Exome Sequencing in Multiplex Autism Families Suggests A Major Role For Heterozygous Truncating Mutations
Exome Sequencing in Multiplex Autism Families Suggests A Major Role For Heterozygous Truncating Mutations
Exome Sequencing in Multiplex Autism Families Suggests A Major Role For Heterozygous Truncating Mutations
ORIGINAL ARTICLE
Exome sequencing in multiplex autism families suggests a
major role for heterozygous truncating mutations
C Toma1,2, B Torrico1,2, A Hervás3, R Valdés-Mas4, A Tristán-Noguero1, V Padillo5, M Maristany5, M Salgado3, C Arenas6, XS Puente4,
M Bayés7 and B Cormand1,2,8
Autism is a severe neurodevelopmental disorder, the aetiology of which remains mainly unknown. Family and twin studies provide
strong evidence that genetic factors have a major role in the aetiology of this disease. Recently, whole exome sequencing (WES)
efforts have focused mainly on rare de novo variants in singleton families. Although these studies have provided pioneering
insights, de novo variants probably explain only a small proportion of the autism risk variance. In this study, we performed exome
sequencing of 10 autism multiplex families with the aim of investigating the role of rare variants that are coinherited in the affected
sibs. The pool of variants selected in our study is enriched with genes involved in neuronal functions or previously reported in
psychiatric disorders, as shown by Gene Ontology analysis and by browsing the Neurocarta database. Our data suggest that rare
truncating heterozygous variants have a predominant role in the aetiology of autism. Using a multiple linear regression model, we
found that the burden of truncating mutations correlates with a lower non-verbal intelligence quotient (NVIQ). Also, the number of
truncating mutations that were transmitted to the affected sibs was significantly higher (twofold) than those not transmitted.
Protein–protein interaction analysis performed with our list of mutated genes revealed that the postsynaptic YWHAZ is the most
interconnected node of the network. Among the genes found disrupted in our study, there is evidence suggesting that YWHAZ and
also the X-linked DRP2 may be considered as novel autism candidate genes.
Molecular Psychiatry (2014) 19, 784–790; doi:10.1038/mp.2013.106; published online 3 September 2013
Keywords: autism spectrum disorder; exome sequencing; multiplex families; novel candidate genes; rare genetic variants;
truncating mutations
Family ID (sib pair ID) Sex (years) Phenotype NVIQ Language delay
& 2014 Macmillan Publishers Limited Molecular Psychiatry (2014), 784 – 790
Exome sequencing in multiplex autism families
C Toma et al
786
Table 2. Gene-disrupting rare variants shared by the affected sibs in each multiplex family
were predicted to be pathogenic. The pool of validated variants is Table 3). Several genes in this category such as CYFIP1, SCN1A,
listed in Supplementary Table 2. Most (83%) are missense DNM2, FLNB, GABRA4, P2RX4, PJA2 and SV2B have been described
changes, whereas variants predicted to cause protein truncation in autism or are involved in neuronal functions. Several GO
(nonsense mutations and frameshift indels) accounted for 16% of biological processes were over-represented in our study and some
the total. Variants altering canonical splice sites or start codons of them may be relevant for autism, such as ‘Neuronal tube
were less represented, at 0.5% each (Supplementary Figure 2). All development’ (P ¼ 0.044) and ‘Regulation of action potential’
these variants were heterozygous, none of them was homozygous (P ¼ 0.044) (Supplementary Table 4). In addition, we used IPA to
or compound heterozygous, and no gene was found mutated in analyze the ‘Top Bio function’ category of ‘Diseases and disorders’
more than one family. Truncating mutations represent remarkable and found that ‘Developmental disorders’ and ‘Neurological
events in the exome and may pinpoint potential candidate genes diseases’, among others, were the most significant groups related
for a disease. Interestingly, in our study this kind of mutation to the genes in our study (Supplementary Table 5). The Neurocarta
represented a substantial proportion of the whole pool of variants, database was used to build a map of psychiatric phenotypes
with 20 indels and 16 nonsense mutations (Table 2). previously linked to these genes (Figure 1). The majority of genes
Chromosomal rearrangements and previously described fully were associated with autism, but also with epilepsy, dyslexia,
penetrant deletions were ruled out in the affected probands, by intellectual disability, attention-deficit hyperactivity disorder and
karyotyping and through a CNV study. This analysis, followed by schizophrenia.
