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Introduction: Autism is a common neurodevelopmental condition with a complex genetic aetiology that includes contributions from monogenic and polygenic factors. Many autistic people have unmet healthcare needs that could be served by genomics-informed research and clinical trials. The primary aim of the European Autism GEnomics Registry (EAGER) is to establish a registry of participants with a diagnosis of autism or an associated rare genetic condition who have undergone whole-genome sequencing. The registry can facilitate recruitment for future clinical trials and research studies, based on genetic, clinical and phenotypic profiles, as well as participant preferences. The secondary aim of EAGER is to investigate the association between mental and physical health characteristics and participants' genetic profiles., Methods and Analysis: EAGER is a European multisite cohort study and registry and is part of the AIMS-2-TRIALS consortium. EAGER was developed with input from the AIMS-2-TRIALS Autism Representatives and representatives from the rare genetic conditions community. 1500 participants with a diagnosis of autism or an associated rare genetic condition will be recruited at 13 sites across 8 countries. Participants will be given a blood or saliva sample for whole-genome sequencing and answer a series of online questionnaires. Participants may also consent to the study to access pre-existing clinical data. Participants will be added to the EAGER registry and data will be shared externally through established AIMS-2-TRIALS mechanisms., Ethics and Dissemination: To date, EAGER has received full ethical approval for 11 out of the 13 sites in the UK (REC 23/SC/0022), Germany (S-375/2023), Portugal (CE-085/2023), Spain (HCB/2023/0038, PIC-164-22), Sweden (Dnr 2023-06737-01), Ireland (230907) and Italy (CET_62/2023, CEL-IRCCS OASI/24-01-2024/EM01, EM 2024-13/1032 EAGER). Findings will be disseminated via scientific publications and conferences but also beyond to participants and the wider community (eg, the AIMS-2-TRIALS website, stakeholder meetings, newsletters)., Competing Interests: Competing interests: In the past 3 years, TC has served as a paid consultant to F. Hoffmann-La Roche and Servier and has received royalties from Sage Publications and Guilford Publications. DGMM has received funding for a PhD studentship from Compass, and for consulting from Jaguar Therapeutics and Hoffman Le Roche. GM receives funding for an investigator-initiated study from Compass Pathways; no financial or other conflict of interest with the present study. SB discloses that he has in the last 3 years acted as an author, consultant, or lecturer for Medice, Roche and Linus Biotechnology. SB receives royalties for textbooks and diagnostic tools from Hogrefe, UTB, Ernst Reinhardt, Kohlhammer, and Liber, and is a partner at NeuroSupportSolutions International AB. CC is a full-time employee of Genentech and owns stocks or RSUs in Roche Holdings. MA is the UK chief investigator for a trial sponsored by Roche (a phase II, randomised, double-blind, placebo-controlled, parallel group study to evaluate the safety, efficacy and pharmacodynamics of 52 weeks of treatment with basmasanil in participants aged 2–14 years old with dup15q syndrome followed by a 2-year optional open-label extension). LB served on an advisory board to Kingdom therapeutics in 2022., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY. Published by BMJ.)

While over 100 genes have been associated with autism, little is known about the prevalence of variants affecting them in individuals without a diagnosis of autism. Nor do we fully appreciate the phenotypic diversity beyond the formal autism diagnosis. Based on data from more than 13,000 individuals with autism and 210,000 undiagnosed individuals, we estimated the odds ratios for autism associated to rare loss-of-function (LoF) variants in 185 genes associated with autism, alongside 2,492 genes displaying intolerance to LoF variants. In contrast to autism-centric approaches, we investigated the correlates of these variants in individuals without a diagnosis of autism. We show that these variants are associated with a small but significant decrease in fluid intelligence, qualification level and income and an increase in metrics related to material deprivation. These effects were larger for autism-associated genes than in other LoF-intolerant genes. Using brain imaging data from 21,040 individuals from the UK Biobank, we could not detect significant differences in the overall brain anatomy between LoF carriers and non-carriers. Our results highlight the importance of studying the effect of the genetic variants beyond categorical diagnosis and the need for more research to understand the association between these variants and sociodemographic factors, to best support individuals carrying these variants., (© 2023. The Author(s).)

Individuals with autism spectrum disorder (henceforth referred to as autism) display significant variation in clinical outcome. For instance, across age, some individuals' adaptive skills naturally improve or remain stable, while others' decrease. To pave the way for 'precision-medicine' approaches, it is crucial to identify the cross-sectional and, given the developmental nature of autism, longitudinal neurobiological (including neuroanatomical and linked genetic) correlates of this variation. We conducted a longitudinal follow-up study of 333 individuals (161 autistic and 172 neurotypical individuals, aged 6-30 years), with two assessment time points separated by ~12-24 months. We collected behavioural (Vineland Adaptive Behaviour Scale-II, VABS-II) and neuroanatomical (structural magnetic resonance imaging) data. Autistic participants were grouped into clinically meaningful "Increasers", "No-changers", and "Decreasers" in adaptive behaviour (based on VABS-II scores). We compared each clinical subgroup's neuroanatomy (surface area and cortical thickness at T1, ∆T (intra-individual change) and T2) to that of the neurotypicals. Next, we explored the neuroanatomical differences' potential genomic associates using the Allen Human Brain Atlas. Clinical subgroups had distinct neuroanatomical profiles in surface area and cortical thickness at baseline, neuroanatomical development, and follow-up. These profiles were enriched for genes previously associated with autism and for genes previously linked to neurobiological pathways implicated in autism (e.g. excitation-inhibition systems). Our findings suggest that distinct clinical outcomes (i.e. intra-individual change in clinical profiles) linked to autism core symptoms are associated with atypical cross-sectional and longitudinal, i.e. developmental, neurobiological profiles. If validated, our findings may advance the development of interventions, e.g. targeting mechanisms linked to relatively poorer outcomes., (© 2023. The Author(s).)

SHANK3-related Phelan-McDermid syndrome (PMS) is caused by a loss of the distal part of chromosome 22, including SHANK3, or by a pathological SHANK3 variant. There is an important genetic and phenotypic diversity among patients who can present with developmental delay, language impairments, autism, epilepsy, and other symptoms. SHANK3, encoding a synaptic scaffolding protein, is deleted in the majority of patients with PMS and is considered a major gene involved in the neurological impairments of the patients. However, differences in deletion size can influence clinical features, and in some rare cases, deletions at the 22q13 locus in individuals with SHANK3-unrelated PMS do not encompass SHANK3. These individuals with SHANK3-unrelated PMS still display a PMS-like phenotype. This suggests the participation of other 22q13 genes in the pathogenesis of PMS. Here, we review the biological function and potential implication in PMS symptoms of 110 genes located in the 22q13 region, focusing on 35 genes with evidence for association with neurodevelopmental disorders, including 13 genes for epilepsy and 11 genes for microcephaly and/or macrocephaly. Our review is restricted to the 22q13 region, but future large-scale studies using whole genome sequencing and deep-phenotyping are warranted to develop predictive models of clinical trajectories and to target specific medical and educational care for each individual with PMS., (Copyright © 2023 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

The substantial phenotypic heterogeneity in autism limits our understanding of its genetic etiology. To address this gap, here we investigated genetic differences between autistic individuals (n max  = 12,893) based on core and associated features of autism, co-occurring developmental disabilities and sex. We conducted a comprehensive factor analysis of core autism features in autistic individuals and identified six factors. Common genetic variants were associated with the core factors, but de novo variants were not. We found that higher autism polygenic scores (PGS) were associated with lower likelihood of co-occurring developmental disabilities in autistic individuals. Furthermore, in autistic individuals without co-occurring intellectual disability (ID), autism PGS are overinherited by autistic females compared to males. Finally, we observed higher SNP heritability for autistic males and for autistic individuals without ID. Deeper phenotypic characterization will be critical in determining how the complex underlying genetics shape cognition, behavior and co-occurring conditions in autism., (© 2022. The Author(s).)

