DOCSLIB.ORG

Mechanisms of Centriole Duplication and Spindle Assembly in Human Cells

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text

Load more

Mechanisms of centriole duplication and spindle assembly in human cells by Qianzhu Wu A thesis submitted in conformity with the requirements for the degree of Doctorate of Philosophy Molecular Genetics University of Toronto © Copyright by Qianzhu Wu (2019) Mechanisms of centriole duplication and spindle assembly in human cells Qianzhu Wu Doctorate of Philosophy Molecular Genetics University of Toronto 2019 Abstract The centrosome is the major microtubule-organizing center in animal cells. Centrosomes duplicate once and only once per cell cycle thereby ensuring that two centrosomes are present in mitosis, each of them organizing one of the two spindle poles. Numerical and structural centrosomal abnormities lead to a number of devastating human diseases, including ciliopathies, microcephaly and cancer. Despite extensive effort at elucidating the molecular mechanisms underpinning centriole assembly, our understanding of this process remains incomplete and many novel factors remain to be identified. Here, using a high-throughput, semi-automated centriole duplication screen, I identified TRIM36 as a novel regulator of centriole duplication and mitotic spindle assembly. TRIM36 is a microtubule-binding RING-type E3 ligase. In centriole duplication, TRIM36 regulates daughter centriole assembly in a manner that is dependent on the RING domain. Using super-resolution microscopy, I discovered that TRIM36 plays a role in the early steps of centriole formation, where it mediates the recruitment of SAS6. TRIM36 depletion ii also caused severe defects in mitotic spindle assembly and chromosome congression. My results indicated that the dual role of TRIM36 in centriole duplication and spindle formation involves, at least in part, the recruitment of γ-tubulin to centrosomes. Co-depletion of RBM14, a negative regulator of centriole duplication, compensated for the loss of γ-tubulin recruitment and rescued both the centriole duplication and spindle assembly defects. These results provide insights into the critical role of the E3 ligase TRIM36 in centriole duplication, mitotic spindle assembly and cell cycle progression. iii Acknowledgments I would like to begin by thanking my supervisor, Dr. Laurence Pelletier. It was truly a wonderful experience working in your lab, with your enthusiasm and guidance, and with a team of talented scientists. I would not go this far without your support and guidance. Thank you! A big thank you to my committee members, Dr. Brian Raught & Dr. John Brumell. You always ask great and thoughtful questions, providing inspiring and insightful. Your comments have been very much appreciated. Thank you to all of our collaborators - Brian, Étienne, Anne-Claude, Frank, Nero and Dan - This work would not have been possible without your contributions. A big shout out to all members of the Pelletier lab, past and present and my friends at the LTRI. You made every day a fun and memorable experience. You have been amazing colleagues, friends and bench-mate (yes talking about you, Suzy). Thank you Sally & Johnny for going through my paper manuscript, which took a huge portion in this thesis. Thank you Ladan for being there in difficult times. Thank you Dave, Bahareh and João for great suggestions. A special thank you to Yi 1, Yi 2 and Gagan for all their support and help in the past years. You always kept me motivated and set me back on track. I will miss the baking days and the lab lunches. Thank you! Thank you to all my friends, Yiwang & Huayun, Kai & Huijuan, who have been supportive and generous with your time during my Ph.D. The entire experience would be so different without you. To my family, THANK YOU! Mom, Dad & Jianjin – I don’t think I can ever express how grateful I am for your love and support. I am incredibly fortunate to have you in my life. You all make me feel deeply blessed. Table of Contents Abstract ...................................................................................................................................... ii Acknowledgments ..................................................................................................................... iv Table of Contents ....................................................................................................................... v List of Tables ............................................................................................................................ ix List of Figures ............................................................................................................................ x List of Abbreviations ............................................................................................................... xiv Chapter 1. Introduction ............................................................................................................... 1 Introduction ........................................................................................................................... 2 1.1 The Centrosomes ............................................................................................................ 2 1.1.1 Organization of the human centrosome ................................................................ 2 1.1.2 Centrosome biogenesis ........................................................................................ 8 1.1.3 Spindle assembly ................................................................................................17 1.1.4 Centrosome structure/numerical abnormalities and diseases ...............................22 1.2 The Ubiquitination System ............................................................................................26 1.2.1 The covalent conjugation of ubiquitin with other cellular proteins regulates diverse eukaryotic cell functions.........................................................................26 1.2.2 Ubiquitination and de-ubiquitination in centriole duplication and spindle assembly ............................................................................................................30 1.3 The TRIM E3 ligases .....................................................................................................34 1.3.1 Sub-classes of TRIM family ...............................................................................34 1.3.2 TRIM36 .............................................................................................................36 1.4 Rationale of the Thesis ...................................................................................................37 Chapter 2. Identification of novel centriole duplication regulators .............................................39 Identification of novel centriole duplication regulators ..........................................................40 2.1 Statement of Contributions ............................................................................................40 2.2 Summary .......................................................................................................................41 2.3 Introduction ...................................................................................................................41 2.3.1 BioID .................................................................................................................41 2.4 Results ...........................................................................................................................44 2.4.1 Generation of the centriole duplication interactome ............................................44 2.4.2 Functional characterization of centriole duplication interactome and identification of novel centriole duplication regulators .......................................54 2.5 Discussion .....................................................................................................................66 2.6 Material and Methods ....................................................................................................68 2.6.1 Cell lines and Tissue culture ...............................................................................68 2.6.2 Sample preparation for BioID .............................................................................69 2.6.3 BioID followed by mass spectrometry ................................................................69 2.6.4 SAINT analysis ..................................................................................................70 2.6.5 U-2 OS S-phase arrested centriole over-duplication assay ..................................70 2.6.6 Statistical methods .............................................................................................71 Chapter 3. TRIM36 is a novel regulator of centriole duplication and microtubule organization factor ....................................................................................................................................72 TRIM36 is a novel regulator of centriole duplication and a microtubule organization factor ....................................................................................................................................73 3.1 Statement of Contributions ............................................................................................73 3.2 Summary .......................................................................................................................73 3.3 Introduction ...................................................................................................................74 vi 3.3.1 TRIM36 participates
Recommended publications
  • Mutations in CDK5RAP2 Cause Seckel Syndrome Go¨ Khan Yigit1,2,3,A, Karen E

