DOCSLIB.ORG

01 Front.Pdf (5.994Mb)

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
01 Front.Pdf (5.994Mb) Copyright is owned by the Author of the thesis. Permission is given for a copy to be downloaded by an individual for the purpose of research and private study only. The thesis may not be reproduced elsewhere without the permission of the Author. Molecular Genetic Analysis for Malignant Hyperthermia A presented to Massey University in partial fulfilment of the requirements for the degree of Master of Science in Biochemistry Keisaku Sato 2004 ACKNOWLEDGEMENTS I especially would like to thank my supervisor, Dr. Kathryn Stowell for her help, advise and patience during my project. M s. Robyn MaL ton helped me for some sample preparations and I obtained many ideas and protocols for experimen tal designs from Dr. Rosemary Brown, although I have never seen her yet ... I would also like 10 thank all members al the Twilight Zone. I have never had any di fficulties or problems in the relationship with them. When I was having trouble and hard time. they were very kind and welcomed me warmly. I love the warm-hearted atmosphere in this laboratory. Finally. I have to than k my mother who always understands and accepts me. ABSTRACT M alignant hyperthermia (MH) is a rare pharmacogenetic disorder in humans caused by inhalational general anaesthetics and depolarising muscle relaxants. An MH reaction shows abnormal calcium homeostasis in skeletal muscle lead ing to a hypermetabolic state and increased muscle contracture. A mutation w ithin the calcium release channel ryanodine receptor of. keletal muscle (RYR I) is one of the causes of MH leading to the abnormally high release of calcium ions into the cytosol during MH reactions. The MH reaction can also be triggered by excess exercise. hea t and stress. A New Zealand male. identified as M 8 I 8, showed a fulminant MH reac tion which resu lted in death. The reaction was caused by exerci. e. and he did not have a family history of MH. As this individual did not have any of the mutations within RYR I found 10 date in New Zealand families. the entire RYRI cDNA was screened for a novel mutation that may result in susceptibility to exercise- induced MH. This patien t may have had a novel RYR I mutation because exercise- induced MH is quite rare. Screening of this gene. however did not identify any mutations within RYR I, suggesting that the M 8 18 patient may have a mutation in another gene because Ml-I is a heterogeneous disorder w ith 40-50% of fam ilies showing linkage to alternati ve loci. Heterogeneity of M H can re. ult in discordance between genotype and phenotype. So me Ml-I susceptible patients do not have a RYR I mutation that is found in other individuals w ith the same kindred. One or more other genes could be associated with M H for these individuals al though alternati ve loci have not been studied in New Zealand fami lies . A genome-wide scan was performed to search for other candidate loc i using a large M l-I kindred known as the CH family w ithin which discordance has been observed. Non-parametric linkage analysis across all chromosomes identified five weak linkages from one branch, and two strong linkages from another branch of the CH family. Secondary linkage analysis was performed on one candidate locus identified in the genome-wide scan, and a weak linkage and recombination was observed within the shorter region. No candidate genes with obvious relevance to calcium homeostasis or signalling were identified w ithin this region. The existence of alternative causative loci in this fami ly cannot be ruled out however, because the loci identified from the genome-w ide scan are very large and contain many genes