Institute, GENSAT Project, The Rockefeller University, New York, New York 10065, and 5Cold Spring Harbor Laboratory, Cold Spring Harbor, New York 11724 Neurogliaform (RELNϩ) and bipolar (VIPϩ) GABAergic interneurons of the mammalian cerebral cortex provide critical inhibition locally within the superficial layers. While these subtypes are known to originate from the embryonic caudal ganglionic eminence (CGE), the specific genetic programs that direct their positioning, maturation, and integration into the cortical network have not been eluci- dated. Here, we report that in mice expression of the transcription factor Prox1 is selectively maintained in postmitotic CGE-derived cortical interneuron precursors and that loss of Prox1 impairs the integration of these cells into superficial layers. Moreover, Prox1 differentially regulates the postnatal maturation of each specific subtype originating from the CGE (RELN, Calb2/VIP, and VIP).
    [Show full text]
  • Meta-Analyses of Expression Profiling Data in the Postmortem
    META-ANALYSES OF EXPRESSION PROFILING DATA IN THE POSTMORTEM HUMAN BRAIN by Meeta Mistry B.Sc., McMaster University, 2005 A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY in THE FACULTY OF GRADUATE STUDIES (Bioinformatics) THE UNIVERSITY OF BRITISH COLUMBIA (Vancouver) July 2012 © Meeta Mistry, 2012 Abstract Schizophrenia is a severe psychiatric illness for which the precise etiology remains unknown. Studies using postmortem human brain have become increasingly important in schizophrenia research, providing an opportunity to directly investigate the diseased brain tissue. Gene expression profiling technologies have been used by a number of groups to explore the postmortem human brain and seek genes which show changes in expression correlated with schizophrenia. While this has been a valuable means of generating hypotheses, there is a general lack of consensus in the findings across studies. Expression profiling of postmortem human brain tissue is difficult due to the effect of various factors that can confound the data. The first aim of this thesis was to use control postmortem human cortex for identification of expression changes associated with several factors, specifically: age, sex, brain pH and postmortem interval. I conducted a meta-analysis across the control arm of eleven microarray datasets (representing over 400 subjects), and identified a signature of genes associated with each factor. These genes provide critical information towards the identification of problematic genes when investigating postmortem human brain in schizophrenia and other neuropsychiatric illnesses. The second aim of this thesis was to evaluate gene expression patterns in the prefrontal cortex associated with schizophrenia by exploring two methods of analysis: differential expression and coexpression.
    [Show full text]
  • Supplementary Table 1: Adhesion Genes Data Set
    Supplementary Table 1: Adhesion genes data set PROBE Entrez Gene ID Celera Gene ID Gene_Symbol Gene_Name 160832 1 hCG201364.3 A1BG alpha-1-B glycoprotein 223658 1 hCG201364.3 A1BG alpha-1-B glycoprotein 212988 102 hCG40040.3 ADAM10 ADAM metallopeptidase domain 10 133411 4185 hCG28232.2 ADAM11 ADAM metallopeptidase domain 11 110695 8038 hCG40937.4 ADAM12 ADAM metallopeptidase domain 12 (meltrin alpha) 195222 8038 hCG40937.4 ADAM12 ADAM metallopeptidase domain 12 (meltrin alpha) 165344 8751 hCG20021.3 ADAM15 ADAM metallopeptidase domain 15 (metargidin) 189065 6868 null ADAM17 ADAM metallopeptidase domain 17 (tumor necrosis factor, alpha, converting enzyme) 108119 8728 hCG15398.4 ADAM19 ADAM metallopeptidase domain 19 (meltrin beta) 117763 8748 hCG20675.3 ADAM20 ADAM metallopeptidase domain 20 126448 8747 hCG1785634.2 ADAM21 ADAM metallopeptidase domain 21 208981 8747 hCG1785634.2|hCG2042897 ADAM21 ADAM metallopeptidase domain 21 180903 53616 hCG17212.4 ADAM22 ADAM metallopeptidase domain 22 177272 8745 hCG1811623.1 ADAM23 ADAM metallopeptidase domain 23 102384 10863 hCG1818505.1 ADAM28 ADAM metallopeptidase domain 28 119968 11086 hCG1786734.2 ADAM29 ADAM metallopeptidase domain 29 205542 11085 hCG1997196.1 ADAM30 ADAM metallopeptidase domain 30 148417 80332 hCG39255.4 ADAM33 ADAM metallopeptidase domain 33 140492 8756 hCG1789002.2 ADAM7 ADAM metallopeptidase domain 7 122603 101 hCG1816947.1 ADAM8 ADAM metallopeptidase domain 8 183965 8754 hCG1996391 ADAM9 ADAM metallopeptidase domain 9 (meltrin gamma) 129974 27299 hCG15447.3 ADAMDEC1 ADAM-like,
    [Show full text]
  • A CRISPR-Based Screen for Hedgehog Signaling Provides Insights Into Ciliary Function and Ciliopathies
    ARTICLES https://doi.org/10.1038/s41588-018-0054-7 A CRISPR-based screen for Hedgehog signaling provides insights into ciliary function and ciliopathies David K. Breslow 1,2,7*, Sascha Hoogendoorn 3,7, Adam R. Kopp2, David W. Morgens4, Brandon K. Vu2, Margaret C. Kennedy1, Kyuho Han4, Amy Li4, Gaelen T. Hess4, Michael C. Bassik4, James K. Chen 3,5* and Maxence V. Nachury 2,6* Primary cilia organize Hedgehog signaling and shape embryonic development, and their dysregulation is the unifying cause of ciliopathies. We conducted a functional genomic screen for Hedgehog signaling by engineering antibiotic-based selection of Hedgehog-responsive cells and applying genome-wide CRISPR-mediated gene disruption. The screen can robustly identify factors required for ciliary signaling with few false positives or false negatives. Characterization of hit genes uncovered novel components of several ciliary structures, including a protein complex that contains δ -tubulin and ε -tubulin and is required for centriole maintenance. The screen also provides an unbiased tool for classifying ciliopathies and showed that many congenital heart disorders are caused by loss of ciliary signaling. Collectively, our study enables a systematic analysis of ciliary function and of ciliopathies, and also defines a versatile platform for dissecting signaling pathways through CRISPR-based screening. he primary cilium is a surface-exposed microtubule-based approach. Indeed, most studies to date have searched for genes that compartment that serves as an organizing center for diverse either intrinsically affect cell growth or affect sensitivity to applied Tsignaling pathways1–3. Mutations affecting cilia cause ciliopa- perturbations16–23. thies, a group of developmental disorders including Joubert syn- Here, we engineered a Hh-pathway-sensitive reporter to enable drome, Meckel syndrome (MKS), nephronophthisis (NPHP), and an antibiotic-based selection platform.
