Epigenetic and transcriptional changes in aging mouse rod photoreceptors [RNA-seq]

Aging-associated functional decline and disease susceptibility are believed to be mediated, at least in part, through alterations in the epigenome. To explore epigenetic changes that might influence visual function with advanced age, we performed whole genome bisulfite sequencing of purified mouse rod photoreceptors at four different ages and identified 2054 genomic regions that gain or lose DNA methylation. Differentially methylated regions (DMRs) clustered at chromosomal hotspots, especially on Chromosome 10 that included a longevity interactome. DMRs were preferentially detected at an early stage of aging in long neuronal genes and in rod-specific regulatory regions containing open chromatin domains and H3K27 acetylation. Integration of methylome to age-related transcriptome changes, chromatin signatures and first order protein-protein interactions uncovered an enrichment of DMRs in pathways associated with aging, longevity, synaptic function, and energy homeostasis. In concordance, we detected reduced mitochondrial maximum reserve capacity with retinal age in ex vivo assays. Our study reveals age-dependent genomic and chromatin features susceptible to DNA methylation changes in rod photoreceptors and identifies associations with established and cell type-specific pathways altered in aging. Total RNA from FACS-purified rod photoreceptors was extracted using TRIzol® (Invitrogen, Carlsbad, CA), treated with DNase and cleaned up using the MagMAX mirVana Total RNA Isolation Kit (Applied Biosystems, Foster City, CA) following the manufacturer’s instructions. Libraries were constructed with SMARTer Stranded Total RNA-Seq Kit v2 – Pico Input Mammalian (Takara Bio USA, Mountain View, CA) with 4 ng of RNA and 13 PCR cycles library amplification. Paired-end reads of 125 base pairs were obtained using the HiSeq 2500 platform (Illumina, San Diego, CA). Four biological replicates from each of the four studied time points (3M, 12M, 18M, and 24M) were sequenced.