Autism spectrum disorder (ASD) is a neurodevelopmental condition characterized by difficulties in social communication, but also great heterogeneity. To offer individualized medicine approaches, we need to better target interventions by stratifying autistic people into subgroups with different biological profiles and/or prognoses. We sought to validate neural responses to faces as a potential stratification factor in ASD by measuring neural (electroencephalography) responses to faces (critical in social interaction) in N = 436 children and adults with and without ASD. The speed of early-stage face processing (N170 latency) was on average slower in ASD than in age-matched controls. In addition, N170 latency was associated with responses to faces in the fusiform gyrus, measured with functional magnetic resonance imaging, and polygenic scores for ASD. Within the ASD group, N170 latency predicted change in adaptive socialization skills over an 18-month follow-up period; data-driven clustering identified a subgroup with slower brain responses and poor social prognosis. Use of a distributional data-driven cutoff was associated with predicted improvements of power in simulated clinical trials targeting social functioning. Together, the data provide converging evidence for the utility of the N170 as a stratification factor to identify biologically and prognostically defined subgroups in ASD.

Objective: Autism spectrum disorder (ASD) is a lifelong neurodevelopmental condition that is associated with significant difficulties in adaptive behavior and variation in clinical outcomes across the life span. Some individuals with ASD improve, whereas others may not change significantly, or regress. Hence, the development of "personalized medicine" approaches is essential. However, this requires an understanding of the biological processes underpinning differences in clinical outcome, at both the individual and subgroup levels, across the lifespan., Methods: The authors conducted a longitudinal follow-up study of 483 individuals (204 with ASD and 279 neurotypical individuals, ages 6-30 years), with assessment time points separated by ∼12-24 months. Data collected included behavioral data (Vineland Adaptive Behavior Scale-II), neuroanatomical data (structural MRI), and genetic data (DNA). Individuals with ASD were grouped into clinically meaningful "increasers," "no-changers," and "decreasers" in adaptive behavior. First, the authors compared neuroanatomy between outcome groups. Next, they examined whether deviations from the neurotypical neuroanatomical profile were associated with outcome at the individual level. Finally, they explored the observed neuroanatomical differences' potential genetic underpinnings., Results: Outcome groups differed in neuroanatomical features (cortical volume and thickness, surface area), including in "social brain" regions previously implicated in ASD. Also, deviations of neuroanatomical features from the neurotypical profile predicted outcome at the individual level. Moreover, neuroanatomical differences were associated with genetic processes relevant to neuroanatomical phenotypes (e.g., synaptic development)., Conclusions: This study demonstrates, for the first time, that variation in clinical (adaptive) outcome is associated with both group- and individual-level variation in anatomy of brain regions enriched for genes relevant to ASD. This may facilitate the move toward better targeted/precision medicine approaches.

The genetics of neurodevelopmental disorders (NDD) has made tremendous progress during the last few decades with the identification of more than 1,500 genes associated with conditions such as intellectual disability and autism. The functional roles of these genes are currently studied to uncover the biological mechanisms influencing the clinical outcome of the mutation carriers. To integrate the data, several databases and curated gene lists have been generated. Here, we provide an overview of the main databases focusing on the genetics of NDD, that are widely used by the medical and scientific communities, and extract a list of high confidence NDD genes (HC-NDD). This gene set can be used as a first filter for interpreting large scale omics dataset or for diagnostic purposes. Overall HC-NDD genes (N = 1,586) are expressed at very early stages of fetal brain development and enriched in several biological pathways such as chromosome organization, cell cycle, metabolism and synaptic function. Among those HC-NDD genes, 204 (12,9%) are listed in the synaptic gene ontology SynGO and are enriched in genes expressed after birth in the cerebellum and the cortex of the human brain. Finally, we point at several limitations regarding the relatively poor standardized information available, especially on the carriers of the mutations. Progress on the phenotypic characterization and genetic profiling of the carriers will be crucial to improve our knowledge on the biological mechanisms and on risk and protective factors for NDD., (Copyright © 2021. Published by Elsevier Inc.)

"Oligogenic inheritance" is used to describe cases where more than one rare pathogenic variant is observed in the same individual. While multiple variants can alter disease presentation, the necessity of multiple variants to instigate pathogenesis has not been addressed in amyotrophic lateral sclerosis (ALS). We sequenced ALS-associated genes in C9orf72-expansion-positive and negative ALS patients, alongside unaffected controls, to test the importance of oligogenicity and variant deleteriousness in ALS. We found that all groups had similar numbers of rare variants, but that variant severity was significantly higher in C9orf72-negative ALS cases, suggesting sufficiency of C9orf72 expansion to cause ALS alone.

The core diagnostic criteria for autism comprise two symptom domains - social and communication difficulties, and unusually repetitive and restricted behaviour, interests and activities. There is some evidence to suggest that these two domains are dissociable, though this hypothesis has not yet been tested using molecular genetics. We test this using a genome-wide association study ( N  = 51,564) of a non-social trait related to autism, systemising, defined as the drive to analyse and build systems. We demonstrate that systemising is heritable and genetically correlated with autism. In contrast, we do not identify significant genetic correlations between social autistic traits and systemising. Supporting this, polygenic scores for systemising are significantly and positively associated with restricted and repetitive behaviour but not with social difficulties in autistic individuals. These findings strongly suggest that the two core domains of autism are genetically dissociable, and point at how to fractionate the genetics of autism., Competing Interests: Competing interestsD.A.H. and the 23andMe Research Team are employees of 23andMe, Inc. The remaining authors declare no competing interests.

[This corrects the article DOI: 10.1038/s41525-017-0035-2.].

Essential Tremor is a prevalent neurological disorder of unknown etiology. Studies suggest that genetic factors contribute to this pathology. To date, no causative mutations in a gene have been reproducibly reported. All three structures of the olivocerebellar motor circuitry have been linked to Essential Tremor. We postulated that genes enriched for their expression in the olivocerebellar circuitry would be more susceptible to harbor mutations in Essential Tremor patients. A list of 11 candidate genes, enriched for their expression in the olivocerebellar circuitry, was assessed for their variation spectrum and frequency in a cohort of Canadian Essential Tremor cases. Our results from this list of 11 candidate genes do not support an association for Essential Tremor in our cohort of Canadian cases. The heterogenic nature of ET and modest size of the cohort used in this study are two confounding factors that could explain these results.