    Mutations in CDK5RAP2 Cause Seckel Syndrome Go¨ Khan Yigit1,2,3,A, Karen E

    ORIGINAL ARTICLE Mutations in CDK5RAP2 cause Seckel syndrome Go¨ khan Yigit1,2,3,a, Karen E. Brown4,a,Hu¨ lya Kayserili5, Esther Pohl1,2,3, Almuth Caliebe6, Diana Zahnleiter7, Elisabeth Rosser8, Nina Bo¨ gershausen1,2,3, Zehra Oya Uyguner5, Umut Altunoglu5, Gudrun Nu¨ rnberg2,3,9, Peter Nu¨ rnberg2,3,9, Anita Rauch10, Yun Li1,2,3, Christian Thomas Thiel7 & Bernd Wollnik1,2,3 1Institute of Human Genetics, University of Cologne, Cologne, Germany 2Center for Molecular Medicine Cologne (CMMC), University of Cologne, Cologne, Germany 3Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany 4Chromosome Biology Group, MRC Clinical Sciences Centre, Imperial College School of Medicine, Hammersmith Hospital, London, W12 0NN, UK 5Department of Medical Genetics, Istanbul Medical Faculty, Istanbul University, Istanbul, Turkey 6Institute of Human Genetics, Christian-Albrechts-University of Kiel, Kiel, Germany 7Institute of Human Genetics, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Germany 8Department of Clinical Genetics, Great Ormond Street Hospital for Children, London, WC1N 3EH, UK 9Cologne Center for Genomics, University of Cologne, Cologne, Germany 10Institute of Medical Genetics, University of Zurich, Schwerzenbach-Zurich, Switzerland Keywords Abstract CDK5RAP2, CEP215, microcephaly, primordial dwarfism, Seckel syndrome Seckel syndrome is a heterogeneous, autosomal recessive disorder marked by pre- natal proportionate short stature, severe microcephaly, intellectual disability, and Correspondence characteristic facial features. Here, we describe the novel homozygous splice-site Bernd Wollnik, Center for Molecular mutations c.383+1G>C and c.4005-9A>GinCDK5RAP2 in two consanguineous Medicine Cologne (CMMC) and Institute of families with Seckel syndrome. CDK5RAP2 (CEP215) encodes a centrosomal pro- Human Genetics, University of Cologne, tein which is known to be essential for centrosomal cohesion and proper spindle Kerpener Str.
  • Molecular and Physiological Basis for Hair Loss in Near Naked Hairless and Oak Ridge Rhino-Like Mouse Models: Tracking the Role of the Hairless Gene

