DOCSLIB.ORG

Copy Number Variation and Huntington's Disease

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Copy Number Variation and Huntington's Disease UNIVERSIDADE DE LISBOA FACULDADE DE MEDICINA DE LISBOA Copy number variation and Huntington’s disease Angelica Vittori Ramo das Ciências Biomédicas Especialidade – Neurociências Outubro 2013 UNIVERSIDADE DE LISBOA FACULDADE DE MEDICINA DE LISBOA Copy number variation and Huntington’s disease Candidata: Angelica Vittori Orientadores: Prof . Doutor Tiago Fleming Outeiro Doutor Flaviano Giorgini Doutor Edward J. Hollox Ramo das Ciências Biomédicas Especialidade – Neurociências Todas as afirmações efectuadas no presente documento são da exclusiva II responsabilidade do seu autor, não cabendo qualquer responsabilidade à faculdade de medicina de lisboa pelos conteúdos nela apresentados. A impressão são da exclusiva está dissertação foi aprovada pelo Conselho Cientifico da Faculdade de Medicina em reunião de 19 de Novembro de 2013. III Resumo A variação de número de cópias (CNV em inglês) é uma modificação de uma sequência de DNA que apresenta uma inserção ou deleção em comparação com um genoma de referência com um número de cópias de N = 2. Com um comprimento variável, desde 50 pares de bases até várias megabases, as CNVs identificadas têm um tamanho médio de ~ 3 Kb e representam cerca de 4% do genoma humano. As CNVs, como outras variações genéticas, podem afetar directamente os níveis de expressão dos genes afectados. Os efeitos indirectos na expressão genética podem ser causados por alterações da posição, interrompendo o quadro de leitura do gene ou posteriormente, perturbando as redes de regulação genética. Foi demonstrado que as CNVs são em grande parte responsáveis pela evolução humana e diversidade genética entre os indivíduos. A relevância das CNVs no genoma humano foi salientada por vários estudos de associação que mostraram o efeito das CNVs na susceptibilidade a doenças neurodegenerativas, doenças de características complexas, e por serem a principal causa do aparecimento de doenças mendelianas ou por conferirem um fenótipo benigno. A doença de Huntington (DH) é uma doença degenerativa e progressiva do cérebro que é fatal. Clínicamente é caracterizada por disfunção motora, distúrbios emocionais / psiquiátricos graves, declínio cognitivo e demência. A DH é monogénica e autossómica dominante, sendo causada por uma expansão anormal do trinucleótido citosina-adenina- guanina (CAG) no terminal 5' do gene da Huntingtina (HTT), localizado no cromossoma 4. Normalmente, a manifestação dos sintomas da doença dá-se por volta dos 40 anos, mas formas mais precoces (início <20 anos) e mais tardias (> 70 anos) também existem. O comprimento da expansão aberrante da repetição CAG no gene HTT está inversamente correlacionado com a idade de ocorrência (IO) da doença. O comprimento desta repetição explica ~ 50-70% na variação da IO sendo o restante da variação explicado por outros fatores genéticos e ambientais. Vários estudos têm como objectivo identificar modificadores genéticos da IO em DH, ou seja, nomeadamente um gene que quando IV mutado tem a sua função alterada e determina o aparecimento do fenótipo associado com a mutação CAG. Há poucos modificadores genéticos identificados através de abordagens multi-translacionais de grandes estudos de coorte da doença para validação em vários modelos experimentais, representando, no entanto uma abordagem válida para identificar novos alvos terapêuticos para a DH. O nosso objetivo neste estudo foi identificar genes candidatos com variação de número de cópias com potencial para modificar a IO na DH. Após a triagem de uma biblioteca de CNVs a partir de análise de dados matriz-CGH (disponível em linha através do navegador genoma UCSC), foram seleccionados vários candidatos: a região da β-defensine (DEFB4) e o transportador de glicose neuronal 3 (SLC2A3). Colocámos a hipótese de que a CNV na região de β-defensine (que varia de 2 a 8 cópias) poderia afectar o início da DH através de uma expressão variável do gene DEFB4 (presente nesta região) que codifica para um péptido antimicrobiano importante. Um aumento do número de cópias, codificantes para um nível mais elevado de proteína, podem contribuir a inflamação neuronal presente na DH e consequentemente piorando a evolução da doença. Assim, investigámos o papel potencial da CNV do gene SLC2A3 (que variam de 1 a 3 cópias), assumindo que o maior número de cópias pode ser protector na DH, codificando para um nível mais elevado do transportador de glicólise (GLUT3). Analisámos a distribuição das CNVs dos nossos genes candidatos em indivíduos do estudo REGISTRY realizado pela Grupo Europeu da Doença de Huntington (EHDN), utilizando o Teste da Razão de Parálogos (PRT). PRT é uma tipagem de elevada capacidade de variação do número de cópias que se baseia em PCR quantitativo e relativo. Determinámos