Background: SMPD1 (acid-sphingomyelinase) variants have been associated with Parkinson's disease in recent studies. The objective of this study was to further investigate the role of SMPD1 mutations in PD., Methods: SMPD1 was sequenced in 3 cohorts (Israel Ashkenazi Jewish cohort, Montreal/Montpellier, and New York), including 1592 PD patients and 975 controls. Additional data were available for 10,709 Ashkenazi Jewish controls. Acid-sphingomyelinase activity was measured by a mass spectrometry-based assay in the New York cohort. α-Synuclein levels were measured in vitro following CRISPR/Cas9-mediated knockout and siRNA knockdown of SMPD1 in HeLa and BE(2)-M17 cells. Lysosomal localization of acid-sphingomyelinase with different mutations was studied, and in silico analysis of their effect on acid-sphingomyelinase structure was performed., Results: SMPD1 mutations were associated with PD in the Ashkenazi Jewish cohort, as 1.4% of PD patients carried the p.L302P or p.fsP330 mutation, compared with 0.37% in 10,709 Ashkenazi Jewish controls (OR, 3.7; 95%CI, 1.6-8.2; P = 0.0025). In the Montreal/Montpellier cohort, the p.A487V variant was nominally associated with PD (1.5% versus 0.14%; P = 0.0065, not significant after correction for multiple comparisons). Among PD patients, reduced acid-sphingomyelinase activity was associated with a 3.5- to 5.8-year earlier onset of PD in the lowest quartile versus the highest quartile of acid-sphingomyelinase activity (P = 0.01-0.001). We further demonstrated that SMPD1 knockout and knockdown resulted in increased α-synuclein levels in HeLa and BE(2)-M17 dopaminergic cells and that the p.L302P and p.fsP330 mutations impair the traffic of acid-sphingomyelinase to the lysosome., Conclusions: Our results support an association between SMPD1 variants, acid-sphingomyelinase activity, and PD. Furthermore, they suggest that reduced acid-sphingomyelinase activity may lead to α-synuclein accumulation. © 2019 International Parkinson and Movement Disorder Society., (© 2019 International Parkinson and Movement Disorder Society.)

Objective: To test for somatic C9orf72 hexanucleotide repeat expansion (HRE) and hexanucleotide repeat length instability in the spinal cord of amyotrophic lateral sclerosis (ALS) cases., Methods: Whole and partial spinal cords of 19 ALS cases were dissected into transversal sections (5 mm thick). The presence of C9orf72 HRE was tested in each independent section using RepeatPrimed PCR and amplicon-size genotyping. Index measures for the testing of mosaicism were obtained through serial dilutions of genomic DNA from an individual carrying a germline C9orf72 HRE in the genomic DNA of an individual without a C9orf72 HRE., Results: None of the sections examined supported the presence of a subpopulation of cells with a C9orf72 HRE. Moreover, the C9orf72 hexanucleotide repeat lengths measured were identical across all the spinal cord sections of each individual patient., Conclusions: We did not observe somatic instability of the C9orf72 HRE in disease relevant tissues of ALS cases.

Autism spectrum disorder (ASD) is a set of neurodevelopmental conditions characterized by early-onset difficulties in social communication and unusually restricted, repetitive behavior and interests. Parental consanguinity may lead to higher risk of ASD and to more severe clinical presentations in the offspring. Studies of ASD families with high inbreeding enable the identification of inherited variants of this disorder particularly those with an autosomal recessive pattern of inheritance. In our study, using copy number variants (CNV) analysis, we identified a rare homozygous deletion in 2p11.2 region that affects ELMOD3, CAPG, and SH2D6 genes in a boy with ASD, intellectual disability (ID), and hearing impairment (HI). This deletion may reveal a new contiguous deletion syndrome in which ELMOD3, known to be implicated in autosomal recessive deafness underlies the HI of the proband and CAPG, member of actin regulatory proteins involved in cytoskeletal dynamic, an important function for brain development and activity, underlies the ASD/ID phenotype. A possible contribution of SH2D6 gene, as a part of a chimeric gene, to the clinical presentation of the patient is discussed. Our result supports the implication of ELMOD3 in hearing loss and highlights the potential clinical relevance of 2p11.2 deletion in autism and/or intellectual disability.

The number of genes associated with autism is increasing, but few studies have been performed on epidemiological cohorts and in isolated populations. Here, we investigated 357 individuals from the Faroe Islands including 36 individuals with autism, 136 of their relatives and 185 non-autism controls. Data from SNP array and whole exome sequencing revealed that individuals with autism had a higher burden of rare exonic copy-number variants altering autism associated genes (deletions ( p   =  0.0352) or duplications ( p   =  0.0352)), higher inbreeding status ( p   =  0.023) and a higher load of rare homozygous deleterious variants ( p   =  0.011) compared to controls. Our analysis supports the role of several genes/loci associated with autism (e.g., NRXN1 , ADNP , 22q11 deletion) and identified new truncating (e.g. , GRIK2 , ROBO1, NINL , and IMMP2L ) or recessive deleterious variants (e.g. , KIRREL3 and CNTNAP2 ) affecting autism-associated genes. It also revealed three genes involved in synaptic plasticity, RIMS4 , KALRN , and PLA2G4A , carrying de novo deleterious variants in individuals with autism without intellectual disability. In summary, our analysis provides a better understanding of the genetic architecture of autism in isolated populations by highlighting the role of both common and rare gene variants and pointing at new autism-risk genes. It also indicates that more knowledge about how multiple genetic hits affect neuronal function will be necessary to fully understand the genetic architecture of autism., Competing Interests: The authors declare no competing interests.

The synaptic protein SHANK3 encodes a multidomain scaffold protein expressed at the postsynaptic density of neuronal excitatory synapses. We previously identified de novo SHANK3 mutations in patients with autism spectrum disorders (ASD) and showed that SHANK3 represents one of the major genes for ASD. Here, we analyzed the pyramidal cortical neurons derived from induced pluripotent stem cells from four patients with ASD carrying SHANK3 de novo truncating mutations. At 40-45 days after the differentiation of neural stem cells, dendritic spines from pyramidal neurons presented variable morphologies: filopodia, thin, stubby and muschroom, as measured in 3D using GFP labeling and immunofluorescence. As compared to three controls, we observed a significant decrease in SHANK3 mRNA levels (less than 50% of controls) in correlation with a significant reduction in dendritic spine densities and whole spine and spine head volumes. These results, obtained through the analysis of de novo SHANK3 mutations in the patients' genomic background, provide further support for the presence of synaptic abnormalities in a subset of patients with ASD.