    Molecular and Physiological Basis for Hair Loss in Near Naked Hairless and Oak Ridge Rhino-Like Mouse Models: Tracking the Role of the Hairless Gene

    University of Tennessee, Knoxville TRACE: Tennessee Research and Creative Exchange Doctoral Dissertations Graduate School 5-2006 Molecular and Physiological Basis for Hair Loss in Near Naked Hairless and Oak Ridge Rhino-like Mouse Models: Tracking the Role of the Hairless Gene Yutao Liu University of Tennessee - Knoxville Follow this and additional works at: https://trace.tennessee.edu/utk_graddiss Part of the Life Sciences Commons Recommended Citation Liu, Yutao, "Molecular and Physiological Basis for Hair Loss in Near Naked Hairless and Oak Ridge Rhino- like Mouse Models: Tracking the Role of the Hairless Gene. " PhD diss., University of Tennessee, 2006. https://trace.tennessee.edu/utk_graddiss/1824 This Dissertation is brought to you for free and open access by the Graduate School at TRACE: Tennessee Research and Creative Exchange. It has been accepted for inclusion in Doctoral Dissertations by an authorized administrator of TRACE: Tennessee Research and Creative Exchange. For more information, please contact [email protected]. To the Graduate Council: I am submitting herewith a dissertation written by Yutao Liu entitled "Molecular and Physiological Basis for Hair Loss in Near Naked Hairless and Oak Ridge Rhino-like Mouse Models: Tracking the Role of the Hairless Gene." I have examined the final electronic copy of this dissertation for form and content and recommend that it be accepted in partial fulfillment of the requirements for the degree of Doctor of Philosophy, with a major in Life Sciences. Brynn H. Voy, Major Professor We have read this dissertation and recommend its acceptance: Naima Moustaid-Moussa, Yisong Wang, Rogert Hettich Accepted for the Council: Carolyn R.
  • Synergistic Genetic Interactions Between Pkhd1 and Pkd1 Result in an ARPKD-Like Phenotype in Murine Models

    Synergistic Genetic Interactions Between Pkhd1 and Pkd1 Result in an ARPKD-Like Phenotype in Murine Models

    BASIC RESEARCH www.jasn.org Synergistic Genetic Interactions between Pkhd1 and Pkd1 Result in an ARPKD-Like Phenotype in Murine Models Rory J. Olson,1 Katharina Hopp ,2 Harrison Wells,3 Jessica M. Smith,3 Jessica Furtado,1,4 Megan M. Constans,3 Diana L. Escobar,3 Aron M. Geurts,5 Vicente E. Torres,3 and Peter C. Harris 1,3 Due to the number of contributing authors, the affiliations are listed at the end of this article. ABSTRACT Background Autosomal recessive polycystic kidney disease (ARPKD) and autosomal dominant polycystic kidney disease (ADPKD) are genetically distinct, with ADPKD usually caused by the genes PKD1 or PKD2 (encoding polycystin-1 and polycystin-2, respectively) and ARPKD caused by PKHD1 (encoding fibrocys- tin/polyductin [FPC]). Primary cilia have been considered central to PKD pathogenesis due to protein localization and common cystic phenotypes in syndromic ciliopathies, but their relevance is questioned in the simple PKDs. ARPKD’s mild phenotype in murine models versus in humans has hampered investi- gating its pathogenesis. Methods To study the interaction between Pkhd1 and Pkd1, including dosage effects on the phenotype, we generated digenic mouse and rat models and characterized and compared digenic, monogenic, and wild-type phenotypes. Results The genetic interaction was synergistic in both species, with digenic animals exhibiting pheno- types of rapidly progressive PKD and early lethality resembling classic ARPKD. Genetic interaction be- tween Pkhd1 and Pkd1 depended on dosage in the digenic murine models, with no significant enhancement of the monogenic phenotype until a threshold of reduced expression at the second locus was breached.
  • A Computational Approach for Defining a Signature of Β-Cell Golgi Stress in Diabetes Mellitus

