ORIGINAL ARTICLE Implication of LRRC4C and DPP6 in Neurodevelopmental Disorders Gilles Maussion,1 Cristiana Cruceanu,1,2 Jill A. Rosenfeld,3 Scott C. Bell,1 Fabrice Jollant,1,4 Jin Szatkiewicz,5 Ryan L. Collins,6,7 Carrie Hanscom,6 Ilaria Kolobova,1 Nicolas Menjot de Champfleur,8 Ian Blumenthal,6 Colby Chiang,9,10 Vanessa Ota,1 Christina Hultman,11 Colm O’Dushlaine,7 Steve McCarroll,7,12 Martin Alda,13 Sebastien Jacquemont,14 Zehra Ordulu,15,16 Christian R. Marshall,17 Melissa T. Carter,18 Lisa G. Shaffer,3 Pamela Sklar,19 Santhosh Girirajan,20 Cynthia C. Morton,7,21,22 James F. Gusella,6,7,12 Gustavo Turecki,1,2 Dimitri J. Stavropoulos,23 Patrick F. Sullivan,5 Stephen W. Scherer,17,24 Michael E. Talkowski,6,7,25 and Carl Ernst1,2* 1Department of Psychiatry, McGill Group for Suicide Studies, and Douglas Mental Health University Institute, Montreal, Canada 2Department of Human Genetics, McGill University, Montreal, Canada 3Signature Genomic Laboratories, PerkinElmer, Inc., Spokane, Washington 4Nıˆmes Academic Hospital (CHU), Nıˆmes, France 5Department of Genetics, University of North Carolina, Chapel Hill, North Carolina 6Center for Human Genetic Research, Massachusetts General Hospital, Boston, Massachusetts 7Broad Institute of MIT and Harvard, Cambridge, Massachusetts 8Department of Neuroradiology, University Hospital Center of Montpellier, Montpellier, France 9Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Charlottesville, Virginia 10McDonnell Genome Institute, Washington University School of Medicine, St. Louis, Missouri 11Department of Medical Epidemiology and Biostatistics, Karolinska Institute, Stockholm, Sweden 12Department of Genetics, Harvard Medical School, Boston, Massachusetts 13Department of Psychiatry Halifax, Dalhousie University, Halifax, Nova Scotia, Canada 14Department of Pediatrics, Sainte-Justine Hospital, University of Montreal, Montreal, Canada 15Department of Obstetrics, Gynecology and Reproductive Biology, Brigham and Women’s Hospital, Boston, Massachusetts 16Harvard Medical School, Boston, Massachusetts 17The Centre for Applied Genomics and Genetics and Genome Biology, The Hospital for Sick Children, Toronto, Canada 18Regional Genetics Program, The Children’s Hospital of Eastern Ontario, Ottawa, Canada 19Departments of Neuroscience, Psychiatry and Genetics and Genome Sciences, Mount Sinai Hospital, New York, New York 20Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, Pennsylvania Gilles Maussion and Cristiana Cruceanu are equally contributing authors. Current address of Jill A. Rosenfeld is Department of Molecular and Human Genetics, Baylor College of Medicine, Houston 77030, TX. Current address of Vanessa Ota is Department of Morphology and Genetics—Genetics Division Federal University of S~ao Paulo—SP 04023-900, Brazil. Current address of Lisa G. Shaffer is Paw Print Genetics, Genetic Veterinary Sciences, Inc., Spokane 99202, WA. Conflicts of interest: The authors declare no conflict of interest. Grant sponsor: Canada Research Chairs program; Grant sponsor: Scottish Rite Charitable Foundation. ÃCorrespondence to: Carl Ernst, Ph.D., Douglas Mental Health University Institute, 6875 LaSalle Boulevard, Frank B. Common Pavilion, Room 2101.2, Verdun H4H 1R3, Quebec, Canada. E-mail: [email protected] Article first published online in Wiley Online Library (wileyonlinelibrary.com): 00 Month 2016 DOI 10.1002/ajmg.a.38021 Ó 2016 Wiley Periodicals, Inc. 1 2 AMERICAN JOURNAL OF MEDICAL GENETICS PART A 21Departments of Obstetrics, Gynecology, and Reproductive Biology and of Pathology, Brigham and Women’s Hospital, and Harvard Medical School, Boston, Massachusetts 22Manchester Academic Health Science Center, University of Manchester, Manchester, United Kingdom 23Genome Diagnostics, Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, University of Toronto, Toronto, Canada 24Department of Molecular Genetics and McLaughlin Centre, University of Toronto, Toronto, Canada 25Department of Neurology, Harvard Medical School, Boston, Massachusetts Manuscript Received: 13 April 2016; Manuscript Accepted: 29 September 2016 We performed whole-genome sequencing on an individual from a family with variable psychiatric phenotypes that had a sensory How to Cite this Article: processing disorder, apraxia, and autism. The proband harbored Maussion G, Cruceanu C, Rosenfeld JA, a maternally inherited balanced translocation (46,XY,t(11;14) Bell SC, Jollant F, Szatkiewicz J, Collins RL, LRRC4C (p12;p12)mat) that disrupted , a member of the highly Hanscom C, Kolobova I, de Champfleur specialized netrin G family of axon guidance molecules. The NM, Blumenthal I, Chiang C, Ota V, proband also inherited a paternally derived chromosomal Hultman C, O’Dushlaine