o número de cópias da β-defensine em 490 indivíduos da nossa coorte de DH. A distribuição de CNVs de β-defensine nesta coorte não é significativamente diferente da população europeia sem DH (valor de p= 0,91) e também não está associada a uma alteração significativa da IO (valor de p = 0,41). Assim, podemos concluir que a CNV de β-defensine não é relevante na patologia da DH durante a fase inicial da progressão da doença. No entanto, com o nosso estudo, não podemos descartar qualquer papel potencial do gene DEFB4 numa outra fase da doença. V Determinámos depois a distribuição de CNVs no gene SLC2A3 em 988 indivíduos com a DH e não foi significativamente diferente da população britânica (valor de p =0.38). Curiosamente, verificou-se uma associação significativa entre a CNV de SLC2A3 e uma variação de IO na nossa coorte de DH (valor de p = 0,028). Indivíduos com três cópias de SLC2A3 mostraram um atraso do IO em comparação com o resto do grupo de até cerca de 6 anos. De modo a investigar a base molecular deste efeito protector da GLUT3 analisámos os níveis de expressão da proteína em linhas linfocíticas imortalizadas obtidas a partir de pacientes da nossa coorte, portadores de 1, 2 e 3 cópias de SLC2A3 (5 por cada genótipo). Pela análise immunoblot detectou-se que um maior número de cópias no gene SLC2A3 está associado a um maior nível de expressão da proteína (valor de p = 0,02). Assim, o efeito protector do aumento do número de cópias de SLC2A3 é devido a uma maior expressão do transportador de glicose. Para investigar a base funcional deste efeito protector, foi utilizado um modelo de Drosophila de DH para estudar o efeito de sobre-ou sub-expressão do funcional homólogo de SLC2A3 em Drosophila, Glut1. Analisámos vários indicadores relevantes da doença, incluindo a neurodegeneração dos fotorreceptores, longevidade e taxa de eclosão. Descobrimos que em moscas com DH a sobreexpressão de Glut1 pode reduzir significativamente a perda de rhabdomeres e portanto a neurodegeneração (valor de p <0,001). Pelo contrário, a redução ou a alteração génica deste transportador estão associados com um pior cenário que se manifesta por uma perda maior de rhabdomeres (valor de p = 0,00012 *), uma redução da longevidade média (valor de p = 0,034) e uma redução da taxa de eclosão (valor de p = 0,015). Alterações do principal transportador de glicose num modelo de Drosophila da DH agrava a progressão da doença numa fase inicial, e sobrerexpressão deste transportador protege da neurodegeneração na mosca adulta, confirmando os resultados na coorte de DH. Do que sabemos, este é o primeiro estudo que investigou a função potencial das CNVs em DH. Conclui-se que a CNV no gene SLC2A3 é um novo modificador genético do IO em grandes coortes de DH (988 amostras), em que un aumento de cópias do gene está associado a um efeito protector na DH. Testámos o efeito molecular da ocorrência desta CNV em linhas celulares obtidas directamente dos pacientes e descobrimos que o evento de VI CNV do gene é responsável por um aumento da expressão de GLUT3 na DH, o que poderá repor a deficiência energética nos neurônios de pacientes com a DH e atrasar a neurodegeneração. Além disso, usámos um modelo de Drosophila de DH para testar a função funcional da homólogo de SLC2A3 em moscas (Glut1), confirmando que uma sobre- expressão deste transportador pode diminuir a progressão da doença na mosca adulta, e a alteração do gene piora a progressão da doença num estadio inicial. É necessária futura investigação para reproduzir esta observação e confirmar o SLC2A3 como um alvo de potencial terapeutico para o tratamento da DH. * os dados referem-se a moscas no dia 1 pós eclosão. Palavras-chave: variação do número de cópia (CNV), doença de Huntington (DH), modificadores genéticos, DEFB4, SLC2A3, Drosophila. VII Index RESUMO ...................................................................................................................................IV INDEX .....................................................................................................................................VIII INDEX OF THE FIGURES .................................................................................................................. X INDEX OF THE TABLES .................................................................................................................. XII ABBREVIATIONS ....................................................................................................................... XIV ABSTRACT ............................................................................................................................. XVII CHAPTER ONE: INTRODUCTION .................................................................... 1 1.1 COPY NUMBER VARIATION .....................................................................................................
