Multiplex Autism Families Research Articles

Identifying rare genetic variants in 21 highly multiplex autism families: the role of diagnosis and autistic traits

Autism is a highly heritable, heterogeneous, neurodevelopmental condition. Large-scale genetic studies, predominantly focussing on simplex families and clinical diagnoses of autism have identified hundreds of genes associated with autism. Yet, the contribution of these classes of genes to multiplex families and autistic traits still warrants investigation. Here, we conducted whole-genome sequencing of 21 highly multiplex autism families, with at least three autistic individuals in each family, to prioritise genes associated with autism. Using a combination of both autistic traits and clinical diagnosis of autism, we identify rare variants in genes associated with autism, and related neurodevelopmental conditions in multiple families. We identify a modest excess of these variants in autistic individuals compared to individuals without an autism diagnosis. Finally, we identify a convergence of the genes identified in molecular pathways related to development and neurogenesis. In sum, our analysis provides initial evidence to demonstrate the value of integrating autism diagnosis and autistic traits to prioritise genes.

Integrative Functional Genomic Analysis in Multiplex Autism Families from Kazakhstan.

The study of extended pedigrees containing autism spectrum disorder- (ASD-) related broader autism phenotypes (BAP) offers a promising approach to the search for ASD candidate variants. Here, a total of 650,000 genetic markers were tested in four Kazakhstani multiplex families with ASD and BAP to obtain data on de novo mutations (DNMs), common, and rare inherited variants that may contribute to the genetic risk for developing autistic traits. The variants were analyzed in the context of gene networks and pathways. Several previously well-described enriched pathways were identified, including ion channel activity, regulation of synaptic function, and membrane depolarization. Perhaps these pathways are crucial not only for the development of ASD but also for ВАР. The results also point to several additional biological pathways (circadian entrainment, NCAM and BTN family interactions, and interaction between L1 and Ankyrins) and hub genes (CFTR, NOD2, PPP2R2B, and TTR). The obtained results suggest that further exploration of PPI networks combining ASD and BAP risk genes can be used to identify novel or overlooked ASD molecular mechanisms.

Game theoretic centrality: a novel approach to prioritize disease candidate genes by combining biological networks with the Shapley value

BackgroundComplex human health conditions with etiological heterogeneity like Autism Spectrum Disorder (ASD) often pose a challenge for traditional genome-wide association study approaches in defining a clear genotype to phenotype model. Coalitional game theory (CGT) is an exciting method that can consider the combinatorial effect of groups of variants working in concert to produce a phenotype. CGT has been applied to associate likely-gene-disrupting variants encoded from whole genome sequence data to ASD; however, this previous approach cannot take into account for prior biological knowledge. Here we extend CGT to incorporate a priori knowledge from biological networks through a game theoretic centrality measure based on Shapley value to rank genes by their relevance–the individual gene’s synergistic influence in a gene-to-gene interaction network. Game theoretic centrality extends the notion of Shapley value to the evaluation of a gene’s contribution to the overall connectivity of its corresponding node in a biological network.ResultsWe implemented and applied game theoretic centrality to rank genes on whole genomes from 756 multiplex autism families. Top ranking genes with the highest game theoretic centrality in both the weighted and unweighted approaches were enriched for pathways previously associated with autism, including pathways of the immune system. Four of the selected genes HLA-A, HLA-B, HLA-G, and HLA-DRB1–have also been implicated in ASD and further support the link between ASD and the human leukocyte antigen complex.ConclusionsGame theoretic centrality can prioritize influential, disease-associated genes within biological networks, and assist in the decoding of polygenic associations to complex disorders like autism.