of unknown function. ABBREVIATIONS A absorbance ATP adenosine triphosphate bp base pair CCD central core di sease cDNA co mpl ementary DNA cM centimorga n 4-CmC 4-chloro-m-cresol C-terminal carboxy terminal DEPC diethyl pyrocarbonate DHPR dihydropyridine receptor DNase deoxyribonuclease DMSO dimethyl su lphox ide d TPs dinucleotide triphosphate EC excitation-co ntraction EDTA ethylene diamine tetra-acetate HEPES -l2-hydroxyethyl]piperazine-N' -[ 4-butanesulphonic acid] IVCT in vitro contracture test kb kilobase MH mali gnant hyperthermia MHE malignant hyperthermia equivocal (European protocol) MHN malignant hyperthermia negative (European protocol) MHS malignant hyperthermia susceptible (European protocol) MH+ malignant hyperthermia positive (North American protocol) MH­ malignant hyperthermia negative (North American protocol) mRNA messenger RNA II MOPS 3-[N-morpholino]propanesulphonic acid NPL non-parametric linkage N-terminal amino terminal oligo(dT) oli godeoxythymidine pC02 carbon dioxide pressure PCR polymerase chain reaction PCr phos phocreati ne Pi in orga ni c phosphorus 3 1PNMR phos phorus-3 1 nucl ear magnetic resonance RFLP res tri ction fragment length pol ymorphi sm R ase ribonuclease RT reverse transcri ptase RT-PCR reverse transc riptase-polymerase chain reaction RYR rya nod ine receptor SIDS sudden infant death syndrome SR sarco pl as mi c reti culum SSCP sin gle stranded co nformational polymorphi sm TAE Tris-acetate-EDTA buffer Taq Therm,us aquaticus TBE Tri s-borate-EDTA buffer TE Tris-EDTA buffer Tm melting temperature Tris tris (hydroxymethyl ) ami nomethane T-tubule transverse tubule UV ultraviolet li ght VNTR variable number of tandem repeat pol ymorphism Ill LIST OF FIGURES Page Figure 1- 1 The excitation-contraction pathway of skeletal muscle 5 Figure 1-2 Proposed mechanism for induction of malignant hyperthermia 7 Figure 1-3 A linear representation of location of RYRI mutations 14 Figure 2- 1 Typical PCR amplification curves 29 Figure 2-2 Concepts of quantificati on using external standard samples 30 Fi gure 3- 1 Quality check of genomic D A 4 1 Figure 3-2 Detection of CACNA IS mutations 42 Figure 3-3 R A electrophoresis -+-+ Figure 3-4 Strategy for the amplification of RYR I cDNA 45 Figure 3-5 PCR products for RYR I screening 47 Figure 3-6 Detection of the Tyrl 006 polymorphism 49 Figure 3-7 The position of polymorphisms found in M8 I 8 cDNA 49 Fi gure 3-8 Detection of the sequencing error A 1832G 50 Fi gure 3-9 A lignment of sequences from severa l species 5 1 Figu re -+- I Principle of linkage analys is 5-+ Figure -+-2 The CH family members selected for the genome-wide scan 60 Figure 4-3 A standard curve used for real-time quantification PCR 6 1 Figure 4-4 DNA quantification using a standard curve 62 Figure 4-5 M elting curve analy is 62 Figure 4-6 A genome-wide scan for the CH fam ily 69 Figure 4-7 Candidate loci iclenti fied from the EA branch 7 1 Figure 4-8 A candidate gene on chromosome 16 71 Figure 4-9 Candidate loci on chromosome 6 and 12 73 Figure 5-1 The EA family used for secondary linkage analysis 76 Figure 5-2 Amplification of microsatellite marker loci 79 Figure 5-3 Genotyping using GeneMapper Software 80 IV Fi gure 5-4 Secondary linkage anal ys is on chromosome 16p1 2 82 Figure 5-5 Haplotype of the EA fa mil y on chro mosome 16pl 2 83 LIST OF TABLES Page Tabl e 1-1 Selected RYRJ and CACNA 1S mutati ons 13 Table 2- 1 Typical co nditi ons of PCR therm al cycl es 2 1 Tabl e 2-2 Typical reacti on components fo r PCR 22 Tab le 2-3 The reacti on components fo r Li ghtCycler quantifica ti on 31 Tab le 2-4 The th erm al cyc les fo r Li ghtCycler qu antifi cati on 32 Tab le 2-5 The therm al cycles fo r mi crosatelli te marker analysis 