DOCSLIB.ORG

Functional Relevance of Epigenetic Modifications in Inflammatory Bowel Disease

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Functional Relevance of Epigenetic Modifications in Inflammatory Bowel Disease Functional relevance of epigenetic modifications in inflammatory bowel disease Dissertation in fulfilment of the requirements for the degree “Dr. rer. nat.” of the Faculty of Mathematics and Natural Sciences at Kiel University submitted by Zhe Feng Kiel, 2013 First referee: Prof. Dr. Dr. Thomas C.G. Bosch . Second referee: Prof. Dr. Philip Rosenstiel . Date of the oral examination: 17.06.2013 . Approved for publication: ........................................................... Signed: Prof. Dr. Wolfgang J. Duschl, Dean ........................................................... "I only know that I know nothing." (Socrates) To my parents Parts of this thesis have been published in Genome Research. Häsler R, Feng Z, Bäckdahl L, Spehlmann ME, Franke A, Teschendorff A, Rakyan VK, Down TA, Wilson GA, Feber A, Beck S, Schreiber S, Rosenstiel P. A functional methylome map of ulcerative colitis. Genome Res. 2012 Nov;22(11):2130-7. doi: 10.1101/gr.138347.112. Epub 2012 Jul 23. The final version of this publication can be found in the appendix (Page 120-127). Table of Contents Table of Contents 1 INTRODUCTION ..................................................................................................................................... 4 1.1 Ulcerative colitis: a multifactorial induced complex disorder ................................................................. 4 1.1.1 Epidemiology of ulcerative colitis ......................................................................................................... 5 1.1.2 Clinical phenotype of ulcerative colitis ................................................................................................. 6 1.1.3 Epithelial barrier dysfunction and autoimmunity ................................................................................. 7 1.1.4 Immunological aspects of UC ................................................................................................................ 8 1.1.5 Genetic background of UC .................................................................................................................... 9 1.1.6 Environmental factors ......................................................................................................................... 10 1.2 Epigenetic regulation in UC .................................................................................................................... 11 1.2.1 Epigenitic mechanisms ........................................................................................................................ 13 1.2.2 DNA methylation patterns .................................................................................................................. 14 1.2.3 Non-coding RNA and other epigenetic mechanisms ........................................................................... 16 1.2.4 Epigenetics and environment .............................................................................................................. 17 1.2.5 Twin studies ........................................................................................................................................ 18 1.3 Hypothesis and Aims of the study .......................................................................................................... 19 2 MATERIALS AND METHODS .......................................................................................................... 20 2.1 Patient group composition and sample preparation ............................................................................. 20 2.1.1 Patient Recruitment ............................................................................................................................ 20 2.1.2 Samples storage and preparation ....................................................................................................... 21 2.1.3 RNA extraction from biopsy ................................................................................................................ 22 2.1.4 DNA extraction from biopsies ............................................................................................................. 22 2.2 Genome-wide scanning of the transcriptional and methylational profiling ......................................... 24 2.2.1 Genome-wide Transcriptome Profiling (Layer I) ................................................................................. 24 2.2.2 Genome-wide Quantification of Differentially Methylated Regions (Layer II) .................................... 25 1 Table of Contents 2.2.3 Genome-wide Quantification of individual Methylation Variable Positions (Layer III) ...................... 27 2.3 Merging of the transcriptional and methylational data (from Layer I,II & III) ...................................... 29 2.4 Validation of the transcriptional and methylational candidates .......................................................... 29 2.4.1 Validation of differential mRNA expression via Real-Time PCR .......................................................... 30 2.4.2 Validation of differential DNAm via Pyrosequencing .......................................................................... 32 2.5 Gene Ontology analysis .......................................................................................................................... 34 2.6 Chromatin- immunoprecipitation following quantitative real-time PCR ............................................. 34 2.6.1 Preparation of samples and control DNA ........................................................................................... 