Background: MAPT haplotypes are associated with PD, but their association with rapid eye movement sleep behavior disorder is unclear., Objective: To study the role of MAPT variants in rapid eye movement sleep behavior disorder., Methods: Two cohorts were included: (A) PD (n = 600), rapid eye movement sleep behavior disorder (n = 613) patients, and controls (n = 981); (B) dementia with Lewy bodies patients with rapid eye movement sleep behavior disorder (n = 271) and controls (n = 950). MAPT-associated variants and the entire coding sequence of MAPT were analyzed. Age-, sex-, and ethnicity-adjusted analyses were performed to examine the association between MAPT, PD, and rapid eye movement sleep behavior disorder., Results: MAPT-H2 variants were associated with PD (odds ratios: 0.62-0.65; P = 0.010-0.019), but not with rapid eye movement sleep behavior disorder. In PD, the H1 haplotype odds ratio was 1.60 (95% confidence interval: 1.12-2.28; P = 0.009), and the H2 odds ratio was 0.68 (95% confidence interval: 0.48-0.96; P = 0.03). The H2/H1 haplotypes were not associated with rapid eye movement sleep behavior disorder., Conclusions: Our results confirm the protective effect of the MAPT-H2 haplotype in PD, and define its components. Furthermore, our results suggest that MAPT does not play a major role in rapid eye movement sleep behavior disorder, emphasizing different genetic background than in PD in this locus. © 2018 International Parkinson and Movement Disorder Society., (© 2018 International Parkinson and Movement Disorder Society.)

Autism Spectrum Disorders (ASD) are heterogeneous neurodevelopmental disorders with a complex genetic architecture. They are characterized by impaired social communication, stereotyped behaviors and restricted interests and are frequently associated with comorbidities such as intellectual disability, epilepsy and severe sleep disorders. Hyperserotonemia and low melatonin levels are among the most replicated endophenotypes reported in ASD, but their genetic causes remain largely unknown. Based on the biochemical profile of 717 individuals including 213 children with ASD, 128 unaffected siblings and 376 parents and other relatives, we estimated the heritability of whole-blood serotonin, platelet N-acetylserotonin (NAS) and plasma melatonin levels, as well as the two enzymes arylalkylamine N-acetyltransferase (AANAT) and acetylserotonin O-methyltransferase (ASMT) activities measured in platelets. Overall, heritability was higher for NAS (0.72 ± 0.091) and ASMT (0.59 ± 0.097) compared with serotonin (0.31 ± 0.078), AANAT (0.34 ± 0.077) and melatonin (0.22 ± 0.071). Bivariate analyses showed high phenotypic and genetic correlations between traits of the second step of the metabolic pathway (NAS, ASMT and melatonin) indicating the contribution of shared genetic factors. A better knowledge of the heritability of the melatonin synthesis variability constitutes an important step to identify the factors that perturb this pathway in individuals with ASD.

Phelan-McDermid syndrome (PMS) is characterized by a variety of clinical symptoms with heterogeneous degrees of severity, including intellectual disability (ID), absent or delayed speech, and autism spectrum disorders (ASD). It results from a deletion of the distal part of chromosome 22q13 that in most cases includes the SHANK3 gene. SHANK3 is considered a major gene for PMS, but the factors that modulate the severity of the syndrome remain largely unknown. In this study, we investigated 85 patients with different 22q13 rearrangements (78 deletions and 7 duplications). We first explored the clinical features associated with PMS, and provide evidence for frequent corpus callosum abnormalities in 28% of 35 patients with brain imaging data. We then mapped several candidate genomic regions at the 22q13 region associated with high risk of clinical features, and suggest a second locus at 22q13 associated with absence of speech. Finally, in some cases, we identified additional clinically relevant copy-number variants (CNVs) at loci associated with ASD, such as 16p11.2 and 15q11q13, which could modulate the severity of the syndrome. We also report an inherited SHANK3 deletion transmitted to five affected daughters by a mother without ID nor ASD, suggesting that some individuals could compensate for such mutations. In summary, we shed light on the genotype-phenotype relationship of patients with PMS, a step towards the identification of compensatory mechanisms for a better prognosis and possibly treatments of patients with neurodevelopmental disorders., Competing Interests: The authors declare that they have no competing financial interests.

Objective: To assess the contribution of variants in STK32B , PPARGC1A , and CTNNA3 as essential tremor (ET) predisposing factors following their association in a 2-stage genome-wide association study (GWAS)., Methods: The coding regions of these genes was examined for the presence of rare variants using two approaches: (1) Looking at whole-exome and whole-genome sequencing data of 14 autosomal dominant multiplex ET families. (2) Conducting a targeted massive parallel sequencing to examine the three genes in cohorts of 269 ET cases and 287 control individuals. The cumulative impact of rare variants was assessed using SKAT-O analyses using (1) all variants, (2) only rare variants, and (3) only the rare variants altering the mRNA., Results: Thirty-four variants were identified. No difference emerged regarding the distributions of individual variants (or gene) between cases and controls., Conclusion: No rare exonic variants further validated one of these genes as a risk factor for ET. The recent GWAS offers promising avenues, but the genetic heterogeneity of ET is nonetheless challenging for the validation of risk factors, and ultimately larger cohorts of cases should help to overcome this task.

Introduction: Mutations in teneurin transmembrane protein 4 were reported to be a risk factor for essential tremor, but the relevance of this across different population remains to be examined. The aim of this study was to determine the frequency and spectrum of variations in teneurin transmembrane protein 4 in a cohort of Canadian essential tremor cases., Methods: The coding portion of teneurin transmembrane protein 4 was sequenced in 269 unrelated essential tremor cases and 288 matched control individuals using a targeted and high-throughput sequencing approach., Results: A total of 157 single nucleotide variations were identified, and from these 99 were a missense or nonsense mutation. A total of 68 cases were carriers of ≥1 rare missense or nonsense mutations, and 39 control individuals were carriers of the same types of variations. Gene-based association tests were used to jointly analyze the single nucleotide variations., Conclusions: Our results do not support a positive association between teneurin transmembrane protein 4 and the Canadian population. © 2017 International Parkinson and Movement Disorder Society., (© 2017 International Parkinson and Movement Disorder Society.)

The present study aimed to examine whether the APOE ε4 allele, associated with dementia with Lewy bodies (DLB), and possibly with dementia in Parkinson's disease (PD), is also associated with idiopathic rapid eye movement sleep behavior disorder (RBD). Two single nucleotide polymorphisms, rs429358 and rs7412, were genotyped in RBD patients (n = 480) and in controls (n = 823). APOE ε4 allele frequency was 0.14 among RBD patients and 0.13 among controls (OR = 1.11, 95% CI: 0.88-1.40, p = 0.41). APOE ε4 allele frequencies were similar in those who converted to DLB (0.14) and those who converted to Parkinson's disease (0.12) or multiple system atrophy (0.14, p = 1.0). The APOE ε4 allele is neither a risk factor for RBD nor it is associated with conversion from RBD to DLB or other synucleinopathies., (Copyright © 2016 Elsevier Inc. All rights reserved.)

DNAJC13 mutations have been suggested to cause Parkinson's disease (PD), yet subsequent studies reported conflicting results on this association. In the present study, we sequenced the coding region of DNAJC13 in a French-Canadian/French cohort of 528 PD patients and 692 controls. A total of 62 (11.7%) carriers of rare DNAJC13 variants were identified among the PD patients compared with 82 (11.8%) among controls (p = 1.0). Two variants that were previously suggested to be associated with PD, p.R1516H and p.L2170W, were identified with similar directions of association as previously reported. The p.R1516H was found in 2 (0.4%) patients versus 6 (0.9%, nonsignificant) controls and the p.L2170W variant was found in 9 (1.7%) patients and 5 (0.7%, nonsignificant) controls. Meta-analysis with previous reports resulted in odds ratios of 0.32 (95% confidence interval = 0.15-0.68, p = 0.0037) and 2.68 (95% confidence interval = 1.32-5.42, p = 0.007), respectively. Our results provide some support for the possibility that specific DNAJC13 variants may play a minor role in PD susceptibility, although studies in additional populations are necessary., (Copyright © 2016 Elsevier Inc. All rights reserved.)