    A Computational Approach for Defining a Signature of Β-Cell Golgi Stress in Diabetes Mellitus

    Page 1 of 781 Diabetes A Computational Approach for Defining a Signature of β-Cell Golgi Stress in Diabetes Mellitus Robert N. Bone1,6,7, Olufunmilola Oyebamiji2, Sayali Talware2, Sharmila Selvaraj2, Preethi Krishnan3,6, Farooq Syed1,6,7, Huanmei Wu2, Carmella Evans-Molina 1,3,4,5,6,7,8* Departments of 1Pediatrics, 3Medicine, 4Anatomy, Cell Biology & Physiology, 5Biochemistry & Molecular Biology, the 6Center for Diabetes & Metabolic Diseases, and the 7Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202; 2Department of BioHealth Informatics, Indiana University-Purdue University Indianapolis, Indianapolis, IN, 46202; 8Roudebush VA Medical Center, Indianapolis, IN 46202. *Corresponding Author(s): Carmella Evans-Molina, MD, PhD ([email protected]) Indiana University School of Medicine, 635 Barnhill Drive, MS 2031A, Indianapolis, IN 46202, Telephone: (317) 274-4145, Fax (317) 274-4107 Running Title: Golgi Stress Response in Diabetes Word Count: 4358 Number of Figures: 6 Keywords: Golgi apparatus stress, Islets, β cell, Type 1 diabetes, Type 2 diabetes 1 Diabetes Publish Ahead of Print, published online August 20, 2020 Diabetes Page 2 of 781 ABSTRACT The Golgi apparatus (GA) is an important site of insulin processing and granule maturation, but whether GA organelle dysfunction and GA stress are present in the diabetic β-cell has not been tested. We utilized an informatics-based approach to develop a transcriptional signature of β-cell GA stress using existing RNA sequencing and microarray datasets generated using human islets from donors with diabetes and islets where type 1(T1D) and type 2 diabetes (T2D) had been modeled ex vivo. To narrow our results to GA-specific genes, we applied a filter set of 1,030 genes accepted as GA associated.
  • Par6c Is at the Mother Centriole and Controls Centrosomal Protein

    Par6c Is at the Mother Centriole and Controls Centrosomal Protein

    860 Research Article Par6c is at the mother centriole and controls centrosomal protein composition through a Par6a-dependent pathway Vale´rian Dormoy, Kati Tormanen and Christine Su¨ tterlin* Department of Developmental and Cell Biology, University of California, Irvine, Irvine, CA 92697-2300, USA *Author for correspondence ([email protected]) Accepted 3 December 2012 Journal of Cell Science 126, 860–870 ß 2013. Published by The Company of Biologists Ltd doi: 10.1242/jcs.121186 Summary The centrosome contains two centrioles that differ in age, protein composition and function. This non-membrane bound organelle is known to regulate microtubule organization in dividing cells and ciliogenesis in quiescent cells. These specific roles depend on protein appendages at the older, or mother, centriole. In this study, we identified the polarity protein partitioning defective 6 homolog gamma (Par6c) as a novel component of the mother centriole. This specific localization required the Par6c C-terminus, but was independent of intact microtubules, the dynein/dynactin complex and the components of the PAR polarity complex. Par6c depletion resulted in altered centrosomal protein composition, with the loss of a large number of proteins, including Par6a and p150Glued, from the centrosome. As a consequence, there were defects in ciliogenesis, microtubule organization and centrosome reorientation during migration. Par6c interacted with Par3 and aPKC, but these proteins were not required for the regulation of centrosomal protein composition. Par6c also associated with Par6a, which controls protein recruitment to the centrosome through p150Glued. Our study is the first to identify Par6c as a component of the mother centriole and to report a role of a mother centriole protein in the regulation of centrosomal protein composition.
  • Supplemental Information Proximity Interactions Among Centrosome

