DNA methylation remodeling across adulthood in the rhesus macaque hippocampus

DNA methylation (DNAm) is a key epigenetic mechanism implicated in brain aging, yet methylation trajectories across the lifespan in the primate brain remain poorly understood. To characterize age-associated DNAm patterns in the hippocampus, we generated reduced representation bisulfite sequencing (RRBS) data from 86 rhesus macaque hippocampus samples spanning ~3-35 years of age (females = 56; males = 30). CpG-level analyses revealed minimal age-associated changes in mean methylation, with sites trending toward intermediate states, but widespread increases in methylation variability, consistent with epigenetic drift and reduced regulatory fidelity. Region-level modeling identified 5163 age-associated methylation regions exhibiting both monotonic and nonlinear trajectories across age. Nonlinear clusters revealed midlife inflections centered around 12.3 and 13.2 years, suggesting that hippocampal epigenetic aging may involve molecular transition periods that shape later-life brain aging trajectories. Integration of these age-associated DNAm regions with matched RNAseq data identified candidate cis-regulatory loci where age-associated methylation may be linked to transcriptional changes, including IGF1R. In addition, direct testing of associations between mean gene methylation and expression revealed a significant repressive directional bias (higher methylation associated with lower expression) highlighting genes involved in neuroinflammation, amyloid processing, and myelin maintenance. Intergenic regions overlapped annotated human brain enhancers, indicating that age-associated methylation remodeling extends to distal cis-regulatory elements. Together, these findings indicate that hippocampal DNAm aging in a long-lived primate is dominated by increasing epigenetic variability at the CpG level alongside regional remodeling of both genic and intergenic regulatory loci, with potential implications for transcriptional remodeling during brain aging.