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. 2017 May 31;12(5):e0178532.
doi: 10.1371/journal.pone.0178532. eCollection 2017.

Integrative genomic analyses for identification and prioritization of long non-coding RNAs associated with autism

Affiliations

Integrative genomic analyses for identification and prioritization of long non-coding RNAs associated with autism

Brian L Gudenas et al. PLoS One. .

Abstract

Genetic studies have identified many risk loci for autism spectrum disorder (ASD) although causal factors in the majority of cases are still unknown. Currently, known ASD risk genes are all protein-coding genes; however, the vast majority of transcripts in humans are non-coding RNAs (ncRNAs) which do not encode proteins. Recently, long non-coding RNAs (lncRNAs) were shown to be highly expressed in the human brain and crucial for normal brain development. We have constructed a computational pipeline for the integration of various genomic datasets to identify lncRNAs associated with ASD. This pipeline utilizes differential gene expression patterns in affected tissues in conjunction with gene co-expression networks in tissue-matched non-affected samples. We analyzed RNA-seq data from the cortical brain tissues from ASD cases and controls to identify lncRNAs differentially expressed in ASD. We derived a gene co-expression network from an independent human brain developmental transcriptome and detected a convergence of the differentially expressed lncRNAs and known ASD risk genes into specific co-expression modules. Co-expression network analysis facilitates the discovery of associations between previously uncharacterized lncRNAs with known ASD risk genes, affected molecular pathways and at-risk developmental time points. In addition, we show that some of these lncRNAs have a high degree of overlap with major CNVs detected in ASD genetic studies. By utilizing this integrative approach comprised of differential expression analysis in affected tissues and connectivity metrics from a developmental co-expression network, we have prioritized a set of candidate ASD-associated lncRNAs. The identification of lncRNAs as novel ASD susceptibility genes could help explain the genetic pathogenesis of ASD.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Fig 1
Fig 1. Differentially expressed genes in the ASD cortex.
The volcano plot displays genes differentially expressed in the ASD cortex. A gene was required to have an absolute value of log2 fold change greater than or equal to one and an adjusted p-value less than 0.05 to be considered differentially expressed.
Fig 2
Fig 2. Enrichment of lncRNAs and ASD genes in brain developmental co-expression modules.
The heatmap shows module-based enrichment of gene lists. “DE LncRNAs” are lncRNAs differentially expressed in the ASD cortex, “SFARI ASD” are known ASD risk genes, and “ME16” is an ASD-associated gene co-expression module identified in an independent study. Enrichment of gene lists was determined by a Fischer’s exact test requiring the FDR-adjusted p-value < 0.05 and an Odds Ratio > 1. Only modules containing at least 1 differentially expressed lncRNAs are shown.
Fig 3
Fig 3. Differential expression in the ASD cortex overlaid onto developmental co-expression modules.
The average log2 fold changes of genes differentially expressed in the ASD cortex were overlaid onto the co-expression modules formed using the BrainSpan Developmental Transcriptome. Any genes that failed to reach significance had their log2 fold changes set to 0. The red circle within each bar plot is the average log2 fold change of 10,000 random gene samplings of equal size to the respective module. Significance of differential expression compared to the permuted distribution (FDR-adjusted p-value < 0.05) is denoted by a black asterisk adjacent to a modules respective bar plot.
Fig 4
Fig 4. Characterization of modules enriched for differentially expressed lncRNAs.
Gene Ontology functional enrichment was analyzed for each module and adjusted for multiple comparisons (FDR < 0.05). The scatterplots show modular developmental expression profiles based on a module eigengene (1st principal component) through developmental time, months PC means months post-conception (2 months post-conception to 1 post-natal year), with the blue vertical line demarcating birth. The trend line of each scatterplot is derived from a locally weighted scatterplot smoothing function.
Fig 5
Fig 5. Candidate ASD-associated lncRNA characteristics.
(A) Module assignment for differentially expressed lncRNAs. Only modules with at least 4 lncRNAs are displayed. In addition, we provide the module function, which is the highest scoring GO biological process for the whole module. (B) Average log2 fold change of expression in the ASD cortex for the differentially expressed lncRNAs from each module. (C) Average fractional expression levels in the brain for the differentially expressed lncRNAs from each module. Fractional brain expression for each lncRNA is calculated using the RNA-seq data from the Genotype-Tissue Expression project as the total expression in brain tissues divided by the sum of expression across all tissue types [28]. The red line at 50% represents the threshold for tissue specificity as defined by Ayupe et al [36]. (D) Overlaps between ASD CNVs and DE lncRNAs from each module.

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This work was supported by a grant from the Self Regional Healthcare Foundation.

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