Load more

Recommended publications
  • (Lcrs) in 22Q11 Mediate Deletions, Duplications, Translocations, and Genomic Instability: an Update and Literature Review Tamim H
    review January/February 2001 ⅐ Vol. 3 ⅐ No. 1 Evolutionarily conserved low copy repeats (LCRs) in 22q11 mediate deletions, duplications, translocations, and genomic instability: An update and literature review Tamim H. Shaikh, PhD1, Hiroki Kurahashi, MD, PhD1, and Beverly S. Emanuel, PhD1,2 Several constitutional rearrangements, including deletions, duplications, and translocations, are associated with 22q11.2. These rearrangements give rise to a variety of genomic disorders, including DiGeorge, velocardiofacial, and conotruncal anomaly face syndromes (DGS/VCFS/CAFS), cat eye syndrome (CES), and the supernumerary der(22)t(11;22) syndrome associated with the recurrent t(11;22). Chromosome 22-specific duplications or low copy repeats (LCRs) have been directly implicated in the chromosomal rearrangements associated with 22q11.2. Extensive sequence analysis of the different copies of 22q11 LCRs suggests a complex organization. Examination of their evolutionary origin suggests that the duplications in 22q11.2 may predate the divergence of New World monkeys 40 million years ago. Based on the current data, a number of models are proposed to explain the LCR-mediated constitutional rearrangements of 22q11.2. Genetics in Medicine, 2001:3(1):6–13. Key Words: duplication, evolution, 22q11, deletion and translocation Although chromosome 22 represents only 2% of the haploid The 22q11.2 deletion syndrome, which includes DGS/ human genome,1 recurrent, clinically significant, acquired, VCFS/CAFS, is the most common microdeletion syndrome. and somatic
    [Show full text]
  • Gentles Et Al 2007 Gen Res.Pdf
    Downloaded from www.genome.org on July 2, 2007 Letter Evolutionary dynamics of transposable elements in the short-tailed opossum Monodelphis domestica Andrew J. Gentles,1,2,6 Matthew J. Wakefield,3 Oleksiy Kohany,2 Wanjun Gu,4 Mark A. Batzer,5 David D. Pollock,4 and Jerzy Jurka2,6 1Department of Radiology, School of Medicine, Stanford University, Stanford, California 94305, USA; 2Genetic Information Research Institute, Mountain View, California 94043, USA; 3ARC Centre for Kangaroo Genomics, Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3050, Australia; 4Department of Biochemistry and Molecular Genetics, University of Colorado Health Sciences Center, Aurora 80045, Colorado, USA; 5Department of Biological Sciences, Biological Computation and Visualization Center, Center for BioModular Multi-Scale Systems, Louisiana State University, Baton Rouge, Louisiana 70803, USA The genome of the gray short-tailed opossum Monodelphis domestica is notable for its large size (∼3.6 Gb). We characterized nearly 500 families of interspersed repeats from the Monodelphis. They cover ∼52% of the genome, higher than in any other amniotic lineage studied to date, and may account for the unusually large genome size. In comparison to other mammals, Monodelphis is significantly rich in non-LTR retrotransposons from the LINE-1, CR1, and RTE families, with >29% of the genome sequence comprised of copies of these elements. Monodelphis has at least four families of RTE, and we report support for horizontal transfer of this non-LTR retrotransposon. In addition to short interspersed elements (SINEs) mobilized by L1, we found several families of SINEs that appear to use RTE elements for mobilization. In contrast to L1-mobilized SINEs, the RTE-mobilized SINEs in Monodelphis appear to shift from G+C-rich to G+C-low regions with time.