C, McCarroll S, DPP6, inversion that disrupted a potassium channel interacting Alda M, Jacquemont S, Ordulu Z, protein. Copy Number (CN) analysis in 14,077 cases with neuro- Marshall CR, Carter MT, Shaffer LG, developmental disorders and 8,960 control subjects revealed Sklar P, Girirajan S, Morton CC, LRRC4C that 60% of cases with exonic deletions in had a second Gusella JF, Turecki G, Stavropoulos DJ, clinically recognizable syndrome associated with variable clini- Sullivan PF, Scherer SW, Talkowski ME, cal phenotypes, including 16p11.2, 1q44, and 2q33.1 CN syn- Ernst C. 2016. Implication of LRRC4C and LRRC4C dromes, suggesting deletion variants may be modifiers DPP6 in neurodevelopmental disorders. of neurodevelopmental disorders. In vitro, functional assess- Am J Med Genet Part A 9999A:1–12. ments modeling patient deletions in LRRC4C suggest a negative regulatory role of these exons found in the untranslated region of LRRC4C, which has a single, terminal coding exon. These data suggest that the proband’s autism may be due to the inheritance a contributor to NDDs and deletions in LRRC4C may be a modifier of disruptions in both DPP6 and LRRC4C, and may highlight the of genetic lesions associated with variable NDD phenotypes. importance of the netrin G family and potassium channel interacting molecules in neurodevelopmental disorders. METHODS Ó 2016 Wiley Periodicals, Inc. Family Recruitment and Assessment Key words: autism; sensory processing; LRRC4C; Netrin G; We obtained blood from all carriers of a BCR identified by DPP6 karyotyping in a multigenerational family (46,XY,t(11;14)(p12; p12)mat), from which the proband, the proband’s mother, and grandmother were available for clinical and genetic testing. Participants or their guardians gave informed consent, using INTRODUCTION consent forms approved by the Douglas Institute Ethics Board. While much progress has been made on the genetics of neuro- The proband was assessed with both the ADOS and ADI as part developmental disorders (NDDs), over 50% of cases assessed clini- of a full clinical work-up done at the Montreal Children’s cally on any genetic platform are considered idiopathic [Krumm Hospital. The mother of the proband confirmed that he met et al., 2014; Sanders et al., 2015]. While more NDD cases would likely criteria for autism on both measures. The mother and the be associated with genetic variation if all cases were assessed using the grandmother of the proband were assessed with the SCID-I full extent of state-of-the-art technology, different strategies need to for Axis-I mental disorders and SCID-II for personality disorders be employed to further unravel the genetics of NDDs. We have according to DSM-IV criteria. Current level of depression was previously shown that balanced chromosomal rearrangement (BCR) assessed by the Beck Depression Inventory and the Hamilton sequencing is a powerful strategy to discover new genes for NDDs Depression Rating Scale (HDRS-24). Cognitive functioning for [Talkowski et al., 2011, 2012c], and here, we report on a child who these two individuals was investigated with the Stroop Color Test inherited two BCRs, one from each parent, each of which disrupted and the Hayling Sentence Completion test for cognitive inhibi- only a single gene: leucine rich repeat containing 4C (LRRC4C [MIM tion, the Trail Making Test (TMT) A and B for flexibility/shifting, 608817]) and dipeptidyl-peptidase 6 (DPP6 [MIM 126141]). Analy- a categorical and semantic verbal fluency test, the National sis of rare copy gains and losses in these genes in thousands of Adult Reading Test (NART) for a raw estimate of verbal IQ, NDD cases suggest that that exon-disrupting CNVs in DPP6 may be the working memory subscale of the WAIS-III, and the Iowa MAUSSION ET AL. 3 Gambling Task (IGT) for decision-making. Individual cognitive and III–2, were read by an experienced radiologist (NM) blinded to performance was then compared to performance from 81 healthy the genetic status of the subjects but aware of their age. A systematic female controls from the GREFEX control group and 72 healthy assessment was run, which included: movement artifacts, atrophy female controls from our own database. (pons, vermian, cortical, corpus callosum), white matter hyper- intensities (infratentorial, cerebellar, pons, dentate nuclei, cerebellar Next Generation Sequencing of BCRs peduncles, mesencephalon, periventricular, deep, juxtacortical, basal ganglia, semi ovale centrum, corticospinal tract, corpus Translocation mapping experiments were performed using cus- callosum), Scheltens and Koedam scores, enlargement (ventricular, tomized large-insert, or “jumping library,” whole genome olfactive sulcus), infarct (territorial, lacunar,