Abstract

Autism is a multifactorial neurodevelopmental disorder affecting more males than females; consequently, under a multifactorial genetic hypothesis, females are affected only when they cross a higher biological threshold. We hypothesize that deleterious variants at conserved residues are enriched in severely affected patients arising from female-enriched multiplex families with severe disease, enhancing the detection of key autism genes in modest numbers of cases. Here we show the use of this strategy by identifying missense and dosage sequence variants in the gene encoding the adhesive junction-associated δ-catenin protein (CTNND2) in female-enriched multiplex families and demonstrating their loss-of-function effect by functional analyses in zebrafish embryos and cultured hippocampal neurons from wild-type and Ctnnd2 null mouse embryos. Finally, through gene expression and network analyses, we highlight a critical role for CTNND2 in neuronal development and an intimate connection to chromatin biology. Our data contribute to the understanding of the genetic architecture of autism and suggest that genetic analyses of phenotypic extremes, such as female-enriched multiplex families, are of innate value in multifactorial disorders.

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Acknowledgements

We acknowledge the participation of all of the families in the AGRE, NIMH and SSC studies that have been a model of public participatory research. The AGRE is a program of Autism Speaks and is supported, in part, by grant 1U24MH081810 from the National Institute of Mental Health. The SSC used here was developed by the following principal investigators: A. Beaudet, R. Bernier, J. Constantino, E. Cook, E. Fombonne, D. Geschwind, D. Grice, A. Klin, D. Ledbetter, C. Lord, C. Martin, D. Martin, R. Maxim, J. Miles, O. Ousley, B. Peterson, J. Piggot, C. Saulnier, M. State, W. Stone, J. Sutcliffe, C. Walsh, E. Wijsman. We thank the Allen Brain Atlas for use of their publicly available developing human brain expression data. Finally, we thank V. Kustanovich (AGRE) for helping with access to Autism Diagnostic Observation Schedule severity score data, D. Arking for sharing DNA from the SSC for Taqman genotyping, S. Maragh for zebrafish complementary DNA (cDNA) libraries and eef1a1l1 primers, A. Kapoor for discussions, Q. Jiang for the translation of ref. 43, and J. A. Rosenfeld, L. G. Shaffer, Y. Shen and B.-L. Wu for sharing CNV data sets. Sequencing services were provided by the Johns Hopkins University Next Generation Sequencing Center, Sidney Kimmel Comprehensive Cancer Center, Illumina Sequencing Services and the Johns Hopkins University Genetic Resources Core Facility. E.C.O. is a National Alliance for Research on Schizophrenia and Depression young investigator. N.K. is a Distinguished George W. Brumley Professor. This work was funded by grants from the Simons Foundation to A.C. and to N.K., NIMH grant MH095867 to M.E.T., NIMH grants 5R25MH071584-07 and MH19961-14 to D.M.D.L. (Malison), National Institutes of Health grant RO1MH074090 to C.L.M., NIMH grant R01MH081754 to A.C. and an Autism Speaks Dennis Weatherstone pre-doctoral fellowship (number 7863) to T.T.

Author information

Affiliations

  1. Center for Complex Disease Genomics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA

    • Tychele N. Turner
    • , Maria X. Sosa
    • , Dallas R. Auer
    • , Vasyl Pihur
    •  & Aravinda Chakravarti
  2. Predoctoral Training Program in Human Genetics and Molecular Biology, McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA

    • Tychele N. Turner
  3. National Institute of Mental Health (NIMH) Autism Centers of Excellence (ACE) Genetics Consortium at the University of California, Los Angeles, Los Angeles, California 90095, USA

    • Tychele N. Turner
    • , Maria X. Sosa
    • , Dallas R. Auer
    • , Stephan J. Sanders
    • , Daniel Moreno-De-Luca
    • , Vasyl Pihur
    • , Christa Lese Martin
    • , Matthew W. State
    •  & Aravinda Chakravarti
  4. Solomon H. Snyder Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA

    • Kamal Sharma
    •  & Richard Huganir
  5. Center for Human Disease Modeling, Duke University, Durham, North Carolina 27710, USA

    • Edwin C. Oh
    • , Yangfan P. Liu
    •  & Nicholas Katsanis
  6. Center for Human Genetic Research, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114, USA

    • Ryan L. Collins
    • , Harrison Brand
    •  & Michael E. Talkowski
  7. Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114 USA

    • Harrison Brand
    •  & Michael E. Talkowski
  8. Department of Psychiatry, University of California, San Francisco, San Francisco, California 94158, USA

    • Stephan J. Sanders
    •  & Matthew W. State
  9. Department of Psychiatry, Yale University, New Haven, Connecticut 06511, USA

    • Daniel Moreno-De-Luca
  10. Leidos Biomedical Research, Inc., Frederick, Maryland 21702, USA

    • Teri Plona
    • , Kristen Pike
    •  & Daniel R. Soppet
  11. National Human Genome Research Institute, Bethesda, Maryland 20892, USA

    • Michael W. Smith
  12. Baylor College of Medicine, Houston, Texas 77030, USA

    • Sau Wai Cheung
  13. Autism & Developmental Medicine Institute, Geisinger Health System, Lewisburg, Pennsylvania 17837, USA

    • Christa Lese Martin
  14. University of Illinois at Chicago, Chicago, Illinois 60608, USA

    • Edwin Cook

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Contributions

Designed the study and wrote the manuscript (T.T., A.C.); edited the manuscript (all authors); examined phenotype data for the female autism patients (T.T., E.C.); MECP2/CTNND2 sequencing and TaqMan genotyping (T.T., M.X.S., T.P., K.P., D.S., M.W.S.); autism exome sequencing (T.T.); Simons exome sequencing analysis (S.S., M.S.); CNV analysis (S.W.C., C.L.M., D.M.D., S.S., R.C.C., H.B., M.E.T, M.S., T.T.); CTNND2 molecular biology (T.T., M.X.S.); zebrafish gastrulation and protein-protein interaction studies (Y.P.L., E.O., N.K.); primary hippocampal neuron experiments and expression analysis (K.S., T.T., D.A.); bioinformatics analyses (T.T.,V.P.).

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Aravinda Chakravarti.

All sequence data have been deposited in the National Database for Autism Research in NDAR Study 367 and are available at http://dx.doi.org/10.15154/1171641.

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https://doi.org/10.1038/nature14186

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