To identify genetic factors contributing to amyotrophic lateral sclerosis (ALS), we conducted whole-exome analyses of 1,022 index familial ALS (FALS) cases and 7,315 controls. In a new screening strategy, we performed gene-burden analyses trained with established ALS genes and identified a significant association between loss-of-function (LOF) NEK1 variants and FALS risk. Independently, autozygosity mapping for an isolated community in the Netherlands identified a NEK1 p.Arg261His variant as a candidate risk factor. Replication analyses of sporadic ALS (SALS) cases and independent control cohorts confirmed significant disease association for both p.Arg261His (10,589 samples analyzed) and NEK1 LOF variants (3,362 samples analyzed). In total, we observed NEK1 risk variants in nearly 3% of ALS cases. NEK1 has been linked to several cellular functions, including cilia formation, DNA-damage response, microtubule stability, neuronal morphology and axonal polarity. Our results provide new and important insights into ALS etiopathogenesis and genetic etiology.

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are overlapping, fatal neurodegenerative disorders in which the molecular and pathogenic basis remains poorly understood. Ubiquitinated protein aggregates, of which TDP-43 is a major component, are a characteristic pathological feature of most ALS and FTD patients. Here we use genome-wide linkage analysis in a large ALS/FTD kindred to identify a novel disease locus on chromosome 16p13.3. Whole-exome sequencing identified a CCNF missense mutation at this locus. Interrogation of international cohorts identified additional novel CCNF variants in familial and sporadic ALS and FTD. Enrichment of rare protein-altering CCNF variants was evident in a large sporadic ALS replication cohort. CCNF encodes cyclin F, a component of an E3 ubiquitin-protein ligase complex (SCF(Cyclin F)). Expression of mutant CCNF in neuronal cells caused abnormal ubiquitination and accumulation of ubiquitinated proteins, including TDP-43 and a SCF(Cyclin F) substrate. This implicates common mechanisms, linked to protein homeostasis, underlying neuronal degeneration.

Juvenile amyotrophic lateral sclerosis (jALS) is characterized by progressive upper and lower motor neuron degeneration leading to facial muscle spasticity, spastic dysarthria, and spastic gait with an early onset (before 25 years old). Unlike adult-onset amyotrophic lateral sclerosis (ALS), patients with jALS tend to have slower progression of motor neuron disease and prolonged survival to a normal life expectancy. Mutations in FUS gene have been reported in jALS,(1) including p.P525L mutation that has been consistently associated with early onset and aggressive presentation.(2) Here, we report a patient carrying p.P525L FUS mutation and experiencing an aggressive course of ALS presenting with dysphonia and diplopia.

Introduction: The most common forms of amyotrophic lateral sclerosis and frontotemporal dementia are caused by a large GGGGCC repeat expansion in the first intron of the C9orf72 gene. The repeat-containing intron should be degraded after being spliced out, however GGGGCC repeat-containing RNA species either accumulate in nuclear foci or are exported to the cytoplasm where they are translated into potentially toxic dipeptide repeat proteins by repeat-associated non-AUG-initiated (RAN) translation., Results: In order to determine the mechanisms of repeat-containing intron misprocessing, we have analyzed C9orf72 transcripts in lymphoblasts from C9orf72 expansion carriers (n = 15) and control individuals (n = 15). We have identified polyadenylated C9orf72 RNA species retaining the repeat-containing intron and in which downstream exons are spliced correctly resulting in a C9orf72 mRNA with an enlarged 5'-UTR containing the GGGGCC repeats. Intron-retaining transcripts are produced from both wild-type and mutant alleles. Intron-retaining C9orf72 transcripts were also detected in brain with a 2.7 fold increase measured in the frontal cortex from heterozygous expansion carriers (n = 11) compared to controls (n = 10). The level of intron-retaining transcripts was increased 5.9 fold in a case homozygous for the expansion. We also show that a large proportion of intron 1-retaining C9orf72 transcripts accumulate in the nucleus., Conclusions: Retention of the repeat-containing intron in mature C9orf72 mRNA can potentially explain nuclear foci formation as well as nuclear export of GGGGCC repeat RNA and suggests that the misprocessing of C9orf72 transcripts initiates the pathogenic process caused by C9orf72 hexanucleotide repeat expansions as well as provides the basis for novel therapeutic strategies.

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disorder characterized by an extensive loss of motor neurons in the primary motor cortex, brainstem, and spinal cord. Genetic studies report a high heritability of ALS. Recently, whole-exome sequencing analysis of familial ALS (FALS) patients allowed the identification of missense variations within the MATR3 gene. MATR3 was previously associated to distal myopathy 2 and encodes for a nuclear matrix and DNA/RNA binding protein that has been shown to interact with TDP43 in an RNA-dependent manner. Here, we assessed the MATR3 mutation frequency in French-Canadian ALS and control individuals (nFALS = 83, sporadic ALS [nSALS] = 164, and ncontrols = 162) and showed that MATR3 mutations were found in 0%, 1.8%, and 0% of FALS, SALS, and controls, respectively. Interestingly, among the mutations identified in SALS, the splicing mutation c.48+1G>T was found to result in the insertion of 24 amino acids in MATR3 protein. These findings further support the role of MATR3 in ALS, and more studies are needed to shed more light on MATR3 proteinopathy., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Jacobsen syndrome (JS) is characterized by intellectual disability and higher risk for autism spectrum disorders (ASD). All patients with JS are carriers of contiguous de novo deletions of 11q24.2-25, but the causative genes remain unknown. Within the critical interval, we hypothesized that haploinsufficiency of the neuronal cell adhesion molecule Neurotrimin (NTM) might increase the risk for ASD and could affect brain structure volumes. We searched for deleterious mutations affecting NTM in 1256 ASD patients and 1287 controls, using SNP arrays, and by direct sequencing of 250 ASD patients and 180 controls. We compared our results to those obtained from independent cohorts of ASD patients and controls. We identified two patients with Copy Number Variants (CNV) encompassing NTM, one with a large de novo deletion, and a clinical phenotype of JS (including macrocephaly), and a second with a paternally inherited duplication, not consistent with JS. Interestingly, no similar CNVs were observed in controls. We did not observe enrichment for deleterious NTM mutations in our cohort. We then explored if the macrocephaly in the patient with JS was associated with a homogeneous increase of brain structures volumes using automatic segmentation. Compared to subjects without NTM micro-rearrangements (n=188), the patient had an increased volume of the sub-cortical structures but a decrease of the occipital gray matter. Finally our explorations could not incriminate NTM as a susceptibility gene for ASD, but provides new information on the impact of the 11q24.2-25 deletion on brain anatomy., (© 2015 Wiley Periodicals, Inc.)

