Investigating epigenetic changes associated with dry age-related macular degeneration

The project focuses on epigenetic dysregulation in age-related macular degeneration (AMD) and follows the largest epigenome-wide association study (EWAS) using ocular tissue (Retinal Pigment Epithelial cells) from donors with dry AMD and controls to date.

The project is based in Liverpool where Dr Louise Porter’s work focuses on genetics and epigenetics in ocular disease and Dr Paraoan’s laboratory focuses on gene expression and regulation in pigmented cells of the eye with importance for the pathophysiology of AMD. Furthermore, the project involves collaboration with the groups of Dr Anneke den Hollander and Dr Eiko de Jong, with extensive expertise in genetics and epigenetics, at the Radboud University, Netherlands.

Age-related macular degeneration (AMD) is the commonest cause of vision loss in the UK. Genetic approaches have identified major common variants conferring AMD risk within complement factor H (CFH) (Y402H) and the age-related maculopathy susceptibility 2 (ARMS2) and adjacent high-temperature requirement factor (HTRA1) genes [1]. While some biochemical explanations for the chromosome 1 association with disease have been investigated [2] and the identification of a number of genetic variants that contribute to either a ‘protective’ or ‘risk’ haplotype [1], it is still unclear what effect these SNPs have on gene expression levels overall.

Epigenetic mechanisms add an additional dimension to gene regulation, influencing expression and function without modification of the base sequence of DNA. Epigenetic modifications include DNA methylation, histone modifications and non-coding RNA and have been implicated in the development of human disease [3]. DNA methylation, particularly at CpG dinucleotides, is an important mechanism of gene silencing [3]. Differential CpG dinucleotide DNA methylation levels in the vicinity of AMD risk loci have been shown to result in altered gene expression levels of target genes [4,5].

The student will be involved in the analysis of a novel dataset of epigenomic data and gain experience in a number of validation methods including pyrosequencing as well as in core genetics and molecular biology techniques. Further functional work will be performed to analyse the effect of differentially methylated genes using cell biological techniques. We have access to a tissue bank of fresh ocular tissue at the University of Liverpool with opportunities and experience in RT-PCR, RNA-sequencing, immunohistochemistry and the culture of primary RPE cells for protein studies (western blotting, immunofluorescence) and RNA analysis. Replication of differentially methylated (DM) loci identified using the above methods will also be attempted in blood-derived DNA of AMD donors, by performing pyrosequencing. Correlations with epigenetic profiles derived from ocular tissue DNA will identify similarities, and eye-specific differences, associated with AMD, and will serve to identify useful biomarkers.

Methods:
• Eye collection, RPE cell harvesting and DNA extraction
• Genome-wide methylation analysis data interpretation
• Pyrosequencing
• Cell culture
• Western blotting
• Immunohistochemistry
• RT-PCR / RNA sequencing

The Institute of Ageing and Chronic Disease is fully committed to promoting gender equality in all activities. We offer a supportive working environment with flexible family support for all our staff and students and applications for part-time study are encouraged. The Institute holds a silver Athena SWAN award in recognition of on-going commitment to ensuring that the Athena SWAN principles are embedded in its activities and strategic initiatives.

This project is most suitable to a student with an undergraduate degree or Master’s degree in the Life Sciences (Biology, Genetics, Biochemistry, Biomedical Sciences, Cell Biology or Medicine with an interest in vision science).