Functional annotation of gene variants in autism risk genes using DNA methylation signatures

  • Awarded: 2021
  • Award Type: Targeted: Genomics of ASD: Pathways to Genetic Therapies
  • Award #: 887172

The role of dysregulated chromatin biology in autism spectrum disorder (ASD) has become increasingly apparent as genome sequencing has implicated numerous epigenetic regulatory genes in ASD etiology. Notably, more than 20 of the 50 prioritized SPARK genes function in chromatin remodeling, including modifications to histone proteins and/or DNA (e.g., DNA methylation).

Rosanna Weksberg’s laboratory has described DNA methylation “signatures” for multiple rare neurodevelopmental disorders caused by pathogenic variants in epigenetic regulatory genes1-5. These DNA methylation signatures consist of unique patterns of genome-wide DNA methylation that represent aberrant downstream signals of the primary pathogenic variant. Weksberg and her colleagues have shown that these signatures identify disruptions in gene regulation that constitute the molecular pathophysiology of each disorder and also reflect the clinical phenotype. From these signatures, Weksberg’s team derived novel functional tests for classifying variants of uncertain significance (VUS) in these genes with high sensitivity and specificity (greater than 95 percent).

The identification of VUS in individuals with ASD is a growing obstacle to definitive molecular diagnoses as the clinical use of genome sequencing increases. While variant databases such as ClinVar and in silico missense prediction tools can provide some insights into variant pathogenicity, they often do not provide definitive answers. Weksberg’s team proposes that the application of DNA methylation signature technology to ASD could ultimately resolve uncertain diagnoses for many affected individuals. Furthermore, the utility of DNA methylation signatures goes beyond VUS classification; they enable the identification of unique DNA methylation profiles for atypical variants (e.g., gain-of-function) and variant clusters/subregions of genes with distinct clinical presentations [i.e., (epi)genotype-phenotype correlations]. Such findings will support a role for this platform as a valuable adjunct to genome sequencing in clinical diagnostics and translational research with potential utility for the development of biomarkers and targeted therapies.

In the current project, Weksberg and colleagues propose to derive DNA methylation signatures for 23 SPARK genes with VUS rates of approximately 28 percent and known or potential functions in chromatin biology. From these signatures, they will develop predictive tools to classify VUS in these genes. They hypothesize that DNA methylation signatures will serve as functional assays for VUS classification and for the definition of critical domains in 23 SPARK genes, thereby enhancing accurate diagnostic classification of gene variants in ASD cohorts.


  1. Choufani S. et al. Commun. 6, 10207 (2015) PubMed
  2. Butcher D.T. et al. J. Hum. Genet. 100, 773-788 (2017) PubMed
  3. Choufani S. et al. J. Hum. Genet. 106, 596-610 (2020) PubMed
  4. Chater-Diehl E et al. Am J Hum Genet. 108, 1359–1366 (2021) PubMed
  5. Awamleh Z. et al. J. Hum. Genet. 30, 695-702 (2022) PubMed
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