    [Show full text]
  • Fine-Scale Survey of X Chromosome Copy Number Variants and Indels Underlying Intellectual Disability
    ARTICLE Fine-Scale Survey of X Chromosome Copy Number Variants and Indels Underlying Intellectual Disability Annabel C. Whibley,1 Vincent Plagnol,1,2 Patrick S. Tarpey,3 Fatima Abidi,4 Tod Fullston,5,6 Maja K. Choma,1 Catherine A. Boucher,1 Lorraine Shepherd,1 Lionel Willatt,7 Georgina Parkin,7 Raffaella Smith,3 P. Andrew Futreal,3 Marie Shaw,8 Jackie Boyle,9 Andrea Licata,4 Cindy Skinner,4 Roger E. Stevenson,4 Gillian Turner,9 Michael Field,9 Anna Hackett,9 Charles E. Schwartz,4 Jozef Gecz,5,6,8 Michael R. Stratton,3 and F. Lucy Raymond1,* Copy number variants and indels in 251 families with evidence of X-linked intellectual disability (XLID) were investigated by array comparative genomic hybridization on a high-density oligonucleotide X chromosome array platform. We identified pathogenic copy number variants in 10% of families, with mutations ranging from 2 kb to 11 Mb in size. The challenge of assessing causality was facilitated by prior knowledge of XLID-associated genes and the ability to test for cosegregation of variants with disease through extended pedigrees. Fine-scale analysis of rare variants in XLID families leads us to propose four additional genes, PTCHD1, WDR13, FAAH2, and GSPT2,as candidates for XLID causation and the identification of further deletions and duplications affecting X chromosome genes but without apparent disease consequences. Breakpoints of pathogenic variants were characterized to provide insight into the underlying mutational mechanisms and indicated a predominance of mitotic rather than meiotic events. By effectively bridging the gap between karyotype-level investigations and X chromosome exon resequencing, this study informs discussion of alternative mutational mechanisms, such as non- coding variants and non-X-linked disease, which might explain the shortfall of mutation yield in the well-characterized International Genetics of Learning Disability (IGOLD) cohort, where currently disease remains unexplained in two-thirds of families.
    [Show full text]
  • ITRAQ-Based Quantitative Proteomic Analysis of Processed Euphorbia Lathyris L
    Zhang et al. Proteome Science (2018) 16:8 https://doi.org/10.1186/s12953-018-0136-6 RESEARCH Open Access ITRAQ-based quantitative proteomic analysis of processed Euphorbia lathyris L. for reducing the intestinal toxicity Yu Zhang1, Yingzi Wang1*, Shaojing Li2*, Xiuting Zhang1, Wenhua Li1, Shengxiu Luo1, Zhenyang Sun1 and Ruijie Nie1 Abstract Background: Euphorbia lathyris L., a Traditional Chinese medicine (TCM), is commonly used for the treatment of hydropsy, ascites, constipation, amenorrhea, and scabies. Semen Euphorbiae Pulveratum, which is another type of Euphorbia lathyris that is commonly used in TCM practice and is obtained by removing the oil from the seed that is called paozhi, has been known to ease diarrhea. Whereas, the mechanisms of reducing intestinal toxicity have not been clearly investigated yet. Methods: In this study, the isobaric tags for relative and absolute quantitation (iTRAQ) in combination with the liquid chromatography-tandem mass spectrometry (LC-MS/MS) proteomic method was applied to investigate the effects of Euphorbia lathyris L. on the protein expression involved in intestinal metabolism, in order to illustrate the potential attenuated mechanism of Euphorbia lathyris L. processing. Differentially expressed proteins (DEPs) in the intestine after treated with Semen Euphorbiae (SE), Semen Euphorbiae Pulveratum (SEP) and Euphorbiae Factor 1 (EFL1) were identified. The bioinformatics analysis including GO analysis, pathway analysis, and network analysis were done to analyze the key metabolic pathways underlying the attenuation mechanism through protein network in diarrhea. Western blot were performed to validate selected protein and the related pathways. Results: A number of differentially expressed proteins that may be associated with intestinal inflammation were identified.
    [Show full text]