experimental validation, revealed six inherited CNVs present in We also investigated the possible correlation between the
five sib-pairs identifying the following genes: COL4A3-MFF, FHIT, severity of the mutations and intellectual disability, by using the
MRPL36-NDUFS6, CTNND2, GRM1 and ASAH1 (Table 3). Structural NVIQ. NVIQ was assessed in our sample of 21 affected siblings,
variants spanning the genes FHIT and CTNND2 had previously with scores ranging from 35 to 139 and a mean of 91
been described in autism.14,26 (Supplementary Figure 3). A multiple linear regression model
To establish whether the pool of variants selected in our exome was applied with NVIQ as a response variable and truncating, non-
study was enriched in potential ASD susceptibility genes, we synonymous (damaging and benign changes according to SIFT
conducted a Gene Ontology study. Cellular component analysis and PolyPhen) and synonymous variants as regressor variables.
showed ‘Cell junction’ to be a significantly enriched category after The results showed that only truncating variants contribute
applying multiple testing corrections (P ¼ 0.043) (Supplementary significantly to NVIQ (P ¼ 0.007). Furthermore, truncating variants
Molecular Psychiatry (2014), 784 – 790 & 2014 Macmillan Publishers Limited
Exome sequencing in multiplex autism families
C Toma et al
787
Table 3. Validated CNVs shared by the affected sibs in each multiplex family
SJD_34.3–34.4 2 228 149 569 228 193 389 q36.3 Gain 43.82 COL4A3, MFF
MT_109.3–109.4 3 60 478 959 60 572 752 p14.2 Loss 93.793 FHIT
MT_76.3–76.4 5 1 742 845 1 849 924 p15.33 Gain 107.079 MRPL36, NDUFS6
SJD_34.3–34.4 5 11 619 568 12 183 983 p15.2 Gain 564.415 CTNND2
SJD_49.3–49.4 6 146 309 359 146 373 644 q24.3 Loss 64.285 GRM1
SJD_50.3–50.4 8 17 908 916 17 946 695 p22 Loss 37.779 ASAH1
Abbreviation: CNV, copy number variant.
a
Positions are indicated according to the GRCh37/hg19 assembly of the UCSC Genome Browser (www.genome.ucsc.edu).
DISCUSSION
were correlated with lower NVIQ scores (correlation coefficient Several exome sequencing reports of autism trios have been
r ¼ –0.517, P ¼ 0.016; Figure 2) and explained 26% of NVIQ published in the past few years. These studies have enabled the
variance in our ASD sample (r2 ¼ 0.267). A simulation study identification of novel candidate genes for ASD by focusing on de
including 50 new simulated analyses of 100 individuals each novo variants. Despite these encouraging results, de novo variants
obtained the same results as in the original sample (for more represent probably o5% of autism risk variance,8 and hence
details see Supplementary Information, Supplementary Table 7 inherited rare variants may account for a considerable proportion
and Supplementary Figure 8). of the missing heritability in autism. Here we present findings from
We subsequently investigated whether truncating mutations the exome sequencing of 10 multiplex families, in which only the
might have a major role in autism aetiology by comparing the inherited rare variants shared by the affected siblings in a family
number of those that were cotransmitted (Table 1) with those that and predicted to be pathogenic were considered. The resulting list
were not transmitted (Supplementary Table 6). We found a of about 220 identified genetic variants was assessed for GO
significant difference, considering the total number of rare enrichment analysis and networks of gene interactions (IPA).