34 2.6.2 Immunoprecipitation of 5-hydroxymethylcytidine enriched DNA fragments .................................... 35 2.6.3 Examination of 5-hydroxymethylcytidine enrichment via qRT PCR .................................................... 35 3 RESULTS .................................................................................................................................................. 37 3.1 Genome-wide profiling of transcription and DNA methylation in ulcerative colitis ............................ 37 3.1.1 Genome-wide transcriptome profiling (Layer I) ................................................................................. 37 3.1.2 Genome-wide methylome profiling (Layer II and layer III) ................................................................. 37 3.1.3 Genome-wide quantification of individual methylated variable positions (Layer III) ......................... 37 3.1.4 Merging of the transcriptional and methylational data (from Layer I,II & III) .................................... 38 3.1.5 Intra-class correlations of the three-layer dataset ............................................................................. 40 3.1.6 Overview: genomic location of the different methylation in ulcerative colitis .................................. 41 3.1.7 Principle component analysis of the significant candidates ............................................................... 41 3.1.8 Correlation between the expressional and methylational data (in cis) .............................................. 42 3.1.9 Gene ontology (GO) analysis of the disease relevant processes ........................................................ 43 3.2 Validation of selected candidate transcripts under potential epigenetic control ................................ 44 3.3 Potential new finding of the DNA hydroxymethylation in the acute inflammation ............................ 49 4 DISCUSSION ........................................................................................................................................... 52 4.1 Genetics and epigenetics in the pathogenesis of ulcerative colitis ...................................................... 52 2 Table of Contents 4.2 Environment and epigenetics in the pathogenesis of ulcerative colitis ................................................ 56 4.3 Ulcerative colitis: a complex epigenetic disorder .................................................................................. 57 4.4 Genome-wide patterns of hypomethylation are a prominent motive in UC ........................................ 59 4.5 DNA demethylation in chronic and acute colitis .................................................................................... 60 4.5.1 Passive mechanisms of DNA demethylation are potentially absent in UC ......................................... 62 4.5.2 Demethylation in mammalian cells: a multi-step process .................................................................. 62 4.5.3 The APOBEC Family: Mediators of 5-mC or 5-hmC Deamination ....................................................... 63 4.5.4 Reduced BER activity may lead to carcinogenesis of UC ..................................................................... 64 4.6 Concluding remarks and future studies ................................................................................................. 66 5 SUMMARY ............................................................................................................................................... 67 6 ZUSAMMENFASSUNG ........................................................................................................................ 68 7 REFERENCES ......................................................................................................................................... 69 8 APPENDIX ..............................................................................................................................................
Load more

Recommended publications
  • Lower Genomic Stability of Induced Pluripotent Stem Cells Reflects
    Zhang et al. Cancer Commun (2018) 38:49 https://doi.org/10.1186/s40880-018-0313-0 Cancer Communications ORIGINAL ARTICLE Open Access Lower genomic stability of induced pluripotent stem cells refects increased non‑homologous end joining Minjie Zhang1,2†, Liu Wang3†, Ke An1,2†, Jun Cai1, Guochao Li1,2, Caiyun Yang1, Huixian Liu1, Fengxia Du1, Xiao Han1,2, Zilong Zhang1,2, Zitong Zhao1,2, Duanqing Pei4, Yuan Long5, Xin Xie5, Qi Zhou3 and Yingli Sun1* Abstract Background: Induced pluripotent stem cells (iPSCs) and embryonic stem cells (ESCs) share many common features, including similar morphology, gene expression and in vitro diferentiation profles. However, genomic stability is much lower in iPSCs than in ESCs. In the current study, we examined whether changes in DNA damage repair in iPSCs are responsible for their greater tendency towards mutagenesis. Methods: Mouse iPSCs, ESCs and embryonic fbroblasts were exposed to ionizing radiation (4 Gy) to introduce dou- ble-strand DNA breaks. At 4 h later, fdelity of DNA damage repair was assessed using whole-genome re-sequencing. We also analyzed genomic stability in mice derived from iPSCs versus ESCs. Results: In comparison to ESCs and embryonic fbroblasts, iPSCs had lower DNA damage repair capacity, more somatic mutations and short indels after irradiation. iPSCs showed greater non-homologous end joining DNA repair and less homologous recombination DNA repair. Mice derived from iPSCs had lower DNA damage repair capacity than ESC-derived mice as well as C57 control mice. Conclusions: The relatively low genomic stability of iPSCs and their high rate of tumorigenesis in vivo appear to be due, at least in part, to low fdelity of DNA damage repair.