Background: Restless legs syndrome (RLS) is a common disorder, with several known genetic risk factors, yet the actual genetic causes are unclear., Methods: Whole-exome sequencing (WES) was performed in seven RLS families, focusing on six known genetic loci: MEIS1, BTBD9, PTPRD, MAP2K5/SKOR1, TOX3, and rs6747972. Genotyping using specific TaqMan assays was performed in two case-control cohorts (627 patients and 410 controls), and in a familial cohort (n = 718)., Results: WES identified two candidate GLO1 variants (within the BTBD9 locus), p.E111A and the promoter variant c.-7C>T, both co-segregated with the disease in four families. The GLO1 p.E111A variant was associated with RLS in the French-Canadian cohort (odds ratio, OR = 1.38, p = 0.02), and demonstrated a similar trend in the US cohort (OR = 1.26, p = 0.09, combined analysis OR = 1.28, p = 0.009). However, the original genome-wide association study (GWAS) marker, BTBD9 rs9357271, had stronger association with RLS (OR = 1.84, p = 0.0003). Conditional haplotype analysis, controlling for the effect of the BTBD9 variant rs9357271, demonstrated that the association of GLO1 p.E111A turned insignificant (p = 0.54). In the familial cohort, the two GLO1 variants were not associated with RLS. Other variants in the SKOR1 (p.W200R and p.A672V) and PTPRD (p.R995C, p.Q447E, p.T781A, p.Q447E, and c.551-4C > G) genes, did not co-segregate with the disease., Conclusions: The GLO1 variations studied here are not the source of association of the BTBD9 locus with RLS. It is likely that the genetic variants affecting RLS susceptibility are located in regulatory regions., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Rapid eye movement sleep behavior disorder and GBA mutations are both associated with Parkinson's disease. The GBA gene was sequenced in idiopathic rapid eye movement sleep behavior disorder patients (n = 265), and compared to controls (n = 2240). Rapid eye movement sleep behavior disorder questionnaire was performed in an independent Parkinson's disease cohort (n = 120). GBA mutations carriers had an OR of 6.24 (10.2% in patients vs. 1.8% in controls, P < 0.0001) for rapid eye movement sleep behavior disorder, and among Parkinson's disease patients, the OR for mutation carriers to have probable rapid eye movement sleep behavior disorder was 3.13 (P = 0.039). These results demonstrate that rapid eye movement sleep behavior disorder is associated with GBA mutations, and that combining genetic and prodromal data may assist in identifying individuals susceptible to Parkinson's disease.

Disruption of endoplasmic reticulum (ER) proteostasis is a salient feature of amyotrophic lateral sclerosis (ALS). Upregulation of ER foldases of the protein disulfide isomerase (PDI) family has been reported in ALS mouse models and spinal cord tissue and body fluids derived from sporadic ALS cases. Although in vitro studies suggest a neuroprotective role of PDIs in ALS, the possible contribution of genetic mutations of these ER foldases in the disease process remains unknown. Interestingly, intronic variants of the PDIA1 gene were recently reported as a risk factor for ALS. Here, we initially screened for mutations in two major PDI genes (PDIA1/P4HB and PDIA3/ERp57) in a US cohort of 96 familial and 96 sporadic ALS patients using direct DNA sequencing. Then, 463 familial and 445 sporadic ALS patients from two independent cohorts were also screened for mutations in these two genes using whole exome sequencing. A total of nine PDIA1 missense variants and seven PDIA3 missense variants were identified in 16 ALS patients. We have identified several novel and rare single nucleotide polymorphisms (SNPs) in both genes that are enriched in ALS cases compared with a large group of control subjects showing a frequency of around 1% in ALS cases. The possible biological and structural impact of these ALS-linked PDI variants is also discussed., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Background: It is currently under debate whether there is a sex effect in LRRK2-associated Parkinson disease (PD), as several studies suggested such effect while others did not., Methods: All case-control studies describing LRRK2 mutations and PD were examined, and papers with data on sex and LRRK2 mutations in both patients and controls were included (n = 17) in a sex-stratified meta-analysis. Additional studies (n = 33) that included data on male:female ratio only in patients with LRRK2 mutations, were included in further analysis of male:female ratio in LRRK2-assocoiated PD patients., Results: Similar risk estimates were calculated for men and women. Among men, LRRK2 mutation carriers had a pooled OR for PD of 4.20 (95% CI 2.95-5.99, p < 0.0001) and among women, LRRK2 mutation carriers had a pooled OR for PD of 4.73 (95% CI 3.26-6.86, p < 0.0001). Similar risk estimates for men and women were also observed when analysing specific LRRK2 mutations. A total of 1080 LRRK2-associated PD patients with sex information were identified. The male:female ratio was 1.02:1.00 (50.6% men and 49.4% women)., Conclusion: While sporadic PD is characterized by a sex effect, with more affected men than women, LRRK2-associated PD lacks a sex effect, as typically seen in autosomal dominant traits., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Rapid eye movement (REM) sleep behavior disorder (RBD) is a prodromal condition for Parkinson's disease (PD) and other synucleinopathies, which often occurs many years before the onset of PD. We analyzed 261 RBD patients and 379 controls for nine PD-associated SNPs and examined their effects, first upon on RBD risk and second, on eventual progression to synucleinopathies in a prospective follow-up in a subset of patients. The SCARB2 rs6812193 (OR = 0.67, 95 % CI = 0.51-0.88, p = 0.004) and the MAPT rs12185268 (OR-0.43, 95 % CI-0.26-0.72, p = 0.001) were associated with RBD in different models. Kaplan-Meier survival analysis in a subset of RBD patients (n = 56), demonstrated that homozygous carriers of the USP25 rs2823357 SNP had progressed to synucleinopathies faster than others (log-rank p = 0.003, Breslow p = 0.005, Tarone-Ware p = 0.004). As a proof-of-concept study, these results suggest that RBD may be associated with at least a subset of PD-associated genes, and demonstrate that combining genetic and prodromal clinical data may help identifying individuals that are either more or less susceptible to develop synucleinopathies. More studies are necessary to replicate these results, and identify more genetic factors affecting progression from RBD to synucleinopathies.

Background: Cohen Syndrome (COH1) is a rare autosomal recessive disorder, principally identified by ocular, neural and muscular deficits. We identified three large consanguineous Pakistani families with intellectual disability and in some cases with autistic traits., Methods: Clinical assessments were performed in order to allow comparison of clinical features with other VPS13B mutations. Homozygosity mapping followed by whole exome sequencing and Sanger sequencing strategies were used to identify disease-related mutations., Results: We identified two novel homozygous deletion mutations in VPS13B, firstly a 1 bp deletion, NM&#95;017890.4:c.6879delT; p.Phe2293Leufs*24, and secondly a deletion of exons 37-40, which co-segregate with affected status. In addition to COH1-related traits, autistic features were reported in a number of family members, contrasting with the "friendly" demeanour often associated with COH1. The c.6879delT mutation is present in two families from different regions of the country, but both from the Baloch sub-ethnic group, and with a shared haplotype, indicating a founder effect among the Baloch population., Conclusion: We suspect that the c.6879delT mutation may be a common cause of COH1 and similar phenotypes among the Baloch population. Additionally, most of the individuals with the c.6879delT mutation in these two families also present with autistic like traits, and suggests that this variant may lead to a distinct autistic-like COH1 subgroup.