    Supplemental Information Proximity Interactions Among Centrosome

    Current Biology, Volume 24 Supplemental Information Proximity Interactions among Centrosome Components Identify Regulators of Centriole Duplication Elif Nur Firat-Karalar, Navin Rauniyar, John R. Yates III, and Tim Stearns Figure S1 A Myc Streptavidin -tubulin Merge Myc Streptavidin -tubulin Merge BirA*-PLK4 BirA*-CEP63 BirA*- CEP192 BirA*- CEP152 - BirA*-CCDC67 BirA* CEP152 CPAP BirA*- B C Streptavidin PCM1 Merge Myc-BirA* -CEP63 PCM1 -tubulin Merge BirA*- CEP63 DMSO - BirA* CEP63 nocodazole BirA*- CCDC67 Figure S2 A GFP – + – + GFP-CEP152 + – + – Myc-CDK5RAP2 + + + + (225 kDa) Myc-CDK5RAP2 (216 kDa) GFP-CEP152 (27 kDa) GFP Input (5%) IP: GFP B GFP-CEP152 truncation proteins Inputs (5%) IP: GFP kDa 1-7481-10441-1290218-1654749-16541045-16541-7481-10441-1290218-1654749-16541045-1654 250- Myc-CDK5RAP2 150- 150- 100- 75- GFP-CEP152 Figure S3 A B CEP63 – – + – – + GFP CCDC14 KIAA0753 Centrosome + – – + – – GFP-CCDC14 CEP152 binding binding binding targeting – + – – + – GFP-KIAA0753 GFP-KIAA0753 (140 kDa) 1-496 N M C 150- 100- GFP-CCDC14 (115 kDa) 1-424 N M – 136-496 M C – 50- CEP63 (63 kDa) 1-135 N – 37- GFP (27 kDa) 136-424 M – kDa 425-496 C – – Inputs (2%) IP: GFP C GFP-CEP63 truncation proteins D GFP-CEP63 truncation proteins Inputs (5%) IP: GFP Inputs (5%) IP: GFP kDa kDa 1-135136-424425-4961-424136-496FL Ctl 1-135136-424425-4961-424136-496FL Ctl 1-135136-424425-4961-424136-496FL Ctl 1-135136-424425-4961-424136-496FL Ctl Myc- 150- Myc- 100- CCDC14 KIAA0753 100- 100- 75- 75- GFP- GFP- 50- CEP63 50- CEP63 37- 37- Figure S4 A siCtl
  • The P53/P73 - P21cip1 Tumor Suppressor Axis Guards Against Chromosomal Instability by Restraining CDK1 in Human Cancer Cells

    The P53/P73 - P21cip1 Tumor Suppressor Axis Guards Against Chromosomal Instability by Restraining CDK1 in Human Cancer Cells