    [Show full text]
  • Abnormal Developmental Control of Replication-Timing Domains in Pediatric Acute Lymphoblastic Leukemia
    Downloaded from genome.cshlp.org on October 1, 2021 - Published by Cold Spring Harbor Laboratory Press Research Abnormal developmental control of replication-timing domains in pediatric acute lymphoblastic leukemia Tyrone Ryba,1,6 Dana Battaglia,1,6 Bill H. Chang,2 James W. Shirley,1 Quinton Buckley,1 Benjamin D. Pope,1 Meenakshi Devidas,3 Brian J. Druker,4,5 and David M. Gilbert1,7 1Department of Biological Science, Florida State University, Tallahassee, Florida 32306, USA; 2Division of Hematology and Oncology, Department of Pediatrics, and OHSU Knight Cancer Institute, Oregon Health & Science University, Portland, Oregon 97239, USA; 3COG and Department of Biostatistics, College of Medicine, University of Florida, Gainesville, Florida 32601, USA; 4Division of Hematology and Medical Oncology, and OHSU Knight Cancer Institute, Oregon Health & Science University, Portland, Oregon 97239, USA; 5Howard Hughes Medical Institute, Chevy Chase, Maryland 20815, USA Abnormal replication timing has been observed in cancer but no study has comprehensively evaluated this misregulation. We generated genome-wide replication-timing profiles for pediatric leukemias from 17 patients and three cell lines, as well as normal B and T cells. Nonleukemic EBV-transformed lymphoblastoid cell lines displayed highly stable replication- timing profiles that were more similar to normal T cells than to leukemias. Leukemias were more similar to each other than to B and T cells but were considerably more heterogeneous than nonleukemic controls. Some differences were patient specific, while others were found in all leukemic samples, potentially representing early epigenetic events. Differences encompassed large segments of chromosomes and included genes implicated in other types of cancer. Remarkably, dif- ferences that distinguished leukemias aligned in register to the boundaries of developmentally regulated replication- timing domains that distinguish normal cell types.
    [Show full text]
  • N6-Methyladenosine (M6a) Is an Endogenous A3 Adenosine Receptor Ligand
    bioRxiv preprint doi: https://doi.org/10.1101/2020.11.21.391136; this version posted November 22, 2020. 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-NC-ND 4.0 International license. N6-methyladenosine (m6A) is an endogenous A3 adenosine receptor ligand Akiko Ogawa,1, 2, 3 Chisae Nagiri,4, 13 Wataru Shihoya,4, 13 Asuka Inoue,5, 6, 7, 13 Kouki Kawakami,5 Suzune Hiratsuka,5 Junken Aoki,7, 8 Yasuhiro Ito,3 Takeo Suzuki,9 Tsutomu Suzuki,9 Toshihiro Inoue,3 Osamu Nureki,4 Hidenobu Tanihara,10 Kazuhito Tomizawa,2, 11 Fan-Yan Wei1, 2, 12, 14 1Department of Modomics Biology & Medicine, Institute of Development, Aging and Cancer (IDAC), Tohoku University, Sendai 980-8575, Japan. 2Department of Molecular Physiology, Faculty of Life Sciences, Kumamoto University, Kumamoto 860-8556, Japan. 3Department of Ophthalmology, Faculty of Life Sciences, Kumamoto University, Kumamoto 860-8556, Japan. 4Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Tokyo 113-0033, Japan. 1 bioRxiv preprint doi: https://doi.org/10.1101/2020.11.21.391136; this version posted November 22, 2020. 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-NC-ND 4.0 International license. 5Laboratory of Molecular and Cellular Biochemistry, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai 980-8578, Japan.
