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DNA methylation analysis of the autistic brain reveals multiple dysregulated biological pathways.

Nardone S, Sams DS, Reuveni E, Getselter D, Oron O, Karpuj M, Elliott E - Transl Psychiatry (2014)

Bottom Line: In BA10 we found a very significant enrichment for genomic areas responsible for immune functions among the hypomethylated CpGs, whereas genes related to synaptic membrane were enriched among hypermethylated CpGs.Finally, we determined the epigenetic dysregulation of the gene HDAC4, and we confirm that the locus encompassing C11orf21/TSPAN32 has multiple hypomethylated CpGs in the autistic brain, as previously demonstrated.Our data suggest a possible role for epigenetic processes in the etiology of ASD.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Medicine, Bar Ilan University, Safed, Israel.

ABSTRACT
Autism spectrum disorders (ASD) are a group of neurodevelopmental conditions characterized by dysfunction in social interaction, communication and stereotypic behavior. Genetic and environmental factors have been implicated in the development of ASD, but the molecular mechanisms underlying their interaction are not clear. Epigenetic modifications have been suggested as molecular mechanism that can mediate the interaction between the environment and the genome to produce adaptive or maladaptive behaviors. Here, using the Illumina 450 K methylation array we have determined the existence of many dysregulated CpGs in two cortical regions, Brodmann area 10 (BA10) and Brodmann area 24 (BA24), of individuals who had ASD. In BA10 we found a very significant enrichment for genomic areas responsible for immune functions among the hypomethylated CpGs, whereas genes related to synaptic membrane were enriched among hypermethylated CpGs. By comparing our methylome data with previously published transcriptome data, and by performing real-time PCR on selected genes that were dysregulated in our study, we show that hypomethylated genes are often overexpressed, and that there is an inverse correlation between gene expression and DNA methylation within the individuals. Among these genes there were C1Q, C3, ITGB2 (C3R), TNF-α, IRF8 and SPI1, which have recently been implicated in synaptic pruning and microglial cell specification. Finally, we determined the epigenetic dysregulation of the gene HDAC4, and we confirm that the locus encompassing C11orf21/TSPAN32 has multiple hypomethylated CpGs in the autistic brain, as previously demonstrated. Our data suggest a possible role for epigenetic processes in the etiology of ASD.

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Dysregulated DNA methylation in the genes HDAC4 and TSPAN32. (a) Gene expression and directional association with β methylation values are depicted for HDAC4. The position of the significant CpGs in relation to the gene, CpG islands and the Illumina 450 K methylation array are shown on the top of the three box plots. Gene expression (left) and inverse (center) and direct (right) correlations between gene expression and CpG methylation levels are shown below. P-values for both expression and Spearman's ρ correlation coefficient were <0.05. (b) Differences in methylation between autistic and control cohorts are described for C11orf21/TSPAN32. The position of significant CpGs in relation to the gene, CpG islands and the Illumina 450 K methylation array are shown above. A bar chart provides a detailed description of each single CpG site for C11orf21/TSPAN32 for both autistic and control cohorts (below) with relative P-value indicated above each pair of bars (*P<0.05; **P<0.01; ***P<0.001). For one CpG site (highlighted in red) is shown a box plot describing the β methylation value in autism and control cohorts, and a Spearman's ρ correlation coefficient between the methylation β values (microarray) and the pyrosequencing methylation values. P-values for both methylation and Spearman's ρ correlation coefficient were <0.05.
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fig4: Dysregulated DNA methylation in the genes HDAC4 and TSPAN32. (a) Gene expression and directional association with β methylation values are depicted for HDAC4. The position of the significant CpGs in relation to the gene, CpG islands and the Illumina 450 K methylation array are shown on the top of the three box plots. Gene expression (left) and inverse (center) and direct (right) correlations between gene expression and CpG methylation levels are shown below. P-values for both expression and Spearman's ρ correlation coefficient were <0.05. (b) Differences in methylation between autistic and control cohorts are described for C11orf21/TSPAN32. The position of significant CpGs in relation to the gene, CpG islands and the Illumina 450 K methylation array are shown above. A bar chart provides a detailed description of each single CpG site for C11orf21/TSPAN32 for both autistic and control cohorts (below) with relative P-value indicated above each pair of bars (*P<0.05; **P<0.01; ***P<0.001). For one CpG site (highlighted in red) is shown a box plot describing the β methylation value in autism and control cohorts, and a Spearman's ρ correlation coefficient between the methylation β values (microarray) and the pyrosequencing methylation values. P-values for both methylation and Spearman's ρ correlation coefficient were <0.05.

Mentions: Among genes with several dysregulated CpGs in the autistic BA10, there was HDAC4. HDAC4 is also an interesting gene in the context of autism because it is known to downregulate the transcription of several synaptic genes,29 and genetic aberrations in this gene cause brachydactyly-mental retardation syndrome, including autistic features.30 Interestingly, HDAC4 did not show a specific methylation trend, with some CpGs being hypomethylated whereas others hypermethylated. We found that HDAC4 was significantly overexpressed in autistic subjects, and that methylation of some CpGs had a positive while others a negative correlation to its gene expression (Figure 4a, Supplementary Table 11). Therefore, there is a complex interaction between DNA methylation and gene expression of HDAC4 in the autistic brain.