    [Show full text]
  • Repetitive Elements in Humans
    International Journal of Molecular Sciences Review Repetitive Elements in Humans Thomas Liehr Institute of Human Genetics, Jena University Hospital, Friedrich Schiller University, Am Klinikum 1, D-07747 Jena, Germany; [email protected] Abstract: Repetitive DNA in humans is still widely considered to be meaningless, and variations within this part of the genome are generally considered to be harmless to the carrier. In contrast, for euchromatic variation, one becomes more careful in classifying inter-individual differences as meaningless and rather tends to see them as possible influencers of the so-called ‘genetic background’, being able to at least potentially influence disease susceptibilities. Here, the known ‘bad boys’ among repetitive DNAs are reviewed. Variable numbers of tandem repeats (VNTRs = micro- and minisatellites), small-scale repetitive elements (SSREs) and even chromosomal heteromorphisms (CHs) may therefore have direct or indirect influences on human diseases and susceptibilities. Summarizing this specific aspect here for the first time should contribute to stimulating more research on human repetitive DNA. It should also become clear that these kinds of studies must be done at all available levels of resolution, i.e., from the base pair to chromosomal level and, importantly, the epigenetic level, as well. Keywords: variable numbers of tandem repeats (VNTRs); microsatellites; minisatellites; small-scale repetitive elements (SSREs); chromosomal heteromorphisms (CHs); higher-order repeat (HOR); retroviral DNA 1. Introduction Citation: Liehr, T. Repetitive In humans, like in other higher species, the genome of one individual never looks 100% Elements in Humans. Int. J. Mol. Sci. alike to another one [1], even among those of the same gender or between monozygotic 2021, 22, 2072.
    [Show full text]
  • Alpha Defensin 1 Antibody / DEFA1 (R32739)
    Alpha Defensin 1 Antibody / DEFA1 (R32739) Catalog No. Formulation Size R32739 0.5mg/ml if reconstituted with 0.2ml sterile DI water 100 ug Bulk quote request Availability 1-3 business days Species Reactivity Human, Rat Format Antigen affinity purified Clonality Polyclonal (rabbit origin) Isotype Rabbit IgG Purity Antigen affinity Buffer Lyophilized from 1X PBS with 2.5% BSA, 0.025% sodium azide UniProt P59665 Applications Western Blot : 0.5-1ug/ml Limitations This Alpha Defensin 1 antibody is available for research use only. Western blot testing of 1) rat testis and 2) human HeLa lysate with Alpha Defensin 1 antibody at 0.5ug/ml. Predicted molecular weight ~10 kDa. Description Defensin, alpha 1, also known as human alpha defensin 1, human neutrophil peptide 1 (HNP-1) or neutrophil defensin 1 is a human protein that is encoded by the DEFA1 gene. Defensins are a family of antimicrobial and cytotoxic peptides thought to be involved in host defense. They are abundant in the granules of neutrophils and also found in the epithelia of mucosal surfaces such as those of the intestine, respiratory tract, urinary tract, and vagina. Members of the defensin family are highly similar in protein sequence and distinguished by a conserved cysteine motif. The protein encoded by this gene, defensin, alpha 1, is found in the microbicidal granules of neutrophils and likely plays a role in phagocyte-mediated host defense. Several alpha defensin genes are clustered on chromosome 8. This gene differs from defensin, alpha 3 by only one amino acid. This gene and the gene encoding defensin, alpha 3 are both subject to copy number variation.