    [Show full text]
  • WO 2012/174282 A2 20 December 2012 (20.12.2012) P O P C T
    (12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization International Bureau (10) International Publication Number (43) International Publication Date WO 2012/174282 A2 20 December 2012 (20.12.2012) P O P C T (51) International Patent Classification: David [US/US]; 13539 N . 95th Way, Scottsdale, AZ C12Q 1/68 (2006.01) 85260 (US). (21) International Application Number: (74) Agent: AKHAVAN, Ramin; Caris Science, Inc., 6655 N . PCT/US20 12/0425 19 Macarthur Blvd., Irving, TX 75039 (US). (22) International Filing Date: (81) Designated States (unless otherwise indicated, for every 14 June 2012 (14.06.2012) kind of national protection available): AE, AG, AL, AM, AO, AT, AU, AZ, BA, BB, BG, BH, BR, BW, BY, BZ, English (25) Filing Language: CA, CH, CL, CN, CO, CR, CU, CZ, DE, DK, DM, DO, Publication Language: English DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, HN, HR, HU, ID, IL, IN, IS, JP, KE, KG, KM, KN, KP, KR, (30) Priority Data: KZ, LA, LC, LK, LR, LS, LT, LU, LY, MA, MD, ME, 61/497,895 16 June 201 1 (16.06.201 1) US MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ, 61/499,138 20 June 201 1 (20.06.201 1) US OM, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SC, SD, 61/501,680 27 June 201 1 (27.06.201 1) u s SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM, TN, TR, 61/506,019 8 July 201 1(08.07.201 1) u s TT, TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, ZW.
    [Show full text]
  • Download Tool
    by Submitted in partial satisfaction of the requirements for degree of in in the GRADUATE DIVISION of the UNIVERSITY OF CALIFORNIA, SAN FRANCISCO Approved: ______________________________________________________________________________ Chair ______________________________________________________________________________ ______________________________________________________________________________ ______________________________________________________________________________ ______________________________________________________________________________ Committee Members Copyright 2019 by Adolfo Cuesta ii Acknowledgements For me, completing a doctoral dissertation was a huge undertaking that was only possible with the support of many people along the way. First, I would like to thank my PhD advisor, Jack Taunton. He always gave me the space to pursue my own ideas and interests, while providing thoughtful guidance. Nearly every aspect of this project required a technique that was completely new to me. He trusted that I was up to the challenge, supported me throughout, helped me find outside resources when necessary. I remain impressed with his voracious appetite for the literature, and ability to recall some of the most subtle, yet most important details in a paper. Most of all, I am thankful that Jack has always been so generous with his time, both in person, and remotely. I’ve enjoyed our many conversations and hope that they will continue. I’d also like to thank my thesis committee, Kevan Shokat and David Agard for their valuable support, insight, and encouragement throughout this project. My lab mates in the Taunton lab made this such a pleasant experience, even on the days when things weren’t working well. I worked very closely with Tangpo Yang on the mass spectrometry aspects of this project. Xiaobo Wan taught me almost everything I know about protein crystallography. Thank you as well to Geoff Smith, Jordan Carelli, Pat Sharp, Yazmin Carassco, Keely Oltion, Nicole Wenzell, Haoyuan Wang, Steve Sethofer, and Shyam Krishnan, Shawn Ouyang and Qian Zhao.