35 Table 2-6 Settin gs fo r linkage analys is 38 Table 3-1 Concent ra ti ons and th e A2601180 ra ti os of ex trac ted RNA samples 44 Table 3-2 PCR and seq uencin g primers fo r screenin g of RYR I cDNA 46 Tabl e 3-3 Polymorphi sms fo un d from RYR 1 cDN A of M8 I 8 48 Table 4-1 Algorithm s and res tricti ons of linkage analys is progra mmes 57 Table 4-2 Factors influencin g co mputati on time 57 Tab le 4-3 The A26011so rat ios and concent rations of 26 indi vid uals 63 Table 4-4 An example of genotyping data 64 Tabl e 4-5 An example of haplotype and all ele frequency 65 Tabl e 4-6 Statu s of indi viduals of the EA branch 70 Table 4-7 Statu s of indi vidu als of the ES branch 72 Tabl e 4-8 Statu s of indi viduals of the TAME branch 74 Tabl e 5-1 Statu s of indi viduals used fo r th e secondary linkage analys is 77 Table 5-2 Microsatellite markers used fo r the second ary linkage analys is 78 Tabl e 5-3 The A26012 so rati os and concentrati ons of additional indi viduals 79 Table 5-4 Primers for CACNG3 screenin g 85 V Table 6-1 The IVCT result of the CH family 92 Table 6-2 Numbers of each disease status in three branches 93 TABLE OF CONTENTS Page Abstract l Abb revi ati011s ll List o/j1gures /l' List o/tables CHAPTER ONE : INTRODUCTION 1.1 Overview of Malignant Hyperthermia 1 I. I. I Introduction 1 1.1.2 Historr 1 1.1.3 Clinical srn1ptoms 2 1.1.4 Triggering agents 2 1.1.5 Mmwgement and treatment 3 1.2 Pathophysiology of MH 3 1.2.J Introduction 3 1.2.2 Excitation-contraction coupling 4 1.2.3 Dihydropyridine receptor 4 1.2.4 Ryanodine receptor 6 1.2.5 The proposed mechanisrn <~f'MH reactions 6 VI 1.3 Diagnosis of MH 7 1.3.
Load more

Recommended publications
  • Autism Multiplex Family with 16P11.2P12.2 Microduplication Syndrome in Monozygotic Twins and Distal 16P11.2 Deletion in Their Brother
    European Journal of Human Genetics (2012) 20, 540–546 & 2012 Macmillan Publishers Limited All rights reserved 1018-4813/12 www.nature.com/ejhg ARTICLE Autism multiplex family with 16p11.2p12.2 microduplication syndrome in monozygotic twins and distal 16p11.2 deletion in their brother Anne-Claude Tabet1,2,3,4, Marion Pilorge2,3,4, Richard Delorme5,6,Fre´de´rique Amsellem5,6, Jean-Marc Pinard7, Marion Leboyer6,8,9, Alain Verloes10, Brigitte Benzacken1,11,12 and Catalina Betancur*,2,3,4 The pericentromeric region of chromosome 16p is rich in segmental duplications that predispose to rearrangements through non-allelic homologous recombination. Several recurrent copy number variations have been described recently in chromosome 16p. 16p11.2 rearrangements (29.5–30.1 Mb) are associated with autism, intellectual disability (ID) and other neurodevelopmental disorders. Another recognizable but less common microdeletion syndrome in 16p11.2p12.2 (21.4 to 28.5–30.1 Mb) has been described in six individuals with ID, whereas apparently reciprocal duplications, studied by standard cytogenetic and fluorescence in situ hybridization techniques, have been reported in three patients with autism spectrum disorders. Here, we report a multiplex family with three boys affected with autism, including two monozygotic twins carrying a de novo 16p11.2p12.2 duplication of 8.95 Mb (21.28–30.23 Mb) characterized by single-nucleotide polymorphism array, encompassing both the 16p11.2 and 16p11.2p12.2 regions. The twins exhibited autism, severe ID, and dysmorphic features, including a triangular face, deep-set eyes, large and prominent nasal bridge, and tall, slender build. The eldest brother presented with autism, mild ID, early-onset obesity and normal craniofacial features, and carried a smaller, overlapping 16p11.2 microdeletion of 847 kb (28.40–29.25 Mb), inherited from his apparently healthy father.
