The completion of a telomere-to-telomere human reference genome, T2T-CHM13, has resolved complex regions of the genome, including repetitive and homologous regions. Here, we present a high-resolution epigenetic study of previously unresolved sequences, representing entire acrocentric chromosome short arms, gene family expansions, and a diverse collection of repeat classes. This resource precisely maps CpG methylation (32.28 million CpGs), DNA accessibility, and short-read datasets (166,058 previously unresolved chromatin immunoprecipitation sequencing peaks) to provide evidence of activity across previously unidentified or corrected genes and reveals clinically relevant paralog-specific regulation. Probing CpG methylation across human centromeres from six diverse individuals generated an estimate of variability in kinetochore localization. This analysis provides a framework with which to investigate the most elusive regions of the human genome, granting insights into epigenetic regulation.

We performed whole-genome sequencing (WGS) of 208 genomes from 53 families affected by simplex autism. For the majority of these families, no copy-number variant (CNV) or candidate de novo gene-disruptive single-nucleotide variant (SNV) had been detected by microarray or whole-exome sequencing (WES). We integrated multiple CNV and SNV analyses and extensive experimental validation to identify additional candidate mutations in eight families. We report that compared to control individuals, probands showed a significant (p = 0.03) enrichment of de novo and private disruptive mutations within fetal CNS DNase I hypersensitive sites (i.e., putative regulatory regions). This effect was only observed within 50 kb of genes that have been previously associated with autism risk, including genes where dosage sensitivity has already been established by recurrent disruptive de novo protein-coding mutations (ARID1B, SCN2A, NR3C2, PRKCA, and DSCAM). In addition, we provide evidence of gene-disruptive CNVs (in DISC1, WNT7A, RBFOX1, and MBD5), as well as smaller de novo CNVs and exon-specific SNVs missed by exome sequencing in neurodevelopmental genes (e.g., CANX, SAE1, and PIK3CA). Our results suggest that the detection of smaller, often multiple CNVs affecting putative regulatory elements might help explain additional risk of simplex autism.

The progressive loss of midbrain (MB) dopaminergic (DA) neurons defines the motor features of Parkinson disease (PD), and modulation of risk by common variants in PD has been well established through genome-wide association studies (GWASs). We acquired open chromatin signatures of purified embryonic mouse MB DA neurons because we anticipated that a fraction of PD-associated genetic variation might mediate the variants' effects within this neuronal population. Correlation with >2,300 putative enhancers assayed in mice revealed enrichment for MB cis-regulatory elements (CREs), and these data were reinforced by transgenic analyses of six additional sequences in zebrafish and mice. One CRE, within intron 4 of the familial PD gene SNCA, directed reporter expression in catecholaminergic neurons from transgenic mice and zebrafish. Sequencing of this CRE in 986 individuals with PD and 992 controls revealed two common variants associated with elevated PD risk. To assess potential mechanisms of action, we screened >16,000 proteins for DNA binding capacity and identified a subset whose binding is impacted by these enhancer variants. Additional genotyping across the SNCA locus identified a single PD-associated haplotype, containing the minor alleles of both of the aforementioned PD-risk variants. Our work posits a model for how common variation at SNCA might modulate PD risk and highlights the value of cell-context-dependent guided searches for functional non-coding variation.