    [Show full text]
  • Family-Based Investigation of the Genetics and Epigenetics of Obesity in Qatar
    Family-Based Investigation of the Genetics and Epigenetics of Obesity in Qatar Mashael Nedham A J Alshafai A Thesis Submitted for the Degree of Doctor of Philosophy Department of Genomics of Common Disease School of Public Health Imperial College London July 2015 1 Abstract Abstract Single nucleotide polymorphisms (SNPs), copy number variations (CNVs) and DNA methylation patterns play a role in the susceptibility to obesity. Despite the alarming figures of obesity in the Arab world, the genetics of obesity remain understudied in Arabs. Here, I recruited ten multigenerational Qatari families segregating obesity, and generated genome-wide SNP genotyping, whole genome sequencing and genome-wide methylation profiling data using Illumina platforms to investigate the role of rare mutations, CNVs, and DNA methylation patterns in the susceptibility to obesity. For the identification of obesity mutations, I first identified candidate obesity regions through linkage and run of homozygosity analyses, and then investigated these regions to detect potential deleterious mutations. These analyses highlighted putative rare variants for obesity risk at PCSK1, NMUR2, CLOCK and RETSAT. The functional impact of the PCSK1 mutation on obesity was previously demonstrated while for the other candidates further work is needed to confirm their impact. For the identification of common obesity CNVs, I performed a genome-wide CNV association analysis with BMI and identified a common duplication of ~5.6kb on 19p13.3 that associates with higher BMI. Moreover, I investigated large (≥500kb) rare CNVs and identified a ~618kb deletion on 16p11.2 in the most extremely obese subject in my samples, which confirms the contribution of the previously reported 16p11.2 deletions to severe obesity beyond European populations.
    [Show full text]
  • Identification of Genetic Factors Underpinning Phenotypic Heterogeneity in Huntington’S Disease and Other Neurodegenerative Disorders
    Identification of genetic factors underpinning phenotypic heterogeneity in Huntington’s disease and other neurodegenerative disorders. By Dr Davina J Hensman Moss A thesis submitted to University College London for the degree of Doctor of Philosophy Department of Neurodegenerative Disease Institute of Neurology University College London (UCL) 2020 1 I, Davina Hensman Moss confirm that the work presented in this thesis is my own. Where information has been derived from other sources, I confirm that this has been indicated in the thesis. Collaborative work is also indicated in this thesis. Signature: Date: 2 Abstract Neurodegenerative diseases including Huntington’s disease (HD), the spinocerebellar ataxias and C9orf72 associated Amyotrophic Lateral Sclerosis / Frontotemporal dementia (ALS/FTD) do not present and progress in the same way in all patients. Instead there is phenotypic variability in age at onset, progression and symptoms. Understanding this variability is not only clinically valuable, but identification of the genetic factors underpinning this variability has the potential to highlight genes and pathways which may be amenable to therapeutic manipulation, hence help find drugs for these devastating and currently incurable diseases. Identification of genetic modifiers of neurodegenerative diseases is the overarching aim of this thesis. To identify genetic variants which modify disease progression it is first necessary to have a detailed characterization of the disease and its trajectory over time. In this thesis clinical data from the TRACK-HD studies, for which I collected data as a clinical fellow, was used to study disease progression over time in HD, and give subjects a progression score for subsequent analysis. In this thesis I show blood transcriptomic signatures of HD status and stage which parallel HD brain and overlap with Alzheimer’s disease brain.
    [Show full text]
  • A Genome Wide Screen in C. Elegans Identifies Cell Non-Autonomous Regulators of Oncogenic Ras Mediated Over-Proliferation DISSER
    A genome wide screen in C. elegans identifies cell non-autonomous regulators of oncogenic Ras mediated over-proliferation DISSERTATION Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Komal Rambani Graduate Program in Biomedical Sciences The Ohio State University 2016 Dissertation Committee: Gustavo Leone, PhD “Advisor" Helen Chamberlin, PhD Gregory Lesinski, PhD Joanna Groden, PhD Jeffrey Parvin, PhD Thomas Ludwig, PhD Copyright by Komal Rambani 2016 ABSTRACT Coordinated proliferative signals from the mesenchymal cells play a crucial role in the regulation of proliferation of epithelial cells during normal development, wound healing and several other normal physiological conditions. However, when epithelial cells acquire a set of malignant mutations, they respond differently to these extrinsic proliferative signals elicited by the surrounding mesenchymal cells. This scenario leads to a pathological signaling microenvironment that enhances abnormal proliferation of mutant epithelial cells and hence tumor growth. Despite mounting evidence that mesenchymal (stromal) cells influence the growth of tumors and cancer progression, it is unclear which specific genes in the mesenchymal cells regulate the molecular signals that promote the over-proliferation of the adjacent mutant epithelial cells. We hypothesized that there are certain genes in the mesenchymal (stromal) cells that regulate proliferation of the adjacent mutant cells. The complexity of various stromal cell types and their interactions in vivo in cancer mouse models and human tumor samples limits our ability to identify mesenchymal genes important in this process. Thus, we took a cross-species approach to use C. elegans vulval development as a model to understand the impact of mesenchymal (mesodermal) cells on the proliferation of epithelial (epidermal) cells.
