Basic Science and Pathogenesis

We have reported a statistical interaction between the African ancestry specific A allele at rs10423769 and APOEε4, where the presence of the A allele is associated with a reduction in AD risk up to 75% in APOEε4 homozygotes. The mechanism by which this variant confers protection could provide insights into new therapeutics for APOEε4 carriers. However, the genomic context of this protective variant is complex with a high number of segmental duplications (SD) and structural variants (SVs). Thus, we have used advanced genomic techniques to investigate the region of ∼2MB between rs10423769_A and APOE to gain further insights into potential AD-protective mechanisms.

We used high-coverage (90x) PacBio HiFi whole genome sequencing on 9 samples of rs10423769_A/A, rs10423769_A/G, and rs10423769_G/G carriers. We performed genome assembly with hifiasm to identify contigs spanning the whole region between the protective allele and APOE. We compared the contigs using minigraph and Bandage. We performed Hi-C sequencing from 13 brain autopsy samples (seven rs10423769_A carriers) and used in-house bioinformatics tools (enhanced Hi-C Capture Analysis - eHiCA) to identify chromatin loops indicative of potential interactions between the protective and APOE loci.

Long read sequencing detected the co-occurrence of an expanded VNTR containing multiple MEF2D transcription factor binding sites within the protective haplotype. A higher number of SVs in the SD region surrounding rs10423769_A haplotype were detected when compared to the reference and are under investigation. eHiCA using rs10423769 or the APOE promoter as the bait suggested that rs10423769 locus and the APOE promoter region have evidence of long-range, physical interactions. Furthermore, both rs10423769 locus and the APOE promoter interact with common regions in between the two baits, including a cluster of zinc finger (ZNF) genes upstream of APOE. The reciprocal eHiCAs suggest high confidence interactions between these loci and with other regions of chr19.

Our results show that the protective haplotype locus of African origin has long-range, physical interactions with the APOE promoter and other regions on chr19. It also carries an expanded VNTR specific to the protective haplotype containing multiple transcription factor binding sites.