variants, between disrupting mutations that were transmitted to These approaches identified interesting categories related to
affected probands (16 nonsense variants and 20 frameshift indels) developmental disorders or involving neuronal functions, and
and those not transmitted (9 nonsense variants and 9 frameshift revealed plausible interactions with previously reported ASD
indels) (Fisher’s exact test, P ¼ 0.015; Supplementary Figure 4). genes. In our study, we identified a substantial number of genes
Subsequently, we investigated the potential contribution to already associated with autism or with other psychiatric
ASD of rare non-synonymous variants predicted to be pathogenic, conditions, suggesting that there is a common genetic
by comparing the group of transmitted (TR) single-nucleotide background for psychiatric disorders.27,28 The most interesting
variants with those that were not transmitted (NT). A multiple finding emerging from our study suggests a major role for
logistic regression analysis (fitted logit model) was performed to truncating mutations in autism. We found that those probands
investigate whether the scores given by SIFT and PolyPhen with a higher number of heterozygous disrupting mutations are
were able to discriminate the single-nucleotide variants in relation those with lower NVIQ scores. In addition, we found more
to their TR or NT status, but the results were not significant truncating mutations that were transmitted and shared between
(SIFT P ¼ 0.122; PolyPhen P ¼ 0.811). To further test the possible the affected sibs (36 mutations) than those that were not
relation of the non-synonymous variants with the disease, transmitted (18 mutations). Interestingly, Iossifov et al.9 analysed
we plotted all missense changes against the PolyPhen scores exome data and described a twofold higher rate of disrupting de
(X axis) and SIFT scores (Y axis) (Supplementary Figure 5) to novo mutations in affected probands compared with unaffected
determine whether there was any difference in the proportion of siblings. Also, a very recent WES study on inherited homozygous
variants predicted to be pathogenic (PolyPhen scores40.5 and or compound heterozygous loss-of-function mutations found a
SIFT scoreo0.05) between the TR and NT groups. No significant twofold enrichment in autism compared to a control group.18
differences were detected (two-sided exact binomial test, Such data suggest a genetic model for autism based on the
P ¼ 0.47). cumulative contribution of truncating alleles and other rare
& 2014 Macmillan Publishers Limited Molecular Psychiatry (2014), 784 – 790
Exome sequencing in multiplex autism families
C Toma et al
788
Figure 2. Distribution of the average number of rare variants per proband for each interval of non-verbal intelligence quotient (NVIQ) in four
variant categories: (1) truncating mutations, which include nonsense and frameshift mutations; (2) non-syn damaging, non-synonymous
mutations predicted to be damaging by SIFT or PolyPhen; (3) non-syn benign, non-synonymous mutations predicted to be benign by SIFT and
PolyPhen; and (4) Syn, rare synonymous variants. The pool of rare variants considered are those inherited by two or three affected sibs in a
family. Multiple linear regression analysis of these data showed that the number of truncating variants contributes significantly to NVIQ
(P ¼ 0.007). In contrast, the contribution of the other mutation types was not significant (P40.385). The correlation coefficients between NVIQ
and each mutation type with their significances are also presented.
Figure 3. Protein–protein interaction analysis (Ingenuity Pathway Analysis, IPA) including all genes found to be mutated in our study. Only
direct interactions among proteins were considered. Proteins in grey or colour represent genes identified in our study, with red indicating a
previous association with autism, green with epilepsy and blue with schizophrenia. Proteins depicted in white are those not present in our
study. Upregulatory effects are represented by outward pointing arrows, downregulatory effects are represented by outward ticks, and
circular arrows indicate homotypic interactions.
Molecular Psychiatry (2014), 784 – 790 & 2014 Macmillan Publishers Limited
Exome sequencing in multiplex autism families
C Toma et al
789
pathogenic variants, coupled with rare structural variants, in which Interestingly, a recent WES study reported the same mutation
the impact of common variants may be less important than (E432*) that we found in DRP2 in one autistic family, which was
previously thought.29 Most genes with truncating mutations absent from a large cohort of controls.18
found in our study have an unknown function and none have This work represents one of the first comprehensive studies of
previously been described in autism. Among these, YWHAZ and multiplex families with ASD by exome sequencing. Similar to other
DRP2 may be considered as strong novel ASD candidate genes. studies using WES technologies, known limitations should be
The NHLBI Exome Sequencing Project (ESP) database (http:// considered. First, the coverage of the exome fraction was not
evs.gs.washington.edu/EVS/) was used to verify the frequency of complete (83.5% of target sequence covered on average); thus, it
disrupting mutations in these genes in about 6500 individuals. No is possible that we missed disease-causing mutations. Second,
truncating mutations were listed in either YWHAZ or DRP2. we used stringent criteria to filter false-positives, and as a
The YWHAZ gene, encoding a postsynaptic protein, is the most consequence we may have missed true aetiological variants that
intriguing candidate in our study. The protein physically interacts were not considered as being pathogenic. Third, we considered
with numerous ASD gene products such as TSC1, TSC2, DISC1, only variants shared by affected probands, although aetiological
UBE3A and CYFIP1 (Supplementary Figure 6). YWHAZ belongs to variants transmitted to only one child are also likely to contribute
the highly conserved 14-3-3 protein family that comprises seven to the disease. Fourth, repetitive elements or variants located on
isoforms (b, g, e, Z, z, s, y) involved in signal transduction by non-coding sequences were not explored in our study. Finally, the
binding specific pSer/pThr motifs. These proteins are involved in a sample under study comprises only 10 families.
