14
views
0
recommends
+1 Recommend
0 collections
    0
    shares
      • Record: found
      • Abstract: found
      • Article: found
      Is Open Access

      Whole exome sequencing coupled with unbiased functional analysis reveals new Hirschsprung disease genes

      research-article
      1 , 2 , 3 , 1 , 4 , 4 , 5 , 6 , 7 , 6 , 8 , 4 , 9 , 4 , 10 , 2 , 5 , 6 , 3 , 11 , 12 , 13 , 14 , 1 , 5 , 6 , 11 , 1 , 13 , 14 , 5 , 6 , 2 , 12 , 1 , 4 , 15 , 1 , 9 , 4 , 4 , 9 , 16 , 13 , 14 , 5 , 6 , 11 , 7 , 13 , 14 , 1 , 1 , , 4 , 10 ,
      Genome Biology
      BioMed Central
      De novo mutations, Hirschsprung disease, Neural crest, ENS

      Read this article at

      Bookmark
          There is no author summary for this article yet. Authors can add summaries to their articles on ScienceOpen to make them more accessible to a non-specialist audience.

          Abstract

          Background

          Hirschsprung disease (HSCR), which is congenital obstruction of the bowel, results from a failure of enteric nervous system (ENS) progenitors to migrate, proliferate, differentiate, or survive within the distal intestine. Previous studies that have searched for genes underlying HSCR have focused on ENS-related pathways and genes not fitting the current knowledge have thus often been ignored. We identify and validate novel HSCR genes using whole exome sequencing (WES), burden tests, in silico prediction, unbiased in vivo analyses of the mutated genes in zebrafish, and expression analyses in zebrafish, mouse, and human.

          Results

          We performed de novo mutation (DNM) screening on 24 HSCR trios. We identify 28 DNMs in 21 different genes. Eight of the DNMs we identified occur in RET, the main HSCR gene, and the remaining 20 DNMs reside in genes not reported in the ENS. Knockdown of all 12 genes with missense or loss-of-function DNMs showed that the orthologs of four genes (DENND3, NCLN, NUP98, and TBATA) are indispensable for ENS development in zebrafish, and these results were confirmed by CRISPR knockout. These genes are also expressed in human and mouse gut and/or ENS progenitors. Importantly, the encoded proteins are linked to neuronal processes shared by the central nervous system and the ENS.

          Conclusions

          Our data open new fields of investigation into HSCR pathology and provide novel insights into the development of the ENS. Moreover, the study demonstrates that functional analyses of genes carrying DNMs are warranted to delineate the full genetic architecture of rare complex diseases.

          Electronic supplementary material

          The online version of this article (doi:10.1186/s13059-017-1174-6) contains supplementary material, which is available to authorized users.

          Related collections

          Most cited references39

          • Record: found
          • Abstract: found
          • Article: not found

          De novo gene disruptions in children on the autistic spectrum.

          Exome sequencing of 343 families, each with a single child on the autism spectrum and at least one unaffected sibling, reveal de novo small indels and point substitutions, which come mostly from the paternal line in an age-dependent manner. We do not see significantly greater numbers of de novo missense mutations in affected versus unaffected children, but gene-disrupting mutations (nonsense, splice site, and frame shifts) are twice as frequent, 59 to 28. Based on this differential and the number of recurrent and total targets of gene disruption found in our and similar studies, we estimate between 350 and 400 autism susceptibility genes. Many of the disrupted genes in these studies are associated with the fragile X protein, FMRP, reinforcing links between autism and synaptic plasticity. We find FMRP-associated genes are under greater purifying selection than the remainder of genes and suggest they are especially dosage-sensitive targets of cognitive disorders. Copyright © 2012 Elsevier Inc. All rights reserved.
            Bookmark
            • Record: found
            • Abstract: found
            • Article: not found

            Efficient multiplex biallelic zebrafish genome editing using a CRISPR nuclease system.

            A simple and robust method for targeted mutagenesis in zebrafish has long been sought. Previous methods generate monoallelic mutations in the germ line of F0 animals, usually delaying homozygosity for the mutation to the F2 generation. Generation of robust biallelic mutations in the F0 would allow for phenotypic analysis directly in injected animals. Recently the type II prokaryotic clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated proteins (Cas) system has been adapted to serve as a targeted genome mutagenesis tool. Here we report an improved CRISPR/Cas system in zebrafish with custom guide RNAs and a zebrafish codon-optimized Cas9 protein that efficiently targeted a reporter transgene Tg(-5.1mnx1:egfp) and four endogenous loci (tyr, golden, mitfa, and ddx19). Mutagenesis rates reached 75-99%, indicating that most cells contained biallelic mutations. Recessive null-like phenotypes were observed in four of the five targeting cases, supporting high rates of biallelic gene disruption. We also observed efficient germ-line transmission of the Cas9-induced mutations. Finally, five genomic loci can be targeted simultaneously, resulting in multiple loss-of-function phenotypes in the same injected fish. This CRISPR/Cas9 system represents a highly effective and scalable gene knockout method in zebrafish and has the potential for applications in other model organisms.
              Bookmark
              • Record: found
              • Abstract: found
              • Article: not found

              De novo mutations in human genetic disease.

