Exploring the Stability of Genomic Imprinting and X-Chromosome Inactivation in the Aged Brain (IMPLICON-Seq)
DNA methylation is a common mechanism regulating monoallelic expression of different genes, including imprinted genes. How age-induced alterations of methylation levels vary across alleles and impact on monoallelic expression is unknown. In the brain, this phenomenon may contribute mechanistically to neuronal dysfunction and disease. In order to investigate the allele-specific transcriptional and epigenetic signatures of aging, we used key brain areas, hippocampus and cerebellum, of juvenile and old hybrid mice obtained from BL6×CAST reciprocal crosses. We set out to explore whether epigenetic drift affects global DNA methylation machinery by investigating 5- methylcytosine (5-mC) and 5-hydroxymethylcytosine (5-hmC) levels. Mass spectrometry analysis showed a significant increase of 5hmC levels in hippocampus of old mice, but this was not accompanied by measurable differences in methylating or demethylating enzymes. The expression of several repetitive elements (LINEs, SINEs, IAP) was also analysed with a reduction of the intracisternal A-particle (IAP) retrotransposons seen in HCP of old mice. We uncovered the allelic-specific DNA methylation profile of aging using genomic imprinting as a read-out by IMPLICON and allelic-expression by RNA sequencing. Here, imprinting is not substantially changed during aging, not affecting their normal monoallelic expression. This work provides the first epigenetic and transcriptional landscape of aging with allelic resolution and confirms that genomic imprinting is a stable epigenetic phenomenon across the whole lifespan. Overall design: We used key brain areas, mostly focused in hippocampus (HCP) and cerebellum (CB), of juvenile and old hybrid mice obtained from BL6×CAST reciprocal crosses. We set out to explore whether epigenetic drift affects global DNA methylation machinery by investigating 5-methylcytosine (5-mC) and 5-hydroxymethylcytosine (5-hmC) levels by Liquid Chromatography with tandem mass spectrometry and the genes enconding for enzymes implicated in this process by RT-PCR and RNAseq, the later in HCP only . The expression of several repetitive elements (LINEs, SINEs, IAP) was also analysed by RTq-PCR and also by RNAseq in HCP . We uncovered the allelic-specific DNA methylation profile of aging using genomic imprinting as a read-out by IMPLICON in several brain areas such as hippocampus (HCP), cerebellum (CB), prefrontal cortex (PFC), hypothalamus (HY), nucleus accumbens (NA) and a control region such as lung. Allelic expression analysis by RNA sequencing was performed in HCP.