Introduction: Several studies proposed that Restless Legs Syndrome (RLS) and Parkinson disease (PD) may be clinically and/or etiologically related. To examine this hypothesis, we aimed to determine whether the known RLS genetic markers may be associated with PD risk, as well as with PD subtype., Methods: Two case-control cohorts from Tel-Aviv and New-York, including 1133 PD patients and 867 controls were genotyped for four RLS-related SNPs in the genes MEIS1, BTBD9, PTPRD and MAP2K5/SKOR1. The association between genotype, PD risk and phenotype was tested using multivariate regression models., Results: None of the tested SNPs was significantly associated with PD risk, neither in any individual cohort nor in the combined analysis after correction for multiple comparisons. The MAP2K5/SKOR1 marker rs12593813 was associated with higher frequency of tremor in the Tel-Aviv cohort (61.0% vs. 46.5%, p = 0.001, dominant model). However, the risk allele for tremor in this gene has been associated with reduced RLS risk. Moreover, this association did not replicate in Tremor-dominant PD patients from New-York., Conclusion: RLS genetic risk markers are not associated with increased PD risk or subtype in the current study. Together with previous genetic, neuropathological and epidemiologic studies, our results further strengthen the notion that RLS and PD are likely to be distinct entities., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Amyotrophic lateral sclerosis (ALS) is a devastating neurological disease with no effective treatment. We report the results of a moderate-scale sequencing study aimed at increasing the number of genes known to contribute to predisposition for ALS. We performed whole-exome sequencing of 2869 ALS patients and 6405 controls. Several known ALS genes were found to be associated, and TBK1 (the gene encoding TANK-binding kinase 1) was identified as an ALS gene. TBK1 is known to bind to and phosphorylate a number of proteins involved in innate immunity and autophagy, including optineurin (OPTN) and p62 (SQSTM1/sequestosome), both of which have also been implicated in ALS. These observations reveal a key role of the autophagic pathway in ALS and suggest specific targets for therapeutic intervention., (Copyright © 2015, American Association for the Advancement of Science.)

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by the selective death of motor neurons. Causative mutations in the global RNA-processing proteins TDP-43 and FUS among others, as well as their aggregation in ALS patients, have identified defects in RNA metabolism as an important feature in this disease. Lethal congenital contracture syndrome 1 and lethal arthrogryposis with anterior horn cell disease are autosomal recessive fetal motor neuron diseases that are caused by mutations in another global RNA-processing protein, hGle1. In this study, we carried out the first screening of GLE1 in ALS patients (173 familial and 760 sporadic) and identified 2 deleterious mutations (1 splice site and 1 nonsense mutation) and 1 missense mutation. Functional analysis of the deleterious mutants revealed them to be unable to rescue motor neuron pathology in zebrafish morphants lacking Gle1. Furthermore, in HeLa cells, both mutations caused a depletion of hGle1 at the nuclear pore where it carries out an essential role in nuclear export of mRNA. These results suggest a haploinsufficiency mechanism and point to a causative role for GLE1 mutations in ALS patients. This further supports the involvement of global defects in RNA metabolism in ALS., (© The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Amyotrophic lateral sclerosis (ALS) is a fatal late onset neurological disorder characterized by motor neuron degeneration in the primary motor cortex, brainstem and spinal cord. The majority of cases are sporadic (SALS) and only 5-10% have a family history (FALS). FALS cases show a high heritability and this has enabled the identification of several genetic triggers, of which mutations in SOD1, FUS, TARDBP and C9ORF72 are the most frequent. While such advances have contributed to our current understanding of the causes of most cases of FALS and their underlying pathophysiological consequences, they only explain a small fraction of SALS with the etiology of most SALS cases remaining unexplained. Here, we review past and current methods used for the identification of FALS and SALS associated genes and propose a risk-based classification for these. We also discuss how the growing number of whole exome/genome sequencing datasets prepared from SALS cases, and control individuals, may reveal novel insights into the genetic etiology of SALS; for instance through revealing increased mutation burden rates across genes or genomic regions that were not previously associated with ALS or through allowing the examination of a potential "oligogenic" mechanism of the disease. Finally we summarize the three most recently discovered 'high risk' genes in ALS., (Copyright © 2014 Elsevier Inc. All rights reserved.)

SHANK genes code for scaffold proteins located at the post-synaptic density of glutamatergic synapses. In neurons, SHANK2 and SHANK3 have a positive effect on the induction and maturation of dendritic spines, whereas SHANK1 induces the enlargement of spine heads. Mutations in SHANK genes have been associated with autism spectrum disorders (ASD), but their prevalence and clinical relevance remain to be determined. Here, we performed a new screen and a meta-analysis of SHANK copy-number and coding-sequence variants in ASD. Copy-number variants were analyzed in 5,657 patients and 19,163 controls, coding-sequence variants were ascertained in 760 to 2,147 patients and 492 to 1,090 controls (depending on the gene), and, individuals carrying de novo or truncating SHANK mutations underwent an extensive clinical investigation. Copy-number variants and truncating mutations in SHANK genes were present in ∼1% of patients with ASD: mutations in SHANK1 were rare (0.04%) and present in males with normal IQ and autism; mutations in SHANK2 were present in 0.17% of patients with ASD and mild intellectual disability; mutations in SHANK3 were present in 0.69% of patients with ASD and up to 2.12% of the cases with moderate to profound intellectual disability. In summary, mutations of the SHANK genes were detected in the whole spectrum of autism with a gradient of severity in cognitive impairment. Given the rare frequency of SHANK1 and SHANK2 deleterious mutations, the clinical relevance of these genes remains to be ascertained. In contrast, the frequency and the penetrance of SHANK3 mutations in individuals with ASD and intellectual disability-more than 1 in 50-warrant its consideration for mutation screening in clinical practice.

Background: The GGGGCC-repeat expansion in C9orf72 is the most frequent mutation found in patients with amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Most of the studies on C9orf72 have relied on repeat-primed PCR (RP-PCR) methods for detection of the expansions. To investigate the inherent limitations of this technique, we compared methods and results of 14 laboratories., Methods: The 14 laboratories genotyped DNA from 78 individuals (diagnosed with ALS or FTD) in a blinded fashion. Eleven laboratories used a combination of amplicon-length analysis and RP-PCR, whereas three laboratories used RP-PCR alone; Southern blotting techniques were used as a reference., Results: Using PCR-based techniques, 5 of the 14 laboratories got results in full accordance with the Southern blotting results. Only 50 of the 78 DNA samples got the same genotype result in all 14 laboratories. There was a high degree of false positive and false negative results, and at least one sample could not be genotyped at all in 9 of the 14 laboratories. The mean sensitivity of a combination of amplicon-length analysis and RP-PCR was 95.0% (73.9-100%), and the mean specificity was 98.0% (87.5-100%). Overall, a sensitivity and specificity of more than 95% was observed in only seven laboratories., Conclusions: Because of the wide range seen in genotyping results, we recommend using a combination of amplicon-length analysis and RP-PCR as a minimum in a research setting. We propose that Southern blotting techniques should be the gold standard, and be made obligatory in a clinical diagnostic setting., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.)