DNA methylation analysis of the autistic brain reveals multiple dysregulated biological pathways.

Nardone S, Sams DS, Reuveni E, Getselter D, Oron O, Karpuj M, Elliott E - Transl Psychiatry (2014)

Dysregulated DNA methylation in the genes HDAC4 and TSPAN32. (a) Gene expression and directional association with β methylation values are depicted for HDAC4. The position of the significant CpGs in relation to the gene, CpG islands and the Illumina 450 K methylation array are shown on the top of the three box plots. Gene expression (left) and inverse (center) and direct (right) correlations between gene expression and CpG methylation levels are shown below. P-values for both expression and Spearman's ρ correlation coefficient were <0.05. (b) Differences in methylation between autistic and control cohorts are described for C11orf21/TSPAN32. The position of significant CpGs in relation to the gene, CpG islands and the Illumina 450 K methylation array are shown above. A bar chart provides a detailed description of each single CpG site for C11orf21/TSPAN32 for both autistic and control cohorts (below) with relative P-value indicated above each pair of bars (*P<0.05; **P<0.01; ***P<0.001). For one CpG site (highlighted in red) is shown a box plot describing the β methylation value in autism and control cohorts, and a Spearman's ρ correlation coefficient between the methylation β values (microarray) and the pyrosequencing methylation values. P-values for both methylation and Spearman's ρ correlation coefficient were <0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4203003&req=5

fig4: Dysregulated DNA methylation in the genes HDAC4 and TSPAN32. (a) Gene expression and directional association with β methylation values are depicted for HDAC4. The position of the significant CpGs in relation to the gene, CpG islands and the Illumina 450 K methylation array are shown on the top of the three box plots. Gene expression (left) and inverse (center) and direct (right) correlations between gene expression and CpG methylation levels are shown below. P-values for both expression and Spearman's ρ correlation coefficient were <0.05. (b) Differences in methylation between autistic and control cohorts are described for C11orf21/TSPAN32. The position of significant CpGs in relation to the gene, CpG islands and the Illumina 450 K methylation array are shown above. A bar chart provides a detailed description of each single CpG site for C11orf21/TSPAN32 for both autistic and control cohorts (below) with relative P-value indicated above each pair of bars (*P<0.05; **P<0.01; ***P<0.001). For one CpG site (highlighted in red) is shown a box plot describing the β methylation value in autism and control cohorts, and a Spearman's ρ correlation coefficient between the methylation β values (microarray) and the pyrosequencing methylation values. P-values for both methylation and Spearman's ρ correlation coefficient were <0.05.
Mentions: Among genes with several dysregulated CpGs in the autistic BA10, there was HDAC4. HDAC4 is also an interesting gene in the context of autism because it is known to downregulate the transcription of several synaptic genes,29 and genetic aberrations in this gene cause brachydactyly-mental retardation syndrome, including autistic features.30 Interestingly, HDAC4 did not show a specific methylation trend, with some CpGs being hypomethylated whereas others hypermethylated. We found that HDAC4 was significantly overexpressed in autistic subjects, and that methylation of some CpGs had a positive while others a negative correlation to its gene expression (Figure 4a, Supplementary Table 11). Therefore, there is a complex interaction between DNA methylation and gene expression of HDAC4 in the autistic brain.

Bottom Line: In BA10 we found a very significant enrichment for genomic areas responsible for immune functions among the hypomethylated CpGs, whereas genes related to synaptic membrane were enriched among hypermethylated CpGs.Finally, we determined the epigenetic dysregulation of the gene HDAC4, and we confirm that the locus encompassing C11orf21/TSPAN32 has multiple hypomethylated CpGs in the autistic brain, as previously demonstrated.Our data suggest a possible role for epigenetic processes in the etiology of ASD.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Medicine, Bar Ilan University, Safed, Israel.

ABSTRACT
Autism spectrum disorders (ASD) are a group of neurodevelopmental conditions characterized by dysfunction in social interaction, communication and stereotypic behavior. Genetic and environmental factors have been implicated in the development of ASD, but the molecular mechanisms underlying their interaction are not clear. Epigenetic modifications have been suggested as molecular mechanism that can mediate the interaction between the environment and the genome to produce adaptive or maladaptive behaviors. Here, using the Illumina 450 K methylation array we have determined the existence of many dysregulated CpGs in two cortical regions, Brodmann area 10 (BA10) and Brodmann area 24 (BA24), of individuals who had ASD. In BA10 we found a very significant enrichment for genomic areas responsible for immune functions among the hypomethylated CpGs, whereas genes related to synaptic membrane were enriched among hypermethylated CpGs. By comparing our methylome data with previously published transcriptome data, and by performing real-time PCR on selected genes that were dysregulated in our study, we show that hypomethylated genes are often overexpressed, and that there is an inverse correlation between gene expression and DNA methylation within the individuals. Among these genes there were C1Q, C3, ITGB2 (C3R), TNF-α, IRF8 and SPI1, which have recently been implicated in synaptic pruning and microglial cell specification. Finally, we determined the epigenetic dysregulation of the gene HDAC4, and we confirm that the locus encompassing C11orf21/TSPAN32 has multiple hypomethylated CpGs in the autistic brain, as previously demonstrated. Our data suggest a possible role for epigenetic processes in the etiology of ASD.

Show MeSH
Related in: MedlinePlus