    [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]
  • Altered Stress-Induced Regulation of Genes in Monocytes in Adults with a History of Childhood Adversity
    Neuropsychopharmacology (2016) 41, 2530–2540 © 2016 American College of Neuropsychopharmacology. All rights reserved 0893-133X/16 www.neuropsychopharmacology.org Altered Stress-Induced Regulation of Genes in Monocytes in Adults with a History of Childhood Adversity Marion Schwaiger1, Marianna Grinberg2, Dirk Moser1, Johannes CS Zang1, Markus Heinrichs3,4, Jan G Hengstler5, Jörg Rahnenführer2, Steve Cole6 and Robert Kumsta*,1 1 2 Department of Genetic Psychology, Faculty of Psychology, Ruhr-University Bochum, Bochum, Germany; Department of Statistics, TU Dortmund University, Dortmund, Germany; 3Department of Psychology, Laboratory for Biological and Personality Psychology, University of Freiburg, Freiburg, Germany; 4Freiburg Brain Imaging Center, University Medical Center, University of Freiburg, Freiburg, Germany; 5Leibniz Research Centre for 6 Working Environment and Human Factors at the Technical University of Dortmund (IfADo), Dortmund, Germany; David Geffen School of Medicine, University of California, Los Angeles, CA, USA Exposure to serious or traumatic events early in life can lead to persistent alterations in physiological stress response systems, including enhanced cross talk between the neuroendocrine and immune system. These programming effects may be mechanistically involved in mediating the effects of adverse childhood experience on disease risk in adulthood. We investigated hormonal and genome-wide mRNA = expression responses in monocytes to acute stress exposure, in a sample of healthy adults (n 30) with a history of early childhood = adversity, and a control group (n 30) without trauma experience. The early adversity group showed altered hypothalamus-pituitary- adrenal axis responses to stress, evidenced by lower ACTH and cortisol responses. Analyses of gene expression patterns showed that stress-responsive transcripts were enriched for genes involved in cytokine activity, cytokine–cytokine receptor interaction, chemokine activity, and G-protein coupled receptor binding.
    [Show full text]
  • Absence of S100A4 in the Mouse Lens Induces an Aberrant Retina-Specific Differentiation Program and Cataract
    www.nature.com/scientificreports OPEN Absence of S100A4 in the mouse lens induces an aberrant retina‑specifc diferentiation program and cataract Rupalatha Maddala1*, Junyuan Gao2, Richard T. Mathias2, Tylor R. Lewis1, Vadim Y. Arshavsky1,3, Adriana Levine4, Jonathan M. Backer4,5, Anne R. Bresnick4 & Ponugoti V. Rao1,3* S100A4, a member of the S100 family of multifunctional calcium‑binding proteins, participates in several physiological and pathological processes. In this study, we demonstrate that S100A4 expression is robustly induced in diferentiating fber cells of the ocular lens and that S100A4 (−/−) knockout mice develop late‑onset cortical cataracts. Transcriptome profling of lenses from S100A4 (−/−) mice revealed a robust increase in the expression of multiple photoreceptor‑ and Müller glia‑specifc genes, as well as the olfactory sensory neuron‑specifc gene, S100A5. This aberrant transcriptional profle is characterized by corresponding increases in the levels of proteins encoded by the aberrantly upregulated genes. Ingenuity pathway network and curated pathway analyses of diferentially expressed genes in S100A4 (−/−) lenses identifed Crx and Nrl transcription factors as the most signifcant upstream regulators, and revealed that many of the upregulated genes possess promoters containing a high‑density of CpG islands bearing trimethylation marks at histone H3K27 and/or H3K4, respectively. In support of this fnding, we further documented that S100A4 (−/−) knockout lenses have altered levels of trimethylated H3K27 and H3K4. Taken together,