Background: Widespread abnormalities in white matter development are frequently reported in cases of autism spectrum disorders (ASD) and could be involved in the disconnectivity suggested in these disorders. Homozygous mutations in the gene coding for fatty-acid 2-hydroxylase (FA2H), an enzyme involved in myelin synthesis, are associated with complex leukodystrophies, but little is known about the functional impact of heterozygous FA2H mutations. We hypothesized that rare deleterious heterozygous mutations of FA2H might constitute risk factors for ASD., Methods: We searched deleterious mutations affecting FA2H, by genotyping 1256 independent patients with ASD genotyped using Genome Wide SNP arrays, and also by sequencing in independent set of 186 subjects with ASD and 353 controls. We then explored the impact of the identified mutations by measuring FA2H enzymatic activity and expression, in transfected COS7 cells., Results: One heterozygous deletion within 16q22.3-q23.1 including FA2H was observed in two siblings who share symptoms of autism and severe cognitive impairment, axial T2-FLAIR weighted MRI posterior periventricular white matter lesions. Also, two rare non-synonymous mutations (R113W and R113Q) were reported. Although predictive models suggested that R113W should be a deleterious, we did not find that FA2H activity was affected by expression of the R113W mutation in cultured COS cells., Conclusions: While our results do not support a major role for FA2H coding variants in ASD, a screening of other genes related to myelin synthesis would allow us to better understand the role of non-neuronal elements in ASD susceptibility.

Mouse ultrasonic vocalisations have been often used as a paradigm to extrapolate vocal communication defects observed in patients with autism spectrum disorders (ASD). The role of these vocalisations as well as their development, structure and informational content, however, remain largely unknown. In the present study, we characterised in depth the emission of pup and adult ultrasonic vocalisations of wild-type mice and their ProSAP1/Shank2(-/-) littermates lacking a synaptic scaffold protein mutated in ASD. We hypothesised that the vocal behaviour of ProSAP1/Shank2(-/-) mice not only differs from the vocal behaviour of their wild-type littermates in a quantitative way, but also presents more qualitative abnormalities in temporal organisation and acoustic structure. We first quantified the rate of emission of ultrasonic vocalisations, and analysed the organisation of vocalisations sequences using Markov models. We subsequently measured duration and peak frequency characteristics of each ultrasonic vocalisation, to characterise their acoustic structure. In wild-type mice, we found a high level of organisation in sequences of ultrasonic vocalisations, suggesting a communicative function in this complex system. Very limited significant sex-related variations were detected in their usage and acoustic structure, even in adult mice. In adult ProSAP1/Shank2(-/-) mice, we found abnormalities in the call usage and the structure of ultrasonic vocalisations. Both ProSAP1/Shank2(-/-) male and female mice uttered less vocalisations with a different call distribution and at lower peak frequency in comparison with wild-type littermates. This study provides a comprehensive framework to characterise abnormalities of ultrasonic vocalisations in mice and confirms that ProSAP1/Shank2(-/-) mice represent a relevant model to study communication defects., (Copyright © 2013 The Authors. Published by Elsevier B.V. All rights reserved.)

Mutations in NLGN4X have been identified in individuals with autism spectrum disorders and other neurodevelopmental disorders. A previous study reported that adult male mice lacking neuroligin4 (Nlgn4) displayed social approach deficits in the three-chambered test, altered aggressive behaviors and reduced ultrasonic vocalizations. To replicate and extend these findings, independent comprehensive analyses of autism-relevant behavioral phenotypes were conducted in later generations of the same line of Nlgn4 mutant mice at the National Institute of Mental Health in Bethesda, MD, USA and at the Institut Pasteur in Paris, France. Adult social approach was normal in all three genotypes of Nlgn4 mice tested at both sites. Reciprocal social interactions in juveniles were similarly normal across genotypes. No genotype differences were detected in ultrasonic vocalizations in pups separated from the nest or in adults during reciprocal social interactions. Anxiety-like behaviors, self-grooming, rotarod and open field exploration did not differ across genotypes, and measures of developmental milestones and general health were normal. Our findings indicate an absence of autism-relevant behavioral phenotypes in subsequent generations of Nlgn4 mice tested at two locations. Testing environment and methods differed from the original study in some aspects, although the presence of normal sociability was seen in all genotypes when methods taken from Jamain et al. (2008) were used. The divergent results obtained from this study indicate that phenotypes may not be replicable across breeding generations, and highlight the significant roles of environmental, generational and/or procedural factors on behavioral phenotypes., (Published 2012. This article is a U.S. Government work and is in the public domain in the USA.)

Recent studies have highlighted the involvement of rare (<1% frequency) copy-number variations and point mutations in the genetic etiology of autism spectrum disorder (ASD); these variants particularly affect genes involved in the neuronal synaptic complex. The SHANK gene family consists of three members (SHANK1, SHANK2, and SHANK3), which encode scaffolding proteins required for the proper formation and function of neuronal synapses. Although SHANK2 and SHANK3 mutations have been implicated in ASD and intellectual disability, the involvement of SHANK1 is unknown. Here, we assess microarray data from 1,158 Canadian and 456 European individuals with ASD to discover microdeletions at the SHANK1 locus on chromosome 19. We identify a hemizygous SHANK1 deletion that segregates in a four-generation family in which male carriers--but not female carriers--have ASD with higher functioning. A de novo SHANK1 deletion was also detected in an unrelated male individual with ASD with higher functioning, and no equivalent SHANK1 mutations were found in >15,000 controls (p = 0.009). The discovery of apparent reduced penetrance of ASD in females bearing inherited autosomal SHANK1 deletions provides a possible contributory model for the male gender bias in autism. The data are also informative for clinical-genetics interpretations of both inherited and sporadic forms of ASD involving SHANK1., (Copyright © 2012 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

Autism spectrum disorders comprise a range of neurodevelopmental disorders characterized by deficits in social interaction and communication, and by repetitive behaviour. Mutations in synaptic proteins such as neuroligins, neurexins, GKAPs/SAPAPs and ProSAPs/Shanks were identified in patients with autism spectrum disorder, but the causative mechanisms remain largely unknown. ProSAPs/Shanks build large homo- and heteromeric protein complexes at excitatory synapses and organize the complex protein machinery of the postsynaptic density in a laminar fashion. Here we demonstrate that genetic deletion of ProSAP1/Shank2 results in an early, brain-region-specific upregulation of ionotropic glutamate receptors at the synapse and increased levels of ProSAP2/Shank3. Moreover, ProSAP1/Shank2(-/-) mutants exhibit fewer dendritic spines and show reduced basal synaptic transmission, a reduced frequency of miniature excitatory postsynaptic currents and enhanced N-methyl-d-aspartate receptor-mediated excitatory currents at the physiological level. Mutants are extremely hyperactive and display profound autistic-like behavioural alterations including repetitive grooming as well as abnormalities in vocal and social behaviours. By comparing the data on ProSAP1/Shank2(-/-) mutants with ProSAP2/Shank3αβ(-/-) mice, we show that different abnormalities in synaptic glutamate receptor expression can cause alterations in social interactions and communication. Accordingly, we propose that appropriate therapies for autism spectrum disorders are to be carefully matched to the underlying synaptopathic phenotype.