Impact of dynamic methylome on variants in ageing

Villoo (Avesthagen Ltd.) introduced the presentation by first describing the Avestagenome Project, which aims to investigate the endogamous Parsi community to identify biomarkers and therapeutics for cancers, neurodegenerative conditions, and rare diseases. The Parsi community arrived in India from Iran approximately 13,000 years ago and generations of marriage within the community has resulted in increased evidence of traits such as greater longevity and susceptibility to certain diseases, including Parkinson’s disease, stroke, heart disease, specific cancers and Alzheimer’s disease. The team plan to sequence over 10,000 Parsi genomes as part of this project.

Villoo’s colleague Kashyap Krishnasamy then provided a brief introduction to epigenetic modifications in the context of DNA methylation, explaining how they are both heritable and dynamic, with strong correlation to age-related disease outcomes. To identify age-related epigenetic signatures, the team performed nanopore sequencing, using the GridION device, of two samples obtained from the same individual taken 12 years apart. Kashyap showed a detailed workflow describing the analysis process, from raw sequence reads through to methylation calling using Nanopolish, and variant calling using Nanocaller. It was demonstrated that the most recently obtained sample accrued far more CpG methylation, distributed across most chromosomes. Kashyap then described how, across both samples, the genic and gene promoter regions had the highest levels of methylation. In total, 26,019 differentially methylated genes were identified between the two samples, which were curated using in-house datasets to yield ‘5,258 genes of critical relevance for further analysis’.

Further analysis identified 103 genes that are significantly hypermethylated and 307 genes that are hypomethylated. It was evident that degree of methylation was different among different protein families, for example, oncogenes were typically hypomethylated.

The next step was the identification of epigenome-genome interactions which have documented pathological significance, where they found clusters of genes that were significantly methylated (hyper- or hypo-) with variants at the CpG sites. The majority of clustered genes were implicated in pathways regulating cancer, neurodegenerative disease, and pathways implicated in cellular senescence and longevity.

Kashyap then presented data showing the identification of over 100,000 variants in the more recently obtained sample. A high prevalence of C>T transitions were noted, especially at CpG sites for gene sets regulating Alzheimer’s and Parkinson’s disease pathways.

The team now plan to extend the study to larger cohorts to identify epigenetic signatures of smoking and validation of significant genome-epigenome interactions in in vitro assays.