    [Show full text]
  • A Translaminar Genetic Logic for the Circuit Identity of Intracortically-Projecting Neurons
    bioRxiv preprint doi: https://doi.org/10.1101/290395; this version posted March 28, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Title A translaminar genetic logic for the circuit identity of intracortically-projecting neurons Authors Esther Klingler1, Andres De la Rossa1,3, Sabine Fièvre1, Denis Jabaudon1,2 1Department of Basic Neurosciences, University of Geneva, Geneva, Switzerland. 2Clinic of Neurology, Geneva University Hospital, Geneva, Switzerland. 3Present address: Department of Cell Biology, University of Geneva, Geneva, Switzerland. Correspondence should be addressed to D.J. ([email protected]). Abstract Distinct subtypes of intracortically-projecting neurons (ICPN) are present in all layers, allowing propagation of information within and across cortical columns. How the molecular identities of ICPN relate to their defining anatomical and functional properties is unknown. Here we show that the transcriptional identities of ICPN primarily reflect their input-output connectivities rather than their birth dates or laminar positions. Thus, conserved circuit-related transcriptional programs are at play across cortical layers, which may preserve canonical circuit features across development and evolution. bioRxiv preprint doi: https://doi.org/10.1101/290395; this version posted March 28, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Introduction Neurons of the neocortex are organized into six radial layers, which have appeared at different times during evolution, with the superficial layers representing a more recent acquisition. Input to the neocortex predominantly reaches superficial layers (SL, i.e.
    [Show full text]
  • Gene Expression Profiles of Patients with Cerebral Hematoma Following Spontaneous Intracerebral Hemorrhage
    MOLECULAR MEDICINE REPORTS 10: 1671-1678, 2014 Gene expression profiles of patients with cerebral hematoma following spontaneous intracerebral hemorrhage TAO YANG1,2, JIANWEN GU1, BIN KONG1,2, YONGQIN KUANG1, LIN CHENG1, JINGMIN CHENG1,2, XUN XIA1,2, YUAN MA1,2 and JUNHAI ZHANG1 1Department of Neurosurgery, Chengdu Military General Hospital, Chengdu, Sichuan 610083; 2Department of Postgraduate, Third Military Medical University, Chongqing 400038, P.R. China Received October 13, 2013; Accepted March 27, 2014 DOI: 10.3892/mmr.2014.2421 Abstract. The present study aimed to investigate the gene ITPR1-RYR2, CAMK2A-RYR2 and ITGA5-LAMB1 interac- functions and expression profiles in perihematomal (PH) tion pairs. The present study provides several potential targets brain regions following spontaneous intracerebral hemor- to decrease hematoma expansion and alleviate neuronal cell rhage. The gene expression profiles were downloaded from death following spontaneous intracerebral hemorrhage. the Gene Expression Omnibus database under accession number GSE24265, which includes 11 brain samples from Introduction different regions, including four samples from PH areas, four from contralateral grey matter (CG) and three from contra- Spontaneous intracerebral hemorrhage (ICH) involves the lateral white matter (CW). The gene expression profiles were extravasation of blood within the brain parenchyma in the pre-processed and the differentially expressed genes (DEGs) absence of trauma or surgery, accounting for 10-30% of all between PH and CG tissue, and PH and CW tissue were identi- strokes worldwide (1). ICH is a complex, dynamic process fied using R packages. The expression of genes in different that consists of three distinct phases (2): i) Initial hemorrhage, tissues was analyzed by hierarchical clustering.