Cellular and molecular characterization of multiplex autism in human induced pluripotent stem cell-derived neurons

BackgroundAutism spectrum disorder (ASD) is a neurodevelopmental disorder with pronounced heritability in the general population. This is largely attributable to the effects of polygenic susceptibility, with inherited liability exhibiting distinct sex differences in phenotypic expression. Attempts to model ASD in human cellular systems have principally involved rare de novo mutations associated with ASD phenocopies. However, by definition, these models are not representative of polygenic liability, which accounts for the vast share of population-attributable risk.MethodsHere, we performed what is, to our knowledge, the first attempt to model multiplex autism using patient-derived induced pluripotent stem cells (iPSCs) in a family manifesting incremental degrees of phenotypic expression of inherited liability (absent, intermediate, severe). The family members share an inherited variant of uncertain significance (VUS) in GPD2, a gene that was previously associated with developmental disability but here is insufficient by itself to cause ASD. iPSCs from three first-degree relatives and an unrelated control were differentiated into both cortical excitatory (cExN) and cortical inhibitory (cIN) neurons, and cellular phenotyping and transcriptomic analysis were conducted.ResultscExN neurospheres from the two affected individuals were reduced in size, compared to those derived from unaffected related and unrelated individuals. This reduction was, at least in part, due to increased apoptosis of cells from affected individuals upon initiation of cExN neural induction. Likewise, cIN neural progenitor cells from affected individuals exhibited increased apoptosis, compared to both unaffected individuals. Transcriptomic analysis of both cExN and cIN neural progenitor cells revealed distinct molecular signatures associated with affectation, including the misregulation of suites of genes associated with neural development, neuronal function, and behavior, as well as altered expression of ASD risk-associated genes.ConclusionsWe have provided evidence of morphological, physiological, and transcriptomic signatures of polygenic liability to ASD from an analysis of cellular models derived from a multiplex autism family. ASD is commonly inherited on the basis of additive genetic liability. Therefore, identifying convergent cellular and molecular phenotypes resulting from polygenic and monogenic susceptibility may provide a critical bridge for determining which of the disparate effects of rare highly deleterious mutations might also apply to common autistic syndromes.

Affected Sib-Pair Analyses Identify Signaling Networks Associated With Social Behavioral Deficits in Autism.

Autism spectrum disorders (ASDs) are characterized by deficits in three core behavioral domains: reciprocal social interactions, communication, and restricted interests and/or repetitive behaviors. Several hundreds of risk genes for autism have been identified, however, it remains a challenge to associate these genes with specific core behavioral deficits. In multiplex autism families, affected sibs often show significant differences in severity of individual core phenotypes. We hypothesize that a higher mutation burden contributes to a larger difference in the severity of specific core phenotypes between affected sibs. We tested this hypothesis on social behavioral deficits in autism. We sequenced synaptome genes (n = 1,886) in affected male sib-pairs (n = 274) in families from the Autism Genetics Research Exchange (AGRE) and identified rare (MAF ≤ 1%) and predicted functional variants. We selected affected sib-pairs with a large (≥10; n = 92 pairs) or a small (≤4; n = 108 pairs) difference in total cumulative Autism Diagnostic Interview-Revised (ADI-R) social scores (SOCT_CS). We compared burdens of unshared variants present only in sibs with severe social deficits and found a higher burden in SOCT_CS≥10 compared to SOCT_CS ≤ 4 (SOCT_CS≥10: 705.1 ± 16.2; SOCT_CS ≤ 4, 668.3 ± 9.0; p = 0.025). Unshared SOCT_CS≥10 genes only in sibs with severe social deficits are significantly enriched in the SFARI gene set. Network analyses of these genes using InWeb_IM, molecular signatures database (MSigDB), and GeNetMeta identified enrichment for phosphoinositide 3-kinase (PI3K)-AKT-mammalian target of rapamycin (mTOR) (Enrichment Score [eScore] p value = 3.36E−07; n = 8 genes) and Nerve growth factor (NGF) (eScore p value = 8.94E−07; n = 9 genes) networks. These studies support a key role for these signaling networks in social behavioral deficits and present a novel approach to associate risk genes and signaling networks with core behavioral domains in autism.

