Investigation of epigenetic regulatory networks associated with autism spectrum disorder (ASD) by integrated global LINE-1 methylation and gene expression profiling analyses.

Chayanin Tangsuwansri,Surangrat Thongkorn,Kanya Suphapeetiporn,Apiwat Mutirangura,Valerie Wailin Hu,Tewarit Sarachana,Thanit Saeliw,Tewin Tencomnao,Weerasak Chonchaiya,Kenji Hashimoto

doi:10.1371/journal.pone.0201071

Abstract

BackgroundThe exact cause and mechanisms underlying the pathobiology of autism spectrum disorder (ASD) remain unclear. Dysregulation of long interspersed element-1 (LINE-1) has been reported in the brains of ASD-like mutant mice and ASD brain tissues. However, the role and methylation of LINE-1 in individuals with ASD remain unclear. In this study, we aimed to investigate whether LINE-1 insertion is associated with differentially expressed genes (DEGs) and to assess LINE-1 methylation in ASD.MethodsTo identify DEGs associated with LINE-1 in ASD, we reanalyzed previously published transcriptome profiles and overlapped them with the list of LINE-1-containing genes from the TranspoGene database. An Ingenuity Pathway Analysis (IPA) of DEGs associated with LINE-1 insertion was conducted. DNA methylation of LINE-1 was assessed via combined bisulfite restriction analysis (COBRA) of lymphoblastoid cell lines from ASD individuals and unaffected individuals, and the methylation levels were correlated with the expression levels of LINE-1 and two LINE-1-inserted DEGs, C1orf27 and ARMC8.ResultsWe found that LINE-1 insertion was significantly associated with DEGs in ASD. The IPA showed that LINE-1-inserted DEGs were associated with ASD-related mechanisms, including sex hormone receptor signaling and axon guidance signaling. Moreover, we observed that the LINE-1 methylation level was significantly reduced in lymphoblastoid cell lines from ASD individuals with severe language impairment and was inversely correlated with the transcript level. The methylation level of LINE-1 was also correlated with the expression of the LINE-1-inserted DEG C1orf27 but not ARMC8.ConclusionsIn ASD individuals with severe language impairment, LINE-1 methylation was reduced and correlated with the expression levels of LINE-1 and the LINE-1-inserted DEG C1orf27. Our findings highlight the association of LINE-1 with DEGs in ASD blood samples and warrant further investigation. The molecular mechanisms of LINE-1 and the effects of its methylation in ASD pathobiology deserve further study.

Highlights

Autism spectrum disorder (ASD) is a group of neurodevelopmental disorders that are characterized by behavioral impairments, including deficits in social interaction and communication, and by restricted interests and repetitive behaviors
DNA methylation of long interspersed element-1 (LINE-1) was assessed via combined bisulfite restriction analysis (COBRA) of lymphoblastoid cell lines from ASD individuals and unaffected individuals, and the methylation levels were correlated with the expression levels of LINE-1 and two LINE-1-inserted differentially expressed genes (DEGs), Chromosome 1 open reading frame 27 (C1orf27) and Armadillo Repeat Containing 8 (ARMC8)
We found that LINE-1 insertion was significantly associated with DEGs in ASD

Summary

Introduction

Autism spectrum disorder (ASD) is a group of neurodevelopmental disorders that are characterized by behavioral impairments, including deficits in social interaction and communication, and by restricted interests and repetitive behaviors. Nguyen et al (2010) conducted a largescale methylation profiling analysis of LCLs derived from monozygotic twins as well as sibling pairs discordant for a diagnosis of autism using CpG island microarrays [6] They found that several genes, including RORA and BCL2, which are involved in biological processes associated with ASD, were differentially methylated in LCLs derived from ASD subjects. These findings suggested that molecular changes observed in the peripheral blood or blood-derived cells may reflect some pathobiological conditions in the brain, further supporting the use of peripheral tissues as a surrogate to identify molecular marker candidates for ASD These studies have focused mainly on the CpG islands of protein-coding regions, while DNA methylation in non-coding regions of the genome, which are thought to be involved in gene regulatory processes, has yet to be investigated. We aimed to investigate whether LINE-1 insertion is associated with differentially expressed genes (DEGs) and to assess LINE-1 methylation in ASD

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