    [Show full text]
  • Experimental Eye Research 129 (2014) 93E106
    Experimental Eye Research 129 (2014) 93e106 Contents lists available at ScienceDirect Experimental Eye Research journal homepage: www.elsevier.com/locate/yexer Transcriptomic analysis across nasal, temporal, and macular regions of human neural retina and RPE/choroid by RNA-Seq S. Scott Whitmore a, b, Alex H. Wagner a, c, Adam P. DeLuca a, b, Arlene V. Drack a, b, Edwin M. Stone a, b, Budd A. Tucker a, b, Shemin Zeng a, b, Terry A. Braun a, b, c, * Robert F. Mullins a, b, Todd E. Scheetz a, b, c, a Stephen A. Wynn Institute for Vision Research, The University of Iowa, Iowa City, IA, USA b Department of Ophthalmology and Visual Sciences, Carver College of Medicine, The University of Iowa, Iowa City, IA, USA c Department of Biomedical Engineering, College of Engineering, The University of Iowa, Iowa City, IA, USA article info abstract Article history: Proper spatial differentiation of retinal cell types is necessary for normal human vision. Many retinal Received 14 September 2014 diseases, such as Best disease and male germ cell associated kinase (MAK)-associated retinitis pigmen- Received in revised form tosa, preferentially affect distinct topographic regions of the retina. While much is known about the 31 October 2014 distribution of cell types in the retina, the distribution of molecular components across the posterior pole Accepted in revised form 4 November 2014 of the eye has not been well-studied. To investigate regional difference in molecular composition of Available online 5 November 2014 ocular tissues, we assessed differential gene expression across the temporal, macular, and nasal retina and retinal pigment epithelium (RPE)/choroid of human eyes using RNA-Seq.
    [Show full text]
  • Co-Expression Network Analysis Identifies Possible Hub Genes in Aging of the Human Prefrontal Cortex
    ARTÍCULOS DE INVESTIGACIÓN CLÍNICA O EXPERIMENTAL Co-Expression Network Analysis Identifies Possible Hub Genes in Aging of the Human Prefrontal Cortex Análisis de redes de coexpresión identifica posibles genes clave en el envejecimiento de la corteza prefrontal humana Análise de redes de co-expressão identifica possíveis genes chave no envelhecimento do córtex pré-frontal humano César Payán-Gómez, MD, MSc, PhD1*; Julián Riaño-Moreno, MD, MSc2; Diana Amador-Muñoz, MD, MSc3; Sandra Ramírez-Clavijo, MSc, PhD4 Received: December 4th, 2018 - Accepted: March 15th, 2019 Doi: http://dx.doi.org/10.12804/10.12804/revistas.urosario.edu.co/revsalud/a.7924 To cite this article: Payán-Gómez C, Riaño-Moreno J, Amador-Muñoz D, Ramírez-Clavijo S. Co-expression Network Analysis Identifies Possible Hub Genes in Aging of the Human Prefrontal Cortex. Rev Cienc Salud. 2019;17(2):201-22. Doi: http://dx.doi.org/10.12804/10.12804/ revistas.urosario.edu.co/revsalud/a.7924 Abstract Introduction: Aging is the main risk factor for the development of chronic diseases such as cancer, diabetes, Parkinson’s disease, and Alzheimer’s disease. The central nervous system is particularly susceptible to progressive functional deterioration associated with age, among the brain regions the prefrontal cortex (PFC) has one of the highest involvements. Transcriptomics studies of this brain region have identified the decrease in synaptic function and activation of neuroglia cells as fundamental characteristics of the aging process. The aim of this study was to identify hub genes in the transcriptomic deregulation in the PFC aging to advance in the knowledge of this process. Materials and methods: A gene co-expression analysis was carried 1 Faculty of Natural Sciences and Mathematics, Universidad del Rosario, Bogota, Colombia.