Social responsiveness, an autism endophenotype: genomewide significant linkage to two regions on chromosome 8.

Autism spectrum disorder is characterized by deficits in social function and the presence of repetitive and restrictive behaviors. Following a previous test of principle, the authors adopted a quantitative approach to discovering genes contributing to the broader autism phenotype by using social responsiveness as an endophenotype for autism spectrum disorder. Linkage analyses using scores from the Social Responsiveness Scale were performed in 590 families from the Autism Genetic Resource Exchange, a largely multiplex autism spectrum disorder cohort. Regional and genomewide association analyses were performed to search for common variants contributing to social responsiveness. Social Responsiveness Scale scores were unimodally distributed in male offspring from multiplex autism families, in contrast with a bimodal distribution observed in female offspring. In correlated analyses differing by Social Responsiveness Scale respondent, genomewide significant linkage for social responsiveness was identified at chr8p21.3 (multipoint LOD=4.11; teacher/parent scores) and chr8q24.22 (multipoint LOD=4.54; parent-only scores), respectively. Genomewide or linkage-directed association analyses did not detect common variants contributing to social responsiveness. The sex-differential distributions of Social Responsiveness Scale scores in multiplex autism families likely reflect mechanisms contributing to the sex ratio for autism observed in the general population and form a quantitative signature of reduced penetrance of inherited liability to autism spectrum disorder among females. The identification of two strong loci for social responsiveness validates the endophenotype approach for the identification of genetic variants contributing to complex traits such as autism spectrum disorder. While causal mutations have yet to be identified, these findings are consistent with segregation of rare genetic variants influencing social responsiveness and underscore the increasingly recognized role of rare inherited variants in the genetic architecture of autism spectrum disorder.

New interview and observation measures of the broader autism phenotype: group differentiation.

To identify the broader autism phenotype (BAP), the Family History Interview subject and informant versions and an observational tool (Impression of Interviewee), were developed. This study investigated whether the instruments differentiated between parents of children with autism, and parents of children with Down syndrome (DS). The BAP scores of parents of 28 multiplex autism families were compared with parents from, 32 DS families. The BAP measures provided good group differentiation but when considered together, the subject interview did not improve group differentiation. The differentiation was better for fathers than mothers. The measures do carry an important degree of validity; whether they can differentiate the BAP from other social disorders should be tested.

Transcriptional Consequences of 16p11.2 Deletion and Duplication in Mouse Cortex and Multiplex Autism Families

Reciprocal copy-number variation (CNV) of a 593 kb region of 16p11.2 is a common genetic cause of autism spectrum disorder (ASD), yet it is not completely penetrant and can manifest in a wide array of phenotypes. To explore its molecular consequences, we performed RNA sequencing of cerebral cortex from mouse models with CNV of the syntenic 7qF3 region and lymphoblast lines from 34 members of 7 multiplex ASD-affected families harboring the 16p11.2 CNV. Expression of all genes in the CNV region correlated well with their DNA copy number, with no evidence of dosage compensation. We observed effects on gene expression outside the CNV region, including apparent positional effects in cis and in trans at genomic segments with evidence of physical interaction in Hi-C chromosome conformation data. One of the most significant positional effects was telomeric to the 16p11.2 CNV and includes the previously described "distal" 16p11.2 microdeletion. Overall, 16p11.2 CNV was associated with altered expression of genes and networks that converge on multiple hypotheses of ASD pathogenesis, including synaptic function (e.g., NRXN1, NRXN3), chromatin modification (e.g., CHD8, EHMT1, MECP2), transcriptional regulation (e.g., TCF4, SATB2), and intellectual disability (e.g., FMR1, CEP290). However, there were differences between tissues and species, with the strongest effects being consistently within the CNV region itself. Our analyses suggest that through a combination of indirect regulatory effects and direct effects on nuclear architecture, alteration of 16p11.2 genes disrupts expression networks that involve other genes and pathways known to contribute to ASD, suggesting an overlap in mechanisms of pathogenesis.