wide range of processes including cell cycle, transcription, In conclusion, our data suggest that inherited disrupting
neuronal development, migration and neurite outgrowth. mutations in multiplex families may have a major role in the
Although ubiquitously expressed, expression levels are highest aetiology of ASD. We highlight novel potential ASD candidate
in the brain.30,31 The members of this family have been associated genes such as YWHAZ and DRP2. Further WES studies of inherited
with several neurodevelopmental disorders, syndromes or rare variants in larger ASD samples are warranted to corroborate
psychiatric diseases. Deletions of the contiguous YWHAE and these results and to gain more insight into the missing heritability
PAFAH1B1 genes are responsible for two distinct Mendelian of ASD.
disorders depending on the size of the deletion: isolated
lissencephaly and Miller–Dieker syndrome.32 Duplications
encompassing only the YWHAE gene are associated with a CONFLICT OF INTEREST
distinct phenotype involving autism and other behavioural The authors declare no conflict of interest.
symptoms.33,34 This gene has also been strongly associated with
schizophrenia and found to be downregulated together
with other 14-3-3 isoforms in the prefrontal cortex of ACKNOWLEDGEMENTS
schizophrenic patients.35,36 Other 14-3-3 members have also We are grateful to all families for their participation in our study. We thank Patricia
been associated with psychiatric phenotypes: heterozygous micro- Romarı́s (Hospital Universitari Mútua de Terrassa) for contributing to clinical
deletions encompassing YWHAG and HIP1 were associated with delineation of patients and Lara Nonell and Eulàlia Puigdecanet (Servei d’Anàlisi de
epilepsy, learning difficulties and intellectual disability,37 whereas Micorarrays, IMIM-Hospital del Mar, Parc de Recerca Biomèdica de Barcelona) for their
YWHAH was associated with bipolar disorder.38,39 Recently, Cheah contribution to the CNV studies. Exome sequencing services were provided by the
et al.40 reported that YWHAZ knockout mice show neurobeha- National Centre for Genomic Analysis (CNAG). CT was supported by the European
vioural and cognitive deficiencies, and aberrant development of Union (Marie Curie, PIEF-GA-2009-254930) and BT by AGAUR (FI grant). Financial
support was received from ‘Fundació La Marató de TV3’ (092010), ‘Fundación Alicia
the hippocampus with migratory defects of pyramidal cells and
Koplowitz’, AGAUR (2009SGR00971) and ‘Ministerio de Economı́a y Competitividad,
granular neurons. Furthermore, neuroproteomic studies showed Spain’ (SAF2012-33484, SAF2010-21165).
decreased expression of YWHAZ in the brain of schizophrenic
patients.41 YWHAZ forms a molecular complex with
DISC1 (Disrupted In Schizophrenia 1), Ndel1 and LIS1 to control
REFERENCES
the development of the hippocampus by coordinating
1 Lord C, Jones RM. Annual research review: re-thinking the classification of autism
neuronal migration, axonal pathfinding and synapse formation.40
spectrum disorders. J Child Psychol Psychiatry 2012; 53: 490–509.