    [Show full text]
  • Genomic Mapping of the MHC Transactivator CIITA Using An
    Wong et al. Genome Biology 2014, 15:494 http://genomebiology.com/2014/15/10/494 RESEARCH Open Access Genomic mapping of the MHC transactivator CIITA using an integrated ChIP-seq and genetical genomics approach Daniel Wong1†, Wanseon Lee1†, Peter Humburg1, Seiko Makino1, Evelyn Lau1, Vivek Naranbhai1, Benjamin P Fairfax1, Kenneth Chan2, Katharine Plant1 and Julian C Knight1* Abstract Background: The master transactivator CIITA is essential to the regulation of Major Histocompatibility Complex (MHC) class II genes and an effective immune response. CIITA is known to modulate a small number of non-MHC genes involved in antigen presentation such as CD74 and B2M but its broader genome-wide function and relationship with underlying genetic diversity has not been resolved. Results: We report the first genome-wide ChIP-seq map for CIITA and complement this by mapping inter-individual variation in CIITA expression as a quantitative trait. We analyse CIITA recruitment for pathophysiologically relevant primary human B cells and monocytes, resting and treated with interferon-gamma, in the context of the epigenomic regulatory landscape and DNA-binding proteins associated with the CIITA enhanceosome including RFX, CREB1/ATF1 and NFY. We confirm recruitment to proximal promoter sequences in MHC class II genes and more distally involving the canonical CIITA enhanceosome. Overall, we map 843 CIITA binding intervals involving 442 genes and find 95% of intervals are located outside the MHC and 60% not associated with RFX5 binding. Binding intervals are enriched for genes involved in immune function and infectious disease with novel loci including major histone gene clusters. We resolve differentially expressed genes associated in trans with a CIITA intronic sequence variant, integrate with CIITA recruitment and show how this is mediated by allele-specific recruitment of NF-kB.
    [Show full text]
  • A High-Throughput Approach to Uncover Novel Roles of APOBEC2, a Functional Orphan of the AID/APOBEC Family
    Rockefeller University Digital Commons @ RU Student Theses and Dissertations 2018 A High-Throughput Approach to Uncover Novel Roles of APOBEC2, a Functional Orphan of the AID/APOBEC Family Linda Molla Follow this and additional works at: https://digitalcommons.rockefeller.edu/ student_theses_and_dissertations Part of the Life Sciences Commons A HIGH-THROUGHPUT APPROACH TO UNCOVER NOVEL ROLES OF APOBEC2, A FUNCTIONAL ORPHAN OF THE AID/APOBEC FAMILY A Thesis Presented to the Faculty of The Rockefeller University in Partial Fulfillment of the Requirements for the degree of Doctor of Philosophy by Linda Molla June 2018 © Copyright by Linda Molla 2018 A HIGH-THROUGHPUT APPROACH TO UNCOVER NOVEL ROLES OF APOBEC2, A FUNCTIONAL ORPHAN OF THE AID/APOBEC FAMILY Linda Molla, Ph.D. The Rockefeller University 2018 APOBEC2 is a member of the AID/APOBEC cytidine deaminase family of proteins. Unlike most of AID/APOBEC, however, APOBEC2’s function remains elusive. Previous research has implicated APOBEC2 in diverse organisms and cellular processes such as muscle biology (in Mus musculus), regeneration (in Danio rerio), and development (in Xenopus laevis). APOBEC2 has also been implicated in cancer. However the enzymatic activity, substrate or physiological target(s) of APOBEC2 are unknown. For this thesis, I have combined Next Generation Sequencing (NGS) techniques with state-of-the-art molecular biology to determine the physiological targets of APOBEC2. Using a cell culture muscle differentiation system, and RNA sequencing (RNA-Seq) by polyA capture, I demonstrated that unlike the AID/APOBEC family member APOBEC1, APOBEC2 is not an RNA editor. Using the same system combined with enhanced Reduced Representation Bisulfite Sequencing (eRRBS) analyses I showed that, unlike the AID/APOBEC family member AID, APOBEC2 does not act as a 5-methyl-C deaminase.