Does epilepsy in multiplex autism pedigrees define a different subgroup in terms of clinical characteristics and genetic risk?

BackgroundAutism spectrum disorders (ASD) and epilepsy frequently occur together. Prevalence rates are variable, and have been attributed to age, gender, comorbidity, subtype of pervasive developmental disorder (PDD) and risk factors. Recent studies have suggested disparate clinical and genetic settings depending on simplex or multiplex autism. The aim of this study was to assess: 1) the prevalence of epilepsy in multiplex autism and its association with genetic and non-genetic risk factors of major effect, intellectual disability and gender; and 2) whether autism and epilepsy cosegregate within multiplex autism families.MethodsWe extracted from the Autism Genetic Resource Exchange (AGRE) database (n = 3,818 children from 1,264 families) all families with relevant medical data (n = 664 children from 290 families). The sample included 478 children with ASD and 186 siblings without ASD. We analyzed the following variables: seizures, genetic and non-genetic risk factors, gender, and cognitive functioning as assessed by Raven’s Colored Progressive Matrices (RCPM) and Vineland Adaptive Behavior Scales (VABS).ResultsThe prevalence of epilepsy was 12.8% in cases with ASD and 2.2% in siblings without ASD (P <10-5). With each RCPM or VABS measure, the risk of epilepsy in multiplex autism was significantly associated with intellectual disability, but not with gender. Identified risk factors (genetic or non-genetic) of autism tended to be significantly associated with epilepsy (P = 0.052). When children with prematurity, pre- or perinatal insult, or cerebral palsy were excluded, a genetic risk factor was reported for 6/59 (10.2%) of children with epilepsy and 12/395 (3.0%) of children without epilepsy (P = 0.002). Finally, using a permutation test, there was significant evidence that the epilepsy phenotype co-segregated within families (P <10-4).ConclusionsEpilepsy in multiplex autism may define a different subgroup in terms of clinical characteristics and genetic risk.

Exome sequencing in multiplex autism families suggests a major role for heterozygous truncating mutations

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.

The Effects of Birth Order and Birth Interval on the Phenotypic Expression of Autism Spectrum Disorder

A rise in the prevalence of diagnosed cases of autism spectrum disorder (ASD) has been reported in several studies in recent years. While this rise in ASD prevalence is at least partially related to increased awareness and broadened diagnostic criteria, the role of environmental factors cannot be ruled out, especially considering that the cause of most cases of ASD remains unknown. The study of families with multiple affected children can provide clues about ASD etiology. While the majority of research on ASD multiplex families has focused on identifying genetic anomalies that may underlie the disorder, the study of symptom severity across ASD birth order may provide evidence for environmental factors in ASD. We compared social and cognitive measures of behavior between over 300 first and second affected siblings within multiplex autism families obtained from the Autism Genetic Resource Exchange dataset. Measures included nonverbal IQ assessed with the Ravens Colored Progressive Matrices, verbal IQ assessed with the Peabody Picture Vocabulary Test, and autism severity assessed with the Social Responsiveness Scale (SRS), an instrument established as a quantitative measure of autism. The results indicated that females were more severely impacted by ASD than males, especially first affected siblings. When first and second affected siblings were compared, significant declines in nonverbal and verbal IQ scores were observed. In addition, SRS results demonstrated a significant increase in autism severity between first and second affected siblings consistent with an overall decline in function as indicated by the IQ data. These results remained significant after controlling for the age and sex of the siblings. Surprisingly, the SRS scores were found to only be significant when the age difference between siblings was less than 2 years. These results suggest that some cases of ASD are influenced by a dosage effect involving unknown epigenetic, environmental, and/or immunological factors.