In our study, the 1-bp insertion found in YWHAZ causes a 2 Elsabbagh M, Divan G, Koh YJ, Kim YS, Kauchali S, Marcin C et al. Global
frame shift leading to a premature stop codon after 18 amino prevalence of autism and other pervasive developmental disorders. Autism Res
acids (Supplementary Figure 7). If nonsense-mediated mRNA 2012; 5: 160–179.
decay does not prevent the degradation of this transcript, it is 3 Ozonoff S, Young GS, Carter A, Messinger D, Yirmiya N, Zwaigenbaum L et al.
possible that this abnormal protein may act in a dominant- Recurrence risk for autism spectrum disorders: a Baby Siblings Research
negative manner altering the normal protein interactions Consortium study. Pediatrics 2011; 128: e488–e495.
or preventing homo- and heterodimerization of the protein with 4 Ronald A, Hoekstra RA. Autism spectrum disorders and autistic traits: a decade of
14-3-3 members. new twin studies. Am J Med Genet B 2011; 156B: 255–274.
5 Sanders SJ, Ercan-Sencicek AG, Hus V, Luo R, Murtha MT, Moreno-De-Luca D et al.
The X-linked dystrophin-related protein 2 gene (DRP2),
Multiple recurrent de novo CNVs, including duplications of the 7q11.23 Williams
expressed mainly in the brain and spinal cord, is also a good syndrome region, are strongly associated with autism. Neuron 2011; 70: 863–885.
ASD candidate. DRP2 forms a complex with periaxin and 6 O’Roak BJ, Vives L, Fu W, Egertson JD, Stanaway IB, Phelps IG et al. Multiplex
dystroglycan, regulating the myelination of Schwann cells. Sher- targeted sequencing identifies recurrently mutated genes in autism spectrum
man et al.42 showed that loss of DRP2 affects the organization of disorders. Science 2012; 338: 1619–1622.
the Schwann cell cytoplasm in the Cajal bands, although other 7 O’Roak BJ, Vives L, Girirajan S, Karakoc E, Krumm N, Coe BP et al. Sporadic autism
members of the dystrophin family may partially supply the exomes reveal a highly interconnected protein network of de novo mutations.
absence of DRP2. No human disease has been associated with the Nature 2012; 485: 246–250.
DRP2 gene, although deletions spanning DRP2 were described in 8 Neale BM, Kou Y, Liu L, Ma’ayan A, Samocha KE, Sabo A et al. Patterns and rates
of exonic de novo mutations in autism spectrum disorders. Nature 2012; 485:
individuals with X-linked agammaglobulinaemia and Mohr–
242–245.
Tranebjaerg syndrome caused by BTK and TIMM8A mutations, 9 Iossifov I, Ronemus M, Levy D, Wang Z, Hakker I, Rosenbaum J et al. De novo gene
respectively. The four patients described with the microdeletion disruptions in children on the autistic spectrum. Neuron 2012; 74: 285–299.
spanning BTK, TIMM8A, TAF7L and DRP2 presented the typical 10 Sanders SJ, Murtha MT, Gupta AR, Murdoch JD, Raubeson MJ, Willsey AJ et al.
X-linked agammaglobulinaemia and Mohr–Tranebjaerg syn- De novo mutations revealed by whole-exome sequencing are strongly associated
drome phenotypes, but also autism and language delay.43–45 with autism. Nature 2012; 485: 237–241.
& 2014 Macmillan Publishers Limited Molecular Psychiatry (2014), 784 – 790
Exome sequencing in multiplex autism families
C Toma et al
790
11 Schaaf CP, Sabo A, Sakai Y, Crosby J, Muzny D, Hawes A et al. Oligogenic 29 Anney R, Klei L, Pinto D, Almeida J, Bacchelli E, Baird G et al. Individual common
heterozygosity in individuals with high-functioning autism spectrum disorders. variants exert weak effects on the risk for autism spectrum disorderspi. Hum Mol
Hum Mol Genet 2011; 20: 3366–3375. Genet 2012; 21: 4781–4792.