    Oncogene (2021) 40:436–451 https://doi.org/10.1038/s41388-020-01524-4 ARTICLE The p53/p73 - p21CIP1 tumor suppressor axis guards against chromosomal instability by restraining CDK1 in human cancer cells 1 1 2 1 2 Ann-Kathrin Schmidt ● Karoline Pudelko ● Jan-Eric Boekenkamp ● Katharina Berger ● Maik Kschischo ● Holger Bastians 1 Received: 2 July 2020 / Revised: 2 October 2020 / Accepted: 13 October 2020 / Published online: 9 November 2020 © The Author(s) 2020. This article is published with open access Abstract Whole chromosome instability (W-CIN) is a hallmark of human cancer and contributes to the evolvement of aneuploidy. W-CIN can be induced by abnormally increased microtubule plus end assembly rates during mitosis leading to the generation of lagging chromosomes during anaphase as a major form of mitotic errors in human cancer cells. Here, we show that loss of the tumor suppressor genes TP53 and TP73 can trigger increased mitotic microtubule assembly rates, lagging chromosomes, and W-CIN. CDKN1A, encoding for the CDK inhibitor p21CIP1, represents a critical target gene of p53/p73. Loss of p21CIP1 unleashes CDK1 activity which causes W-CIN in otherwise chromosomally stable cancer cells. fi Vice versa 1234567890();,: 1234567890();,: Consequently, induction of CDK1 is suf cient to induce abnormal microtubule assembly rates and W-CIN. , partial inhibition of CDK1 activity in chromosomally unstable cancer cells corrects abnormal microtubule behavior and suppresses W-CIN. Thus, our study shows that the p53/p73 - p21CIP1 tumor suppressor axis, whose loss is associated with W-CIN in human cancer, safeguards against chromosome missegregation and aneuploidy by preventing abnormally increased CDK1 activity.
  • Cep57, a NEDD1-Binding Pericentriolar Material Component, Is Essential for Spindle Pole Integrity

    Cep57, a NEDD1-Binding Pericentriolar Material Component, Is Essential for Spindle Pole Integrity

    Cell Research (2012) :1-12. © 2012 IBCB, SIBS, CAS All rights reserved 1001-0602/12 $ 32.00 npg ORIGINAL ARTICLE www.nature.com/cr Cep57, a NEDD1-binding pericentriolar material component, is essential for spindle pole integrity Qixi Wu1, *, Runsheng He1, *, Haining Zhou1, Albert CH Yu2, Bo Zhang1, Junlin Teng1, Jianguo Chen1, 3 1The State Key Laboratory of Biomembrane and Membrane Bioengineering and The Key Laboratory of Cell Proliferation and Dif- ferentiation of Ministry of Education, College of Life Sciences, Peking University, Beijing 100871, China; 2Department of Neurobi- ology, Neuroscience Research Institute, School of Basic Medical Sciences, Peking University, Beijing 100191, China; 3The Center for Theoretical Biology, Peking University, Beijing 100871, China Formation of a bipolar spindle is indispensable for faithful chromosome segregation and cell division. Spindle in- tegrity is largely dependent on the centrosome and the microtubule network. Centrosome protein Cep57 can bundle microtubules in mammalian cells. Its related protein (Cep57R) in Xenopus was characterized as a stabilization factor for microtubule-kinetochore attachment. Here we show that Cep57 is a pericentriolar material (PCM) component. Its interaction with NEDD1 is necessary for the centrosome localization of Cep57. Depletion of Cep57 leads to unaligned chromosomes and a multipolar spindle, which is induced by PCM fragmentation. In the absence of Cep57, cen- trosome microtubule array assembly activity is weakened, and the spindle length and microtubule density decrease. As a spindle microtubule-binding protein, Cep57 is also responsible for the proper organization of the spindle micro- tubule and localization of spindle pole focusing proteins. Collectively, these results suggest that Cep57, as a NEDD1- binding centrosome component, could function as a spindle pole- and microtubule-stabilizing factor for establishing robust spindle architecture.
  • A Genome-Wide Association Study Identifies New Susceptibility Loci for Esophageal Adenocarcinoma and Barrett's Esophagus

    A Genome-Wide Association Study Identifies New Susceptibility Loci for Esophageal Adenocarcinoma and Barrett's Esophagus