A common X-linked inborn error of carnitine biosynthesis may be a risk factor for nondysmorphic autism.

We recently reported a deletion of exon 2 of the trimethyllysine hydroxylase epsilon (TMLHE) gene in a proband with autism. TMLHE maps to the X chromosome and encodes the first enzyme in carnitine biosynthesis, 6-N-trimethyllysine dioxygenase. Deletion of exon 2 of TMLHE causes enzyme deficiency, resulting in increased substrate concentration (6-N-trimethyllysine) and decreased product levels (3-hydroxy-6-N-trimethyllysine and γ-butyrobetaine) in plasma and urine. TMLHE deficiency is common in control males (24 in 8,787 or 1 in 366) and was not significantly increased in frequency in probands from simplex autism families (9 in 2,904 or 1 in 323). However, it was 2.82-fold more frequent in probands from male-male multiplex autism families compared with controls (7 in 909 or 1 in 130; P = 0.023). Additionally, six of seven autistic male siblings of probands in male-male multiplex families had the deletion, suggesting that TMLHE deficiency is a risk factor for autism (metaanalysis Z-score = 2.90 and P = 0.0037), although with low penetrance (2-4%). These data suggest that dysregulation of carnitine metabolism may be important in nondysmorphic autism; that abnormalities of carnitine intake, loss, transport, or synthesis may be important in a larger fraction of nondysmorphic autism cases; and that the carnitine pathway may provide a novel target for therapy or prevention of autism.

Association of GTF2i in the Williams-Beuren Syndrome Critical Region with Autism Spectrum Disorders

Duplications of 7q11.23, deleted in Williams-Beuren Syndrome, have been implicated in autism spectrum disorders (ASDs). A 1.5 Mb duplication was identified in one girl with severe expressive language deficits and anxiety among 1,142 ASD individuals screened for this duplication. Family-based association studies of Tag-SNPs in three genes (STX1A , CYLN2 and GTF2i) in two multiplex autism family cohorts revealed strong association of two GTF2i SNPs and their haplotype in Cohort 1 and the combined families. The risk alleles and haplotype were associated with severe problems in social interaction and excessive repetitive behaviors. Our findings suggest the GTF2i gene is important in the etiology of autism in individuals with this duplication and in non-duplication cases with severe social interaction problems and repetitive behaviors.

Microduplications in an autism multiplex family narrow the region of susceptibility for developmental disorders on 15q24 and implicate 7p21

Copy number variations (CNVs) play a crucial role in the intricate genetics of autism spectrum disorders. A region on chromosome 15q24 vulnerable to both deletions and duplications has been previously implicated in a range of phenotypes including autism, Asperger's syndrome, delayed development, and mild to severe mental retardation. Prior studies have delineated a minimal critical region of approximately 1.33 Mb. In this study, a multiplex autism family was evaluated for CNVs using genotyping data from the Illumina 1 M BeadChip and analyzed with the PennCNV algorithm. Variants were then identified that co-segregate with autism features in this family. Here, we report autistic first cousins who carry two microduplications concordant with disease. Both duplications were inherited maternally and found to be identical by descent. The first is an approximately 10,000 base pair microduplication within the minimal region on 15q24 that falls across a single gene, ubiquitin-like 7. This is the smallest duplication in the region to result in a neuropsychiatric disorder, potentially narrowing the critical region for susceptibility to developmental and autism spectrum disorders. The second is a novel, 352 kb tandem duplication on 7p21 that replicates part of the neurexophilin 1 and islet cell autoantigen 1 genes. The breakpoint junction falls within the intronic regions of these genes and demonstrates a microhomology of four base pairs. Each of these microduplications may contribute to the complex etiology of autism spectrum disorders.

