Epigenetic Dysregulation of SHANK3 in Brain Tissues from Individuals with Autism Spectrum Disorders
Although a genetic component is strongly implicated in the etiology of autism spectrum disorders (ASD), the molecular basis remains poorly understood in the majority of cases; single gene mutations and chromosomal microdeletions or duplications are found only in about 10-20% of idiopathic ASD cases. The increasing prevalence of ASD also points to the role of gene and environment interaction in ASD susceptibility. Through analysis of the Angelman and Prader-Willi syndrome imprinting region in brain tissue from individuals with ASD, we proposed a mixed genetic and epigenetic model for the etiology of ASD. A similar model has also been reported by others. Evidence from recent genetic studies suggests that dysfunction of brain synapses underlies the pathogenesis of ASD which led us to hypothesize that epigenetic dysregulation of synaptic genes may be implicated in the molecular basis of ASD.
To test a hypothesis that ASD have an epigenetic cause by performing DNA methylation profiling of 5 CpG islands (CGI-1 to CGI-5) in the SHANK3 gene in post mortem brain tissues from 54 ASD patients and 43 controls.
We used bisulfite genomic sequencing and pyrosequencing methods to determine the level of DNA methylation of SHANK3 CGIs in post mortem brain tissues from individuals with ASD and controls. We also performed expression analysis in brain tissues and treated the cultured cells with DNA inhibitor and promotor
We found significantly increased overall DNA methylation (epimutation) in three intragenic CGIs (CGI-2, CGI-3, and CGI-4). The increased methylation was clustered in the CGI-2 and CGI-4 in ~15% of ASD brain tissues. SHANK3 has an extensive array of mRNA splice variants resulting from combinations of 5 intragenic promoters and alternative splicing of coding exons. Altered expression and alternative splicing of SHANK3 isoforms were observed in brain tissues with increased methylation of SHANK3 CGIs in ASD brain tissues. A DNA methylation inhibitor modified the methylation of CGIs and altered the isoform-specific expression of SHANK3 in cultured cells.
This study is the first to find altered methylation patterns in SHANK3 in ASD brain samples. Our finding provides evidence to support an alternative approach to investigating the molecular basis of ASD. The ability to alter the epigenetic modification and expression of SHANK3 by environmental factors suggests that SHANK3 may be a valuable biomarker for dissecting the role of gene and environment interaction in the etiology of ASD.