    [Show full text]
  • The Human Canonical Core Histone Catalogue David Miguel Susano Pinto*, Andrew Flaus*,†
    bioRxiv preprint doi: https://doi.org/10.1101/720235; this version posted July 30, 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 4.0 International license. The Human Canonical Core Histone Catalogue David Miguel Susano Pinto*, Andrew Flaus*,† Abstract Core histone proteins H2A, H2B, H3, and H4 are encoded by a large family of genes dis- tributed across the human genome. Canonical core histones contribute the majority of proteins to bulk chromatin packaging, and are encoded in 4 clusters by 65 coding genes comprising 17 for H2A, 18 for H2B, 15 for H3, and 15 for H4, along with at least 17 total pseudogenes. The canonical core histone genes display coding variation that gives rise to 11 H2A, 15 H2B, 4 H3, and 2 H4 unique protein isoforms. Although histone proteins are highly conserved overall, these isoforms represent a surprising and seldom recognised variation with amino acid identity as low as 77 % between canonical histone proteins of the same type. The gene sequence and protein isoform diversity also exceeds com- monly used subtype designations such as H2A.1 and H3.1, and exists in parallel with the well-known specialisation of variant histone proteins. RNA sequencing of histone transcripts shows evidence for differential expression of histone genes but the functional significance of this variation has not yet been investigated. To assist understanding of the implications of histone gene and protein diversity we have catalogued the entire human canonical core histone gene and protein complement.
    [Show full text]
  • A Multiprotein Occupancy Map of the Mrnp on the 3 End of Histone
    Downloaded from rnajournal.cshlp.org on October 6, 2021 - Published by Cold Spring Harbor Laboratory Press A multiprotein occupancy map of the mRNP on the 3′ end of histone mRNAs LIONEL BROOKS III,1 SHAWN M. LYONS,2 J. MATTHEW MAHONEY,1 JOSHUA D. WELCH,3 ZHONGLE LIU,1 WILLIAM F. MARZLUFF,2 and MICHAEL L. WHITFIELD1 1Department of Genetics, Dartmouth Geisel School of Medicine, Hanover, New Hampshire 03755, USA 2Integrative Program for Biological and Genome Sciences, University of North Carolina, Chapel Hill, North Carolina 27599, USA 3Department of Computer Science, University of North Carolina, Chapel Hill, North Carolina 27599, USA ABSTRACT The animal replication-dependent (RD) histone mRNAs are coordinately regulated with chromosome replication. The RD-histone mRNAs are the only known cellular mRNAs that are not polyadenylated. Instead, the mature transcripts end in a conserved stem– loop (SL) structure. This SL structure interacts with the stem–loop binding protein (SLBP), which is involved in all aspects of RD- histone mRNA metabolism. We used several genomic methods, including high-throughput sequencing of cross-linked immunoprecipitate (HITS-CLIP) to analyze the RNA-binding landscape of SLBP. SLBP was not bound to any RNAs other than histone mRNAs. We performed bioinformatic analyses of the HITS-CLIP data that included (i) clustering genes by sequencing read coverage using CVCA, (ii) mapping the bound RNA fragment termini, and (iii) mapping cross-linking induced mutation sites (CIMS) using CLIP-PyL software. These analyses allowed us to identify specific sites of molecular contact between SLBP and its RD-histone mRNA ligands. We performed in vitro crosslinking assays to refine the CIMS mapping and found that uracils one and three in the loop of the histone mRNA SL preferentially crosslink to SLBP, whereas uracil two in the loop preferentially crosslinks to a separate component, likely the 3′hExo.
    [Show full text]