12 Leblond CS, Heinrich J, Delorme R, Proepper C, Betancur C, Huguet G et al. 30 Kleppe R, Martinez A, Doskeland SO, Haavik J. The 14-3-3 proteins in regulation of
Genetic and functional analyses of SHANK2 mutations suggest a multiple hit cellular metabolism. Semin Cell Dev Biol 2011; 22: 713–719.
model of autism spectrum disorders. PLoS Genet 2012; 8: e1002521. 31 Foote M, Zhou Y. 14-3-3 Proteins in neurological disorders. Int J Biochem Mol Biol
13 Bailey A, Le Couteur A, Gottesman I, Bolton P, Simonoff E, Yuzda E et al. Autism as 2012; 3: 152–164.
a strongly genetic disorder: evidence from a British twin study. Psychol Med 1995; 32 Cardoso C, Leventer RJ, Ward HL, Toyo-Oka K, Chung J, Gross A et al. Refinement
25: 63–77. of a 400-kb critical region allows genotypic differentiation between isolated lis-
14 Sebat J, Lakshmi B, Malhotra D, Troge J, Lese-Martin C, Walsh T et al. Strong sencephaly, Miller–Dieker syndrome, and other phenotypes secondary to dele-
association of de novo copy number mutations with autism. Science 2007; 316: tions of 17p13.3. Am J Hum Genet 2003; 72: 918–930.
445–449. 33 Bruno DL, Anderlid BM, Lindstrand A, van Ravenswaaij-Arts C, Ganesamoorthy D,
15 Marshall CR, Noor A, Vincent JB, Lionel AC, Feuk L, Skaug J et al. Structural Lundin J et al. Further molecular and clinical delineation of co-locating 17p13.3
variation of chromosomes in autism spectrum disorder. Am J Hum Genet 2008; 82: microdeletions and microduplications that show distinctive phenotypes. J Med
477–488. Genet 2010; 47: 299–311.
16 Chahrour MH, Yu TW, Lim ET, Ataman B, Coulter ME, Hill RS et al. Whole-exome 34 Capra V, Mirabelli-Badenier M, Stagnaro M, Rossi A, Tassano E, Gimelli S et al.
sequencing and homozygosity analysis implicate depolarization-regulated Identification of a rare 17p13.3 duplication including the BHLHA9 and YWHAE
neuronal genes in autism. PLoS Genet 2012; 8: e1002635. genes in a family with developmental delay and behavioural problems. BMC Med
17 Puffenberger EG, Jinks RN, Wang H, Xin B, Fiorentini C, Sherman EA et al. Genet 2012; 13: 93.
A homozygous missense mutation in HERC2 associated with global develop- 35 Ikeda M, Hikita T, Taya S, Uraguchi-Asaki J, Toyo-oka K, Wynshaw-Boris A et al.
mental delay and autism spectrum disorder. Hum Mutat 2012; 33: 1639–1646. Identification of YWHAE, a gene encoding 14-3-3epsilon, as a possible suscept-
18 Lim ET, Raychaudhuri S, Sanders SJ, Stevens C, Sabo A, Macarthur DG et al. Rare ibility gene for schizophrenia. Hum Mol Genet 2008; 17: 3212–3222.
complete knockouts in humans: population distribution and significant role in 36 Middleton FA, Peng L, Lewis DA, Levitt P, Mirnics K. Altered expression of 14-3-3
autism spectrum disorders. Neuron 2013; 77: 235–242. genes in the prefrontal cortex of subjects with schizophrenia. Neuropsycho-
19 Li H, Durbin R. Fast and accurate short read alignment with Burrows–Wheeler pharmacology 2005; 30: 974–983.
transform. Bioinformatics 2009; 25: 1754–1760. 37 Ramocki MB, Bartnik M, Szafranski P, Kolodziejska KE, Xia Z, Bravo J et al.