    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by White Rose Research Online This is an author produced version of A genome-wide association study identifies new susceptibility loci for esophageal adenocarcinoma and Barrett's esophagus.. White Rose Research Online URL for this paper: http://eprints.whiterose.ac.uk/105073/ Article: Levine, D.M., Ek, W.E., Zhang, R. et al. (31 more authors) (2013) A genome-wide association study identifies new susceptibility loci for esophageal adenocarcinoma and Barrett's esophagus. Nature Genetics, 45 (12). pp. 1487-1493. ISSN 1061-4036 https://doi.org/10.1038/ng.2796 promoting access to White Rose research papers [email protected] http://eprints.whiterose.ac.uk/ HHS Public Access Author manuscript Author Manuscript Author ManuscriptNat Genet Author Manuscript. Author manuscript; Author Manuscript available in PMC 2014 June 01. Published in final edited form as: Nat Genet. 2013 December ; 45(12): 1487–1493. doi:10.1038/ng.2796. A Genome-Wide Association Study Identifies New Susceptibility Loci for Esophageal Adenocarcinoma and Barrett’ s Esophagus David M. Levine1, Weronica E. Ek2, Rui Zhang1, Xinxue Liu3, Lynn Onstad4, Cassandra Sather5, Pierre Lao-Sirieix3, Marilie D. Gammon6, Douglas A. Corley7, Nicholas J. Shaheen8, Nigel C. Bird9, Laura J. Hardie10, Liam J. Murray11, Brian J. Reid4,12, Wong-Ho Chow13, Harvey A. Risch14, Olof Nyrén15, Weimin Ye15, Geoffrey Liu16, Yvonne Romero17,18, Leslie Bernstein19, Anna H. Wu20, Alan G. Casson21, Stephen Chanock22, Patricia Harrington23,24,25, Isabel Caldas25, Irene Debiram-Beecham3, Carlos Caldas25,26, Nicholas K. Hayward27, Paul Pharoah23,24,25, Rebecca Fitzgerald3, Stuart MacGregor2, David C.
  • Genetic and Genomic Analysis of Hyperlipidemia, Obesity and Diabetes Using (C57BL/6J × TALLYHO/Jngj) F2 Mice

    Genetic and Genomic Analysis of Hyperlipidemia, Obesity and Diabetes Using (C57BL/6J × TALLYHO/Jngj) F2 Mice

    University of Tennessee, Knoxville TRACE: Tennessee Research and Creative Exchange Nutrition Publications and Other Works Nutrition 12-19-2010 Genetic and genomic analysis of hyperlipidemia, obesity and diabetes using (C57BL/6J × TALLYHO/JngJ) F2 mice Taryn P. Stewart Marshall University Hyoung Y. Kim University of Tennessee - Knoxville, [email protected] Arnold M. Saxton University of Tennessee - Knoxville, [email protected] Jung H. Kim Marshall University Follow this and additional works at: https://trace.tennessee.edu/utk_nutrpubs Part of the Animal Sciences Commons, and the Nutrition Commons Recommended Citation BMC Genomics 2010, 11:713 doi:10.1186/1471-2164-11-713 This Article is brought to you for free and open access by the Nutrition at TRACE: Tennessee Research and Creative Exchange. It has been accepted for inclusion in Nutrition Publications and Other Works by an authorized administrator of TRACE: Tennessee Research and Creative Exchange. For more information, please contact [email protected]. Stewart et al. BMC Genomics 2010, 11:713 http://www.biomedcentral.com/1471-2164/11/713 RESEARCH ARTICLE Open Access Genetic and genomic analysis of hyperlipidemia, obesity and diabetes using (C57BL/6J × TALLYHO/JngJ) F2 mice Taryn P Stewart1, Hyoung Yon Kim2, Arnold M Saxton3, Jung Han Kim1* Abstract Background: Type 2 diabetes (T2D) is the most common form of diabetes in humans and is closely associated with dyslipidemia and obesity that magnifies the mortality and morbidity related to T2D. The genetic contribution to human T2D and related metabolic disorders is evident, and mostly follows polygenic inheritance. The TALLYHO/ JngJ (TH) mice are a polygenic model for T2D characterized by obesity, hyperinsulinemia, impaired glucose uptake and tolerance, hyperlipidemia, and hyperglycemia.
  • Supplemental Information

