Haplotype-resolved DNA methylation at the APOE locus identifies allele-specific epigenetic signatures relevant to Alzheimer’s disease risk

Abstract The APOE gene encodes a lipid transport protein central to Alzheimer’s disease (AD) pathogenesis. Three common alleles—ε2 (rs7412(C > T)), ε3 (reference), and ε4 (rs429358(T > C))—arise from two coding variants in exon 4 and confer distinct AD risk profiles, with ε4 increasing risk and ε2 being protective. The ε3-linked APOE variant rs769455[T] has also been associated with increased AD risk among individuals of African ancestry who also carry the APOE ε4 allele. Determining how genetic variation influences CpG methylation requires methQTL-type analyses, but conventional bisulfite and array-based approaches offer limited resolution for distinguishing allele-specific effects. Here, we use high-accuracy long-read sequencing to generate haplotype-resolved methylation profiles across the APOE locus in 332 postmortem brain tissue samples from ancestrally diverse cohorts, including 201 samples from individuals of European ancestry and 131 samples from individuals of African and African admixed ancestry. Treating each haplotype as an independent observation, OLS regression identified 18 novel differentially methylated CpG sites associated with ε2, ε4, and rs769455[T] across the APOE locus ( TOMM40, APOE, APOC1 , and APOC4 - APOC2 genes). These findings reveal distinct allele-specific methylation signatures and demonstrate the utility of long-read sequencing for resolving epigenetic variation relevant to AD risk.