    [Show full text]
  • Recent Advance in Our Understanding of the Molecular Nature of Chromosomal Abnormalities
    Journal of Human Genetics (2009) 54, 253–260 & 2009 The Japan Society of Human Genetics All rights reserved 1434-5161/09 $32.00 www.nature.com/jhg REVIEW Recent advance in our understanding of the molecular nature of chromosomal abnormalities Hiroki Kurahashi, Hasbaira Bolor, Takema Kato, Hiroshi Kogo, Makiko Tsutsumi, Hidehito Inagaki and Tamae Ohye The completion of the human genome project has enabled researchers to characterize the breakpoints for various chromosomal structural abnormalities including deletions, duplications or translocations. This in turn has shed new light on the molecular mechanisms underlying the onset of gross chromosomal rearrangements. On the other hand, advances in genetic manipulation technologies for various model organisms has increased our knowledge of meiotic chromosome segregation, errors which, contribute to chromosomal aneuploidy. This review focuses on the current understanding of germ line chromosomal abnormalities and provides an overview of the mechanisms involved. We refer to our own recent data and those of others to illustrate some of the new paradigms that have arisen in this field. We also discuss some perspectives on the sexual dimorphism of some of the pathways that leads to these chromosomal abnormalities. Journal of Human Genetics (2009) 54, 253–260; doi:10.1038/jhg.2009.35; published online 17 April 2009 Keywords: aneuploidy; cruciform; deletion; gross chromosomal rearrangement; low-copy repeat; non-disjunction; palindrome; translocation THE RANDOM NATURE OF GROSS CHROMOSOMAL inverted repeats and the critical break points often reside within these REARRANGEMENTS regions. When a DSB occurs within one copy of a repeat segment, The development of chromosomal structural abnormalities, also illegitimate DSB repair through the HR pathway using another copy of known as gross chromosomal rearrangements (GCR), is essentially the repeat segments may be induced, and this is the most likely dependent on two distinct processes: double-strand breaks (DSB) and mechanism of recurrent GCR.
    [Show full text]
  • DEFA1 Rabbit Pab
    Leader in Biomolecular Solutions for Life Science DEFA1 Rabbit pAb Catalog No.: A6897 Basic Information Background Catalog No. Defensins are a family of antimicrobial and cytotoxic peptides thought to be involved in host A6897 defense. They are abundant in the granules of neutrophils and also found in the epithelia of mucosal surfaces such as those of the intestine, respiratory tract, urinary tract, and vagina. Observed MW Members of the defensin family are highly similar in protein sequence and distinguished by a 10KDa conserved cysteine motif. The protein encoded by this gene, defensin, alpha 1, is found in the microbicidal granules of neutrophils and likely plays a role in phagocyte-mediated host defense. Several alpha defensin genes are clustered on chromosome 8. This gene differs Calculated MW from defensin, alpha 3 by only one amino acid. This gene and the gene encoding defensin, 10kDa alpha 3 are both subject to copy number variation. Category Primary antibody Applications WB, IHC, IF Cross-Reactivity Human, Mouse, Rat Recommended Dilutions Immunogen Information WB 1:500 - 1:2000 Gene ID Swiss Prot 1667 P59665 IHC 1:50 - 1:100 Immunogen IF 1:50 - 1:100 Recombinant fusion protein containing a sequence corresponding to amino acids 1-94 of human DEFA1 (NP_004075.1). Synonyms DEFA1;DEF1;DEFA2;HNP-1;HP-1;HP1;MRS Contact Product Information 400-999-6126 Source Isotype Purification Rabbit IgG Affinity purification [email protected] Storage www.abclonal.com.cn Store at -20℃. Avoid freeze / thaw cycles. Buffer: PBS with 0.02% sodium azide,50% glycerol,pH7.3. Validation Data Western blot analysis of extracts of various cell lines, using DEFA1 antibody (A6897) at 1:1000 dilution.