    [Show full text]
  • Stargazin and Other Transmembrane AMPA Receptor Regulating Proteins Interact with Synaptic Scaffolding Protein MAGI-2 in Brain
    The Journal of Neuroscience, July 26, 2006 • 26(30):7875–7884 • 7875 Cellular/Molecular Stargazin and Other Transmembrane AMPA Receptor Regulating Proteins Interact with Synaptic Scaffolding Protein MAGI-2 in Brain Fang Deng, Maureen G. Price, Caleb F. Davis, Mayra Mori, and Daniel L. Burgess Department of Neurology, Baylor College of Medicine, Houston, Texas 77030 The spatial coordination of neurotransmitter receptors with other postsynaptic signaling and structural molecules is regulated by a diverse array of cell-specific scaffolding proteins. The synaptic trafficking of AMPA receptors by the stargazin protein in some neurons, for example, depends on specific interactions between the C terminus of stargazin and the PDZ [postsynaptic density-95 (PSD-95)/Discs large/zona occludens-1] domains of membrane-associated guanylate kinase scaffolding proteins PSD-93 or PSD-95. Stargazin [Cacng2 (Ca 2ϩ channel ␥2 subunit)] is one of four closely related proteins recently categorized as transmembrane AMPA receptor regulating proteins (TARPs) that appear to share similar functions but exhibit distinct expression patterns in the CNS. We used yeast two-hybrid screeningtoidentifyMAGI-2(membraneassociatedguanylatekinase,WWandPDZdomaincontaining2)asanovelcandidateinteractor with the cytoplasmic C termini of the TARPs. MAGI-2 [also known as S-SCAM (synaptic scaffolding molecule)] is a multi-PDZ domain scaffolding protein that interacts with several different ligands in brain, including PTEN (phosphatase and tensin homolog), dasm1 (dendrite arborization and synapse maturation 1), dendrin, axin, ␤- and ␦-catenin, neuroligin, hyperpolarization-activated cation channels, ␤1-adrenergic receptors, and NMDA receptors. We confirmed that MAGI-2 coimmunoprecipitated with stargazin in vivo from mouse cerebral cortex and used in vitro assays to localize the interaction to the C-terminal -TTPV amino acid motif of stargazin and the PDZ1, PDZ3, and PDZ5 domains of MAGI-2.
    [Show full text]
  • Linkage and Association Analysis of CACNG3 in Childhood Absence Epilepsy
    1 Linkage and association analysis of CACNG3 in childhood absence epilepsy Kate Everett1,*, Barry Chioza1,*, Jean Aicardi2, Harald Aschauer3, Oebele Brouwer4, Petra Callenbach4, Athanasios Covanis5, Olivier Dulac6, Orvar Eeg-Olofsson7, Martha Feucht8, Mogens Friis9, Françoise Goutieres10, Renzo Guerrini11, Armin Heils12, Marianne Kjeldsen13, Anna-Elina Lehesjoki14, Andrew Makoff15, Rima Nabbout6, Ingrid Olsson16, Thomas Sander17,18, Auli Sirén19, Paul McKeigue20, Robert Robinson1,†, Nichole Taske1‡, Michele Rees1, Mark Gardiner1 *These two authors contributed equally to the work. †Present affiliation: Paediatric Neurology, Great Ormond Street Hospital, London, UK. ‡Present affiliation: National Institute for Health and Clinical Excellence (NICE), London, UK. 1Department of Paediatrics and Child Health, Royal Free and University College Medical School, University College London, UK; 2Hôpital Robert Debré, Paris Cedex 19, France; 3Department of General Psychiatry, Medical University Vienna, Austria; 4University Medical Centre Groningen, University of Groningen, The Netherlands; 5Neurology Department, The Children’s Hospital ‘Agia Sophia’, Athens, Greece; 6Neuropaediatrics Department, Hôpital Necker Enfant Malades, Paris, France; 7Department of Women’s and Children’s Health/Neuropaediatrics, Uppsala University, Sweden; 8Department of Paediatrics, Medical University Vienna, Austria; 9Department of Neurology, Sygehus Vestsjaelland, Holbaek, Denmark; 10No institutional affiliation; 11Division of Child Neurology and Psychiatry, University of
    [Show full text]