20 Li H, Handsaker B, Wysoker A, Fennell T, Ruan J, Homer N et al. The Sequence Recurrent distal 7q11.23 deletion including HIP1 and YWHAG identified in
Alignment/Map format and SAMtools. Bioinformatics 2009; 25: 2078–2079. patients with intellectual disabilities, epilepsy, and neurobehavioral problems. Am
21 Puente XS, Pinyol M, Quesada V, Conde L, Ordonez GR, Villamor N et al. J Hum Genet 2010; 87: 857–865.
Whole-genome sequencing identifies recurrent mutations in chronic lymphocytic 38 Grover D, Verma R, Goes FS, Mahon PL, Gershon ES, McMahon FJ et al. Family-
leukaemia. Nature 2011; 475: 101–105. based association of YWHAH in psychotic bipolar disorder. Am J Med Genet B
22 Fuentes Fajardo KV, Adams D, Mason CE, Sincan M, Tifft C, Toro C et al. Detecting 2009; 150B: 977–983.
false-positive signals in exome sequencing. Hum Mutat 2012; 33: 609–613. 39 Pers TH, Hansen NT, Lage K, Koefoed P, Dworzynski P, Miller ML et al. Meta-
23 Kumar P, Henikoff S, Ng PC. Predicting the effects of coding non-synonymous analysis of heterogeneous data sources for genome-scale identification of risk
variants on protein function using the SIFT algorithm. Nat Protoc 2009; 4: genes in complex phenotypes. Genet Epidemiol 2011; 35: 318–332.
1073–1081. 40 Cheah PS, Ramshaw HS, Thomas PQ, Toyo-Oka K, Xu X, Martin S et al. Neurode-
24 Adzhubei IA, Schmidt S, Peshkin L, Ramensky VE, Gerasimova A, Bork P et al. velopmental and neuropsychiatric behaviour defects arise from 14-3-3zeta defi-
A method and server for predicting damaging missense mutations. Nat Methods ciency. Mol Psychiatry 2012; 17: 451–466.
2010; 7: 248–249. 41 English JA, Pennington K, Dunn MJ, Cotter DR. The neuroproteomics of schizo-
25 Medina I, Carbonell J, Pulido L, Madeira SC, Goetz S, Conesa A et al. Babelomics: phrenia. Biol Psychiatry 2011; 69: 163–172.
an integrative platform for the analysis of transcriptomics, proteomics and 42 Sherman DL, Wu LM, Grove M, Gillespie CS, Brophy PJ. Drp2 and periaxin form
genomic data with advanced functional profiling. Nucleic Acids Res 2010; 38: Cajal bands with dystroglycan but have distinct roles in Schwann cell growth.
W210–W213. J Neurosci 2012; 32: 9419–9428.
26 He WZ, Liu WQ, Zhong XQ, Chen XL, Li SY, Zhang HM et al. Analysis of de novo 43 Sediva A, Smith CI, Asplund AC, Hadac J, Janda A, Zeman J et al. Contiguous
copy number variations in a family affected with autism spectrum disorders using X-chromosome deletion syndrome encompassing the BTK, TIMM8A, TAF7L, and
high-resolution array-based comparative genomic hybridization. Zhonghua Yi Xue DRP2 genes. J Clin Immunol 2007; 27: 640–646.
Yi Chuan Xue Za Zhi 2012; 29: 266–269. 44 Jyonouchi H, Geng L, Toruner GA, Vinekar K, Feng D, Fitzgerald-Bocarsly P.
27 Guilmatre A, Dubourg C, Mosca AL, Legallic S, Goldenberg A, Drouin-Garraud V et Monozygous twins with a microdeletion syndrome involving BTK, DDP1, and
al. Recurrent rearrangements in synaptic and neurodevelopmental genes and two other genes; evidence of intact dendritic cell development and TLR
shared biologic pathways in schizophrenia, autism, and mental retardation. Arch responses. Eur J Pediatr 2008; 167: 317–321.
Gen Psychiatry 2009; 66: 947–956. 45 Arai T, Zhao M, Kanegane H, van Zelm MC, Futatani T, Yamada M et al. Genetic
28 Brooks-Kayal A. Epilepsy and autism spectrum disorders: are there common analysis of contiguous X-chromosome deletion syndrome encompassing the BTK
developmental mechanisms? Brain Dev 2010; 32: 731–738. and TIMM8A genes. J Hum Genet 2011; 56: 577–582.
Supplementary Information accompanies the paper on the Molecular Psychiatry website (http://www.nature.com/mp)
Molecular Psychiatry (2014), 784 – 790 & 2014 Macmillan Publishers Limited