    [Show full text]
  • Papain-Like Protease Regulates SARS-Cov-2 Viral Spread and Innate Immunity
    Article Papain-like protease regulates SARS-CoV-2 viral spread and innate immunity https://doi.org/10.1038/s41586-020-2601-5 Donghyuk Shin1,2,3, Rukmini Mukherjee1,2, Diana Grewe2, Denisa Bojkova4, Kheewoong Baek5, Anshu Bhattacharya1,2, Laura Schulz6, Marek Widera4, Ahmad Reza Mehdipour6, Georg Tascher1, Received: 30 April 2020 Paul P. Geurink7, Alexander Wilhelm4,8, Gerbrand J. van der Heden van Noort7, Huib Ovaa7,13, Accepted: 23 July 2020 Stefan Müller1, Klaus-Peter Knobeloch9, Krishnaraj Rajalingam10, Brenda A. Schulman5, Jindrich Cinatl4, Gerhard Hummer6,11, Sandra Ciesek4,8,12 & Ivan Dikic1,2,3,12 ✉ Published online: 29 July 2020 Check for updates The papain-like protease PLpro is an essential coronavirus enzyme that is required for processing viral polyproteins to generate a functional replicase complex and enable viral spread1,2. PLpro is also implicated in cleaving proteinaceous post-translational modifcations on host proteins as an evasion mechanism against host antiviral immune responses3–5. Here we perform biochemical, structural and functional characterization of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) PLpro (SCoV2-PLpro) and outline diferences with SARS-CoV PLpro (SCoV-PLpro) in regulation of host interferon and NF-κB pathways. SCoV2-PLpro and SCoV-PLpro share 83% sequence identity but exhibit diferent host substrate preferences; SCoV2-PLpro preferentially cleaves the ubiquitin-like interferon-stimulated gene 15 protein (ISG15), whereas SCoV-PLpro predominantly targets ubiquitin chains. The crystal structure of SCoV2-PLpro in complex with ISG15 reveals distinctive interactions with the amino-terminal ubiquitin-like domain of ISG15, highlighting the high afnity and specifcity of these interactions.
    [Show full text]
  • DEFA1) Antibody Catalogue No.:Abx005247
    Datasheet Version: 1.0.0 Revision date: 19 Nov 2020 Defensin Alpha 1, Neutrophil (DEFA1) Antibody Catalogue No.:abx005247 Western blot analysis of extracts of various cell lines, using DEFA1 antibody (abx005247) at 1/1000 dilution. Immunohistochemistry of paraffin-embedded rat heart using DEFA1 antibody (abx005247) at dilution of 1/100 (40x lens). Immunofluorescence analysis of HeLa cells using DEFA1 antibody (abx005247). Blue: DAPI for nuclear staining. DEFA1 Antibody is a Rabbit Polyclonal antibody against DEFA1. Defensins are a family of antimicrobial and cytotoxic peptides thought to be involved in host defense. They are abundant in the granules of neutrophils and also found in the epithelia of mucosal surfaces such as those of the intestine, respiratory tract, urinary tract, and vagina. Members of the defensin family are highly similar in protein sequence and distinguished by a conserved cysteine motif. The protein encoded by this gene, defensin, alpha 1, is found in the microbicidal granules of neutrophils and likely plays a role in phagocyte-mediated host defense. Several alpha defensin genesFor are clustered on chromosomeReference 8. This gene differs from defensin, alpha Only 3 by only one amino acid. This gene and the gene encoding defensin, alpha 3 are both subject to copy number variation. Target: DEFA1 Clonality: Polyclonal Reactivity: Human, Mouse, Rat v1.0.0 Abbexa Ltd, Cambridge, UK · Phone: +44 1223 755950 · Fax: +44 1223 755951 1 Abbexa LLC, Houston, TX, USA · Phone: +1 832 327 7413 www.abbexa.com · Email: [email protected] Datasheet Version: 1.0.0 Revision date: 19 Nov 2020 Tested Applications: WB, IHC, IF/ICC Host: Rabbit Recommended dilutions: WB: 1/500 - 1/2000, IHC: 1/50 - 1/100, IF/ICC: 1/50 - 1/100.