A family with autism and rare copy number variants disrupting the Duchenne/Becker muscular dystrophy gene DMD and TRPM3

Autism spectrum disorder is a genetically complex and clinically heterogeneous neurodevelopmental disorder. A recent study by the Autism Genome Project (AGP) used 1M single-nucleotide polymorphism arrays to show that rare genic copy number variants (CNVs), possibly acting in tandem, play a significant role in the genetic aetiology of this condition. In this study, we describe the phenotypic and genomic characterisation of a multiplex autism family from the AGP study that was found to harbour a duplication of exons 31–44 of the Duchenne/Becker muscular dystrophy gene DMD and also a rare deletion involving exons 1–9 of TRPM3. Further characterisation of these extremely rare CNVs was carried out using quantitative PCR, fluorescent in situ hybridisation, long-range PCR amplification and sequencing of junction fragments. The maternal chrX:32,097,213-32,321,945 tandem duplication and paternal chr9:72,480,413-73,064,196 deletion (NCBI build 36 coordinates) were transmitted to both affected boys, potentially signifying a multi-hit mechanism. The DMD reading frame rule predicts a Becker phenotype, characterised by later onset and milder symptoms. When last evaluated, neither child had developed signs of muscular dystrophy. These data are consistent with a degree of comorbidity between autism and muscular dystrophy and suggest that genomic background as well as the position of the mutation within the DMD gene may impact on the neurological correlates of Duchenne/Becker muscular dystrophy. Finally, communicating unexpected findings such as these back to families raises a number of ethical questions, which are discussed.Electronic supplementary materialThe online version of this article (doi:10.1007/s11689-011-9076-5) contains supplementary material, which is available to authorized users.

Allowing for sex differences increases power in a GWAS of multiplex Autism families

Current genomewide association studies account for only a small fraction of the estimated heritabilities of genetically complex neuropsychiatric disorders, indicating they are likely to result from the small effects of numerous predisposing variants, many of which have gone undetected. The statistical power to detect associations of common variants with small effects is increased by conducting joint association tests of a single nucleotide polymorphism (SNP), an additional risk factor (F), and their interaction. F can represent an environmental exposure, another genotype or any source of genetic heterogeneity. In case and control studies, logistic regression makes joint tests straightforward. This analytic method cannot be employed directly when SNP transmission tests are used to detect associations in parent/affected child trios and multiplex families. However, the method can be implemented using the case/pseudocontrol approach. We applied this approach to analyze data from a genomewide association study of multiplex families ascertained for Autism Spectrum Disorder, where sex was used to define the F. Joint analyses revealed two associations exceeding genomewide significance. One novel gene, Ryandine Receptor 2, implicated in calcium channel defects, was identified with a joint P-value of 3.9E-11. Calcium channel defects have been connected to Autism spectrum disorder (ASD) by Timothy Syndrome, which is Mendelian, and a previous targeted sex-specific association analysis of idiopathic Autism. A second gene, uridine phosphorylase 2, with a joint P-value of 2.3E-9, has been previously linked and associated with Autism in independent samples. These findings highlight two Autism candidate genes for follow-up studies.

Autistic traits in simplex and multiplex autism families: Focus on unaffected relatives

Autistic traits in simplex and multiplex autism families: Focus on unaffected relatives