              New mutations have long been known to cause genetic disease, but their true contribution to the disease burden can only now be determined using family-based whole-genome or whole-exome sequencing approaches. In this Review we discuss recent findings suggesting that de novo mutations play a prominent part in rare and common forms of neurodevelopmental diseases, including intellectual disability, autism and schizophrenia. De novo mutations provide a mechanism by which early-onset reproductively lethal diseases remain frequent in the population. These mutations, although individually rare, may capture a significant part of the heritability for complex genetic diseases that is not detectable by genome-wide association studies.
                Bookmark

                Author and article information

                Contributors
                +852-28315073 , mmgarcia@hku.hk
                +31-10-7037643 , r.hofstra@erasmusmc.nl
                Journal
                Genome Biol
                Genome Biol
                Genome Biology
                BioMed Central (London )
                1474-7596
                1474-760X
                8 March 2017
                8 March 2017
                2017
                : 18
                : 48
                Affiliations
                [1 ]ISNI 0000000121742757, GRID grid.194645.b, Department of Surgery, Li Ka Shing Faculty of Medicine, , The University of Hong Kong, ; Hong Kong, SAR China
                [2 ]ISNI 0000000121742757, GRID grid.194645.b, Centre for Genomic Sciences, Li Ka Shing Faculty of Medicine, , The University of Hong Kong, ; Hong Kong, SAR China
                [3 ]Department of Neuroscience, section Medical Physiology, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands
                [4 ]ISNI 000000040459992X, GRID grid.5645.2, Department of Clinical Genetics, , Erasmus University Medical Center, ; PO Box 2040, 3000CA Rotterdam, The Netherlands
                [5 ]ISNI 0000 0000 9542 1158, GRID grid.411109.c, Department of Genetics, Reproduction and Fetal Medicine, Institute of Biomedicine of Seville (IBIS), , University Hospital Virgen del Rocío/CSIC/University of Seville, ; Seville, Spain
                [6 ]Centre for Biomedical Network Research on Rare Diseases (CIBERER), Seville, Spain
                [7 ]ISNI 0000 0001 2171 9311, GRID grid.21107.35, McKusick-Nathans Institute of Genetic Medicine, , Johns Hopkins University School of Medicine, ; Baltimore, USA
                [8 ]ISNI 0000000121885934, GRID grid.5335.0, Department of Medicine, School of Clinical Medicine, , University of Cambridge, Addenbrooke’s Hospital, ; Cambridge, UK
                [9 ]ISNI 000000040459992X, GRID grid.5645.2, , Erasmus Center for Biomics, Erasmus Medical Center, ; Rotterdam, The Netherlands
                [10 ]ISNI 0000000121901201, GRID grid.83440.3b, , Stem Cells and Regenerative Medicine, Birth Defects Research Centre, UCL Institute of Child Health, ; London, UK
                [11 ]UOC Genetica Medica, Istituto Gaslini, Genoa, Italy
                [12 ]ISNI 0000 0001 0941 6502, GRID grid.189967.8, Department of Biology, , Emory University, ; Atlanta, USA
                [13 ]ISNI 0000000121866389, GRID grid.7429.8, Laboratory of embryology and genetics of human malformations, , INSERM UMR 1163, Institut Imagine, ; Paris, France
                [14 ]ISNI 0000 0004 0593 9113, GRID grid.412134.1, Department of Genetics, , Paris Descartes-Sorbonne Paris Cité University, Hôpital Necker-Enfants Malades (APHP), ; Paris, France
                [15 ]ISNI 0000 0004 1796 1481, GRID grid.11553.33, Department of Biochemistry and Molecular Biology, Faculty of Medicine, , Universitas Padjadjaran, ; Bandung, Indonesia
                [16 ]Department of Genetics, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands
                Article
                1174
                10.1186/s13059-017-1174-6
                5343413
                28274275
                5b79f9c6-205c-4ac8-9ffa-e7c5d2910b98
                © The Author(s). 2017

                Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License ( http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver ( http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

                History
                : 7 October 2016
                : 17 February 2017
                Funding
                Funded by: FundRef http://dx.doi.org/10.13039/501100001826, ZonMW;
                Award ID: 40-00812-98-10042
                Funded by: FundRef http://dx.doi.org/10.13039/501100005847, Health and Medical Research Fund;
                Award ID: 02131866
                Award Recipient :
                Funded by: FundRef http://dx.doi.org/10.13039/501100004587, Instituto de Salud Carlos III;
                Award ID: PI13/01560; EXP00052887; CTS-7447
                Award Recipient :
                Funded by: the Maag Lever Darm stichting
                Award ID: WO09-62
                Funded by: FundRef http://dx.doi.org/10.13039/100000009, Foundation for the National Institutes of Health;
                Award ID: R37 HD28088
                Award Recipient :
                Funded by: General Research Fund
                Award ID: HKU 777612M)
                Funded by: FundRef http://dx.doi.org/10.13039/100000065, National Institute of Neurological Disorders and Stroke;
                Award ID: 5R21NS082546
                Funded by: Italian Ministry of Health
                Award ID: Cinque per mille
                Award Recipient :
                Categories
                Research
                Custom metadata
                © The Author(s) 2017

                Genetics
                de novo mutations,hirschsprung disease,neural crest,ens
                Genetics
                de novo mutations, hirschsprung disease, neural crest, ens

                Comments

                Comment on this article