    [Show full text]
  • Downloaded from the Tranche Distributed File System (Tranche.Proteomecommons.Org) and Ftp://Ftp.Thegpm.Org/Data/Msms
    Research Article Title: The shrinking human protein coding complement: are there now fewer than 20,000 genes? Authors: Iakes Ezkurdia1*, David Juan2*, Jose Manuel Rodriguez3, Adam Frankish4, Mark Diekhans5, Jennifer Harrow4, Jesus Vazquez 6, Alfonso Valencia2,3, Michael L. Tress2,*. Affiliations: 1. Unidad de Proteómica, Centro Nacional de Investigaciones Cardiovasculares, CNIC, Melchor Fernández Almagro, 3, rid, 28029, MadSpain 2. Structural Biology and Bioinformatics Programme, Spanish National Cancer Research Centre (CNIO), Melchor Fernández Almagro, 3, 28029, Madrid, Spain 3. National Bioinformatics Institute (INB), Spanish National Cancer Research Centre (CNIO), Melchor Fernández Almagro, 3, 28029, Madrid, Spain 4. Wellcome Trust Sanger Institute, Wellcome Trust Campus, Hinxton, Cambridge CB10 1SA, UK 5. Center for Biomolecular Science and Engineering, School of Engineering, University of California Santa Cruz (UCSC), 1156 High Street, Santa Cruz, CA 95064, USA 6. Laboratorio de Proteómica Cardiovascular, Centro Nacional de Investigaciones Cardiovasculares, CNIC, Melchor Fernández Almagro, 3, 28029, Madrid, Spain *: these two authors wish to be considered as joint first authors of the paper. Corresponding author: Michael Tress, [email protected], Tel: +34 91 732 80 00 Fax: +34 91 224 69 76 Running title: Are there fewer than 20,000 protein-coding genes? Keywords: Protein coding genes, proteomics, evolution, genome annotation Abstract Determining the full complement of protein-coding genes is a key goal of genome annotation. The most powerful approach for confirming protein coding potential is the detection of cellular protein expression through peptide mass spectrometry experiments. Here we map the peptides detected in 7 large-scale proteomics studies to almost 60% of the protein coding genes in the GENCODE annotation the human genome.
    [Show full text]
  • Human Genetics Lecture #1 March 25, 2014
    Bio 5491 - Advanced Genetics Human Genetics Lecture #1 March 25, 2014 Cristina de Guzman Strong, Ph.D. Department of Medicine Dermatology/Pharmacogenomics McDonnell Basic Sciences 770 [email protected] 362-7695 Human Genetics • Karyotype • Mutations in Regulatory • Genetic Variants Elements • Mendelian Diseases • Copy number variation • Linkage, positional cloning diseases • Penetrance/Expressivity • Mitochondrial genetics • Human Genome Project • Human-specific variation • GWAS/Next-Gen • Future sequencing (Exome) • Undiagnosed Diseases • Epigenetics/ENCODE What is Human Genetics? The relationship between natural DNA sequence variation(s) and human phenotypic traits What is different about Human Genetics? • Imprinting……..uniquely mammalian. • Trinucleotide repeat diseases…….anticipation. • One can study complex behaviours and cognition. • Extensive sequence variation leads to common/ complex disease 1. Common disease – common variant hypothesis 2. Large # of small-effect variants 3. Large # of large-effect rare variants 4. Combo of genotypic, environmental, epigenetic interactions Greg Gibson, Nature Review Gen 2012 Human Genome (Karyotype) 22 autosomes/ XY sex chromosomes 02_13.jpg Human genome is ~41% GC, but that is non-randomly distributed. Dark G-bands are lower GC (and lower gene content) Genetic variation: Single Base Pair C T – SNP (single nucleotide polymorphism) - Freq > 0.01 – Can also be 1 insertion or 1 deletion, “indel” – Alleles with Freq < 0.01 – called rare variants OR SNPs – Mutations: usually, really RARE. Alter
    [Show full text]
  • Anti-Alpha Defensin 1 Antibody (ARG65200)