Genomic and epigenetic evidence for oxytocin receptor deficiency in autism

BackgroundAutism comprises a spectrum of behavioral and cognitive disturbances of childhood development and is known to be highly heritable. Although numerous approaches have been used to identify genes implicated in the development of autism, less than 10% of autism cases have been attributed to single gene disorders.MethodsWe describe the use of high-resolution genome-wide tilepath microarrays and comparative genomic hybridization to identify copy number variants within 119 probands from multiplex autism families. We next carried out DNA methylation analysis by bisulfite sequencing in a proband and his family, expanding this analysis to methylation analysis of peripheral blood and temporal cortex DNA of autism cases and matched controls from independent datasets. We also assessed oxytocin receptor (OXTR) gene expression within the temporal cortex tissue by quantitative real-time polymerase chain reaction (PCR).ResultsOur analysis revealed a genomic deletion containing the oxytocin receptor gene, OXTR (MIM accession no.: 167055), previously implicated in autism, was present in an autism proband and his mother who exhibits symptoms of obsessive-compulsive disorder. The proband's affected sibling did not harbor this deletion but instead may exhibit epigenetic misregulation of this gene through aberrant gene silencing by DNA methylation. Further DNA methylation analysis of the CpG island known to regulate OXTR expression identified several CpG dinucleotides that show independent statistically significant increases in the DNA methylation status in the peripheral blood cells and temporal cortex in independent datasets of individuals with autism as compared to control samples. Associated with the increase in methylation of these CpG dinucleotides is our finding that OXTR mRNA showed decreased expression in the temporal cortex tissue of autism cases matched for age and sex compared to controls.ConclusionTogether, these data provide further evidence for the role of OXTR and the oxytocin signaling pathway in the etiology of autism and, for the first time, implicate the epigenetic regulation of OXTR in the development of the disorder.See the related commentary by Gurrieri and Neri:

A genome-wide linkage and association scan reveals novel loci for autism

SummaryAlthough autism is a highly heritable neurodevelopmental disorder, attempts to identify specific susceptibility genes have thus far met with limited success 1. Genome-wide association studies (GWAS) using half a million or more markers, particularly those with very large sample sizes achieved through meta-analysis, have shown great success in mapping genes for other complex genetic traits (http://www.genome.gov/26525384). Consequently, we initiated a linkage and association mapping study using half a million genome-wide SNPs in a common set of 1,031 multiplex autism families (1,553 affected offspring). We identified regions of suggestive and significant linkage on chromosomes 6q27 and 20p13, respectively. Initial analysis did not yield genome-wide significant associations; however, genotyping of top hits in additional families revealed a SNP on chromosome 5p15 (between SEMA5A and TAS2R1) that was significantly associated with autism (P = 2 × 10−7). We also demonstrated that expression of SEMA5A is reduced in brains from autistic patients, further implicating SEMA5A as an autism susceptibility gene. The linkage regions reported here provide targets for rare variation screening while the discovery of a single novel association demonstrates the action of common variants.

Locations and patterns of meiotic recombination in two-generation pedigrees

BackgroundMeiotic crossovers are the major mechanism by which haplotypes are shuffled to generate genetic diversity. Previously available methods for the genome-wide, high-resolution identification of meiotic crossover sites are limited by the laborious nature of the assay (as in sperm typing).MethodsSeveral methods have been introduced to identify crossovers using high density single nucleotide polymorphism (SNP) array technologies, although programs are not widely available to implement such analyses.ResultsHere we present a two-generation "reverse pedigree analysis" method (analyzing the genotypes of two children relative to each parent) and a web-accessible tool to determine and visualize inheritance differences among siblings and crossover locations on each parental gamete. This approach is complementary to existing methods and uses informative markers which provide high resolution for locating meiotic crossover sites. We introduce a segmentation algorithm to identify crossover sites, and used a synthetic data set to determine that the segmentation algorithm specificity was 92% and sensitivity was 89%. The use of reverse pedigrees allows the inference of crossover locations on the X chromosome in a maternal gamete through analysis of two sons and their father. We further analyzed genotypes from eight multiplex autism families, observing a 1.462 maternal to paternal recombination ratio and no significant differences between affected and unaffected children. Meiotic recombination results from pediSNP can also be used to identify haplotypes that are shared by probands within a pedigree, as we demonstrated with a multiplex autism family.ConclusionUsing "reverse pedigrees" and defining unique sets of genotype markers within pedigree data, we introduce a method that identifies inherited allelic differences and meiotic crossovers. We implemented the method in the pediSNP software program, and we applied it to several data sets. This approach uses data from two generations to identify crossover sites, facilitating studies of recombination in disease. pediSNP is available online at .