    Supplemental Information

    Supplemental information Dissection of the genomic structure of the miR-183/96/182 gene. Previously, we showed that the miR-183/96/182 cluster is an intergenic miRNA cluster, located in a ~60-kb interval between the genes encoding nuclear respiratory factor-1 (Nrf1) and ubiquitin-conjugating enzyme E2H (Ube2h) on mouse chr6qA3.3 (1). To start to uncover the genomic structure of the miR- 183/96/182 gene, we first studied genomic features around miR-183/96/182 in the UCSC genome browser (http://genome.UCSC.edu/), and identified two CpG islands 3.4-6.5 kb 5’ of pre-miR-183, the most 5’ miRNA of the cluster (Fig. 1A; Fig. S1 and Seq. S1). A cDNA clone, AK044220, located at 3.2-4.6 kb 5’ to pre-miR-183, encompasses the second CpG island (Fig. 1A; Fig. S1). We hypothesized that this cDNA clone was derived from 5’ exon(s) of the primary transcript of the miR-183/96/182 gene, as CpG islands are often associated with promoters (2). Supporting this hypothesis, multiple expressed sequences detected by gene-trap clones, including clone D016D06 (3, 4), were co-localized with the cDNA clone AK044220 (Fig. 1A; Fig. S1). Clone D016D06, deposited by the German GeneTrap Consortium (GGTC) (http://tikus.gsf.de) (3, 4), was derived from insertion of a retroviral construct, rFlpROSAβgeo in 129S2 ES cells (Fig. 1A and C). The rFlpROSAβgeo construct carries a promoterless reporter gene, the β−geo cassette - an in-frame fusion of the β-galactosidase and neomycin resistance (Neor) gene (5), with a splicing acceptor (SA) immediately upstream, and a polyA signal downstream of the β−geo cassette (Fig.
  • The Complete Genome Sequence of Mycobacterium Avium Subspecies Paratuberculosis

    The Complete Genome Sequence of Mycobacterium Avium Subspecies Paratuberculosis

    The complete genome sequence of Mycobacterium avium subspecies paratuberculosis Lingling Li*†‡, John P. Bannantine‡§, Qing Zhang*†‡, Alongkorn Amonsin*†‡, Barbara J. May*†, David Alt§, Nilanjana Banerji†¶, Sagarika Kanjilal†‡¶, and Vivek Kapur*†‡ʈ *Department of Microbiology, †Biomedical Genomics Center, and ¶Department of Medicine, University of Minnesota, St. Paul, MN 55108; and §National Animal Disease Center, U.S. Department of Agriculture–Agriculture Research Service, Ames, IA 50010 Communicated by Harley W. Moon, Iowa State University, Ames, IA, July 13, 2005 (received for review March 18, 2005) We describe here the complete genome sequence of a common Map and Mav (11–13). Therefore, it is widely recognized that the clone of Mycobacterium avium subspecies paratuberculosis (Map) development of rapid, sensitive, and specific assays to identify strain K-10, the causative agent of Johne’s disease in cattle and infected animals is essential to the formulation of rational other ruminants. The K-10 genome is a single circular chromosome strategies to control the spread of Map. of 4,829,781 base pairs and encodes 4,350 predicted ORFs, 45 As a first step toward elucidating the molecular basis of Map’s tRNAs, and one rRNA operon. In silico analysis identified >3,000 physiology and virulence, and providing a foundation for the genes with homologs to the human pathogen, M. tuberculosis development of the next generation of Map diagnostic tests and (Mtb), and 161 unique genomic regions that encode 39 previously vaccines, we report the complete genome sequence of a common unknown Map genes. Analysis of nucleotide substitution rates clone of Map, strain K-10. with Mtb homologs suggest overall strong selection for a vast majority of these shared mycobacterial genes, with only 68 ORFs Materials and Methods with a synonymous to nonsynonymous substitution ratio of >2.