    Product datasheet [email protected] ARG65200 Package: 100 μg anti-alpha Defensin 1 antibody Store at: -20°C Summary Product Description Goat Polyclonal antibody recognizes alpha Defensin 1 Tested Reactivity Hu Tested Application IHC-P, WB Specificity This antibody is expected to equally recognize DEFA1 precursor and DEFA3 precursor. Host Goat Clonality Polyclonal Isotype IgG Target Name alpha Defensin 1 Antigen Species Human Immunogen C-DESLAPKHPGSRKN Conjugation Un-conjugated Alternate Names DEF1; Neutrophil defensin 1; HNP-1; HNP-2; HP-2; HP1; Defensin, alpha 1; MRS; DEFA2; HP2; HP-1 Application Instructions Application table Application Dilution IHC-P 3 - 6 µg/ml WB 1 - 3 µg/ml Application Note IHC-P: Antigen Retrieval: Steam tissue section in Citrate buffer (pH 6.0). WB: Recommend incubate at RT for 1h. * The dilutions indicate recommended starting dilutions and the optimal dilutions or concentrations should be determined by the scientist. Calculated Mw 10 kDa Properties Form Liquid Purification Purified from goat serum by ammonium sulphate precipitation followed by antigen affinity chromatography using the immunizing peptide. Buffer Tris saline (pH 7.3), 0.02% Sodium azide and 0.5% BSA Preservative 0.02% Sodium azide Stabilizer 0.5% BSA Concentration 0.5 mg/ml Storage instruction For continuous use, store undiluted antibody at 2-8°C for up to a week. For long-term storage, aliquot and store at -20°C or below. Storage in frost free freezers is not recommended. Avoid repeated www.arigobio.com 1/2 freeze/thaw cycles. Suggest spin the vial prior to opening. The antibody solution should be gently mixed before use.
    [Show full text]
  • Novel Methods to Study Genomic Fragility and Structural Variation
    From the Department of Medical Biochemistry and Biophysics Karolinska Institutet, Stockholm, Sweden NOVEL METHODS TO STUDY GENOMIC FRAGILITY AND STRUCTURAL VARIATION Reza Mirzazadeh Stockholm 2019 All previously published papers were reproduced with permission from the publisher. Published by Karolinska Institutet. Printed by Arkitektkopia AB, 2019 © Reza Mirzazadeh, 2019 ISBN 978-91-7831-615-1 Novel methods to study genomic fragility and structural variation THESIS FOR DOCTORAL DEGREE (Ph.D.) Defended at Karolinska Institutet, Biomedicum seminar room 1, Solnavägen 9, Stockholm. December 6th 2019, at 10:00 a.m. By Reza Mirzazadeh Principal Supervisor: Opponent: Assistant Professor Nicola Crosetto Assistant Professor Vicente Pelechano Karolinska Institutet Karolinska Institutet Department of Medical Biochemistry Department of Microbiology, and Biophysics Tumor and Cell Biology Co-supervisor(s): Examination Board: Associate Professor Theodoros Foukakis Professor Ulf Landegren Karolinska Institutet Uppsala University Department of Oncology-Pathology Department of Immunology, Genetics and Pathology Professor Qiang Pan-Hammarström Karolinska Institutet Professor Mattias Mannervik Department of Biosciences and Nutrition Stockholm University Department of Molecular Biosciences, The Wenner-Gren Institute Professor Richard Rosenquist Brandell Karolinska Institutet Department of Molecular Medicine and Surgery Dedicated to family, friends, colleagues, my wonderful wife, Nana, and my amazing daughter Elsa, for their support and love. ABSTRACT DNA double-strand breaks (DSBs) are major DNA lesions that when repaired unfaithfully can give rise to loss of genetic information, chromosomal rearrange- ments such as insertions/deletions (indels) and copy number alterations (CNAs), which in turn lead to genomic instability that is characteristic of almost all cancer types. In this context, it is thought that genomic instability has critical roles in cancer initiation, progression and intra-tumor heterogeneity (ITH).
    [Show full text]