  • Identifying Nootropic Drug Targets Via Large-Scale Cognitive GWAS and Transcriptomics
    bioRxiv preprint doi: https://doi.org/10.1101/2020.02.06.934752; this version posted February 6, 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. Title: Identifying Nootropic Drug Targets via Large-Scale Cognitive GWAS and Transcriptomics Max Lam1, 2, 3,4, Chia-Yen, Chen3,5,6, Xia Yan7,8, W. David Hill9, 10, Joey W. Trampush11, Jin Yu1, Emma Knowles12,13,14, Gail Davies9, 10, Eli Stahl15, 16, Laura Huckins15, 16, David C. Liewald10, Srdjan Djurovic17, 18, Ingrid Melle18, 19, Andrea Christoforou20, Ivar Reinvang21, Pamela DeRosse1, 22, 23, Astri J. Lundervold24, Vidar M. Steen18, 20, Thomas Espeseth19, 21, Katri Räikkönen25, Elisabeth Widen26, Aarno Palotie26, 27, 28, Johan G. Eriksson29, 30, 31, Ina Giegling32, Bettina Konte32, Annette M. Hartmann32, Panos Roussos15, 16, 33, Stella Giakoumaki34, Katherine E. Burdick15, 33, 35, Antony Payton36, William Ollier37, 38, Ornit Chiba- Falek39, Deborah K. Koltai39, 40 , Anna C. Need41, Elizabeth T. Cirulli42, Aristotle N. Voineskos43, Nikos C. Stefanis44, 45, 46, Dimitrios Avramopoulos47, 48, Alex Hatzimanolis44, 45, 46, Nikolaos Smyrnis44, 45, Robert M. Bilder49, Nelson A. Freimer49, Tyrone D. Cannon50, 51, Edythe London49, Russell A. Poldrack52, Fred W. Sabb53, Eliza Congdon49, Emily Drabant Conley54, Matthew A. Scult55, Dwight Dickinson56, Richard E. Straub57, Gary Donohoe58, Derek Morris58, Aiden Corvin59, Michael Gill59, Ahmad R. Hariri55, Daniel R. Weinberger57, Neil Pendleton60, Panos Bitsios61, Dan Rujescu32, Jari Lahti25, 62, Stephanie Le Hellard18, 20, Matthew C.
    [Show full text]
  • Neurons Derived from ASD Induced Pluripotent Stem Cells Sahar Avazzadeh1, Katya Mcdonagh1, Jamie Reilly1, Yanqin Wang1,2, Stephanie D
    Avazzadeh et al. Molecular Autism (2019) 10:52 https://doi.org/10.1186/s13229-019-0303-3 RESEARCH Open Access Increased Ca2+ signaling in NRXN1α+/− neurons derived from ASD induced pluripotent stem cells Sahar Avazzadeh1, Katya McDonagh1, Jamie Reilly1, Yanqin Wang1,2, Stephanie D. Boomkamp1, Veronica McInerney3, Janusz Krawczyk4, Jacqueline Fitzgerald5, Niamh Feerick5, Matthew O’Sullivan5, Amirhossein Jalali6, Eva B. Forman7, Sally A. Lynch8,9,10, Sean Ennis11, Nele Cosemans12, Hilde Peeters10, Peter Dockery13, Timothy O’Brien1, Leo R. Quinlan14, Louise Gallagher5* and Sanbing Shen1* Abstract Background: Autism spectrum disorder (ASD) is a neurodevelopmental disorder with a high co-morbidity of epilepsy and associated with hundreds of rare risk factors. NRXN1 deletion is among the commonest rare genetic factors shared by ASD, schizophrenia, intellectual disability, epilepsy, and developmental delay. However, how NRXN1 deletions lead to different clinical symptoms is unknown. Patient-derived cells are essential to investigate the functional consequences of NRXN1 lesions to human neurons in different diseases. Methods: Skin biopsies were donated by five healthy donors and three ASD patients carrying NRXN1α+/− deletions. Seven control and six NRXN1α+/− iPSC lines were derived and differentiated into day 100 cortical excitatory neurons using dual SMAD inhibition. Calcium (Ca2+) imaging was performed using Fluo4-AM, and the properties of Ca2+ transients were compared between two groups of neurons. Transcriptome analysis was carried out to undercover molecular pathways associated with NRXN1α+/− neurons. Results: NRXN1α+/− neurons were found to display altered calcium dynamics, with significantly increased frequency, duration, and amplitude of Ca2+ transients. Whole genome RNA sequencing also revealed altered ion transport and transporter activity, with upregulated voltage-gated calcium channels as one of the most significant pathways in NRXN1α+/− neurons identified by STRING and GSEA analyses.
    [Show full text]