A brain region-specific predictive gene map for autism derived by profiling a reference gene set

Abstract

Molecular underpinnings of complex psychiatric disorders such as autism spectrum disorders (ASD) remain largely unresolved. Increasingly, structural variations in discrete chromosomal loci are implicated in ASD, expanding the search space for its disease etiology. We exploited the high genetic heterogeneity of ASD to derive a predictive map of candidate genes by an integrated bioinformatics approach. Using a reference set of 84 Rare and Syndromic candidate ASD genes (AutRef84), we built a composite reference profile based on both functional and expression analyses. First, we created a functional profile of AutRef84 by performing Gene Ontology (GO) enrichment analysis which encompassed three main areas: 1) neurogenesis/projection, 2) cell adhesion, and 3) ion channel activity. Second, we constructed an expression profile of AutRef84 by conducting DAVID analysis which found enrichment in brain regions critical for sensory information processing (olfactory bulb, occipital lobe), executive function (prefrontal cortex), and hormone secretion (pituitary). Disease specificity of this dual AutRef84 profile was demonstrated by comparative analysis with control, diabetes, and non-specific gene sets. We then screened the human genome with the dual AutRef84 profile to derive a set of 460 potential ASD candidate genes. Importantly, the power of our predictive gene map was demonstrated by capturing 18 existing ASD-associated genes which were not part of the AutRef84 input dataset. The remaining 442 genes are entirely novel putative ASD risk genes. Together, we used a composite ASD reference profile to generate a predictive map of novel ASD candidate genes which should be prioritized for future research.

Figure 1. Integrated analysis of reference sets of genes Linked to ASD.

A reference dataset of ASD-linked genes (AutRef84) was assembled from the Rare and Syndromic categories of AutDB (http://www.mindspec.org/autdb.html), a publicly available portal for ongoing collection of genes linked to ASD. A) Distribution of genetic categories in AutDB. B) Reference sets were analyzed using structured biological knowledge provided by Gene Ontology (GO) consortium .

Figure 2. AutRef84 functional profile: graphical representation of over-represented Biological Process (BP) categories.

Using Bioconductor, we generated directed acyclic graphs based on GO knowledge structure. Enriched GO categories of AutRef84 are represented by rectangular boxes. Terminal nodes are illustrated in yellow. The largest structural component of the BP GO Tree is connected to neuron projection development, which includes the enriched GO categories of neuron differentiation (9 genes, P = 6.0×10⁻⁵), neurogenesis (10 genes, 4.0×10⁻⁵), and central nervous system development (9 genes, P = 4.0×10⁻⁵). Other enriched terminal nodes relate to ion channel activity or cell adhesion.

Figure 3. AutRef84 expression profile: region-specific enrichment of gene expression.

Analysis of tissue expression profiles for AutRef84 genes using the DAVID bioinformatics tool (http://david.abcc.ncifcrf.gov/) demonstrates region-specific enrichment with high statistical significance (p<0.0001) in four areas of the central nervous system: olfactory bulb, occipital lobe, prefrontal cortex, and pituitary. Whereas the olfactory bulb and occipital lobe are involved in sensory processing (smell and vision, respectively), the prefrontal cortex controls executive function and the pituitary gland directs hormone secretion. None of the enriched regions overlapped with those of diabetes or non-specific disease gene sets. * = Bonferroni corrected.

Figure 4. Network analysis of the AutRef84 expression profile.

In this visual representation of the network, each group of gene nodes is spatially positioned near the brain region or regions in which the genes are expressed. The color of each group of gene nodes was derived by averaging red, green, and blue values of the colors of the linked brain region nodes.

Figure 5. Genome-wide screening with the functional AutRef84 profile.

The functional profile of AutRef84 was used to predict ASD genes and map them to their appropriate location on the chromosome. To perform this data mining, we used the biomaRT package of Bioconductor from human genome at the Ensembl database (http://www.ensembl.org/Homo_sapiens) to create a graphical representation of chromosomal locations of genes matching with the functional AutRef84 profile. The complete list of 1185 matching genes is provided as Table S7. This map indicates uneven distribution with dense packing of matched genes in discrete chromosomal regions, 13 of which reached statistical significance (Table S8).

Figure 6. Network representation of ASD predictive gene map matching the dual profile of AutRef84.

After initially identifying 1185 genes matching the AutRef84 functional profile, we filtered this set by performing tissue-specific enrichment analysis and network representation of its shared brain regions within the AutRef84 expression profile. Using this method of dual profiling, we defined a prioritized subset of 460 genes predicted to be mutated in individuals with ASD. Within this subset, 159 genes are expressed in all four enriched brain regions of AutRef84, 62 genes were common to three of the enriched brain regions, 89 genes overlapped in two of the enriched regions, and 150 genes were expressed in only one enriched brain region (Table S9). Node placement and coloring were determined as described in Figure 4 .