Exploring the role of epigenetic mechanisms in the manifestation of Huntington's disease

Lead Research Organisation: UNIVERSITY OF EXETER
Department Name: Clinical and Biomedical Sciences


Huntington's disease (HD) is a neurodegenerative disease caused by an expansion of typically 40 or more repeats of the DNA code "CAG" in the Huntingtin (HTT) gene. The disease is characterised by movement disturbances, cognitive impairments, and psychiatric symptoms and there is currently no disease-modifying treatment. The size of the CAG repeat is closely associated with the age of symptom onset, with individuals with high numbers of repeats developing the disease at a young age. However, there is variation in the age of symptom onset between individuals with the same CAG repeat length. It is known that the expression of genes relies not only on a person's specific DNA code (their genome) but can also be altered by an extra level of information called the "epigenome". Epigenetic processes are chemical tags added to the DNA or histone proteins that turn genes on and off and can be influenced by external factors. We have recently shown robust alterations in two epigenetic marks (DNA methylation (DNAm) and H3K27ac) in Alzheimer's disease (AD). We have also seen DNAm differences in a pilot study of HD brain. We hypothesise that epigenetic mechanisms contribute to the manifestation of HD and plan to use state-of-the art genomic technology and computational approaches to undertake the most comprehensive study of epigenetic mechanisms in HD brain to date. We have the following complimentary work-packages:

We will use cutting-edge long-read sequencing technology to measure the length of the CAG repeat and extent of DNAm across that region of the HTT gene. This will allow us to determine exactly where DNAm is seen in the CAG repeat in HD brain samples. We plan to study two brain regions: the striatum and prefrontal cortex, which are affected at different stages of the disease. It is reported that the CAG repeat length can increase in some cells with age, which is termed somatic mosaicism. One advantage of using long-read sequencing is that we can measure both the CAG repeat length and DNAm on the same molecule, which will allow us to explore this phenomenon in our samples.

We will perform the first genome-scale assessment of epigenetic variation in HD brain samples, profiling DNAm, H3K27ac, chromatin accessibility and genetic variation in the striatum and prefrontal cortex. We will investigate whether DNAm, H3K27ac or areas of open chromatin are seen at specific gene regions in HD. As we have generated similar datasets in other neurodegenerative diseases, we can then examine whether there is any overlap in the epigenetic changes we identify in HD to, for example, AD. Using integrative computational approaches, we will explore the relationship between different layers of epigenetic information. By integrating genetic data, we can identify quantitative trait loci (QTLs), where genetic differences alter the epigenetic marks, and then investigate if these QTLs are enriched in genes we know are affected in HD, and other related disorders.

We will isolate nuclei from different cell types in the prefrontal cortex using fluorescence-activated nuclei sorting (FANS), including inhibitory (GABA) and excitatory (glutamatergic) neurons that are known to be affected in disease, as well as glial cells such as microglia, oligodendrocytes and astrocytes. We will determine which cell types are responsible for the epigenetic changes we observed in the earlier WP in a subset of the cohort with severe pathology (N=20), moderate pathology (N=20) or no pathology (N=20)

We will use long-read sequencing to measure expression of the HTT gene, and other genes we have identified. The advantage of this technology is that we can identify completely novel isoforms, which we have previously done in AD.

Technical Summary

Although the size of the CAG repeat expansion in Huntington's disease (HD) is negatively correlated with age of symptom onset, there is variation amongst individuals with the same CAG repeat length. We hypothesise that epigenetic mechanisms contribute to the manifestation of HD.

In a well-characterised cohort of matched striatum and prefrontal cortex HD and control brain samples, we will undertake four complimentary work-packages (WPs) to address our hypothesis and nine important research questions (RQs). First, we will use cutting-edge long-read sequencing to characterise exon 1 of HTT at a genetic and epigenetic level (WP1). This will allow us to investigate the extent of DNA methylation (DNAm) in the CAG repeat expansion (RQ1), and whether there is somatic mosaicism in the CAG repeat, with respect to DNAm and length (RQ2). Second, we will profile DNAm, H3K27 acetylation and chromatin accessibility in our brain samples (WP2). This will allow us to identify epigenetic signatures in HD, and how these relate to variation in CAG repeat length and age of onset (RQ3). As we have generated similar datasets in other neurodegenerative diseases, we will explore the disease specificity of our loci (RQ4). Data integration will allow us to explore the relationship between the data modalities in HD (RQ5). Furthermore, by incorporating genomic data we will be able to identify quantitative trait loci (QTLs) and explore whether these are enriched in genome-wide association study (GWAS) loci (RQ6). Third, we will determine the cell specificity of these changes using flow cytometry (WP3). This will allow us to assess whether DNAm alterations are driven by specific cell types in the brain (RQ7). Finally, we will use targeted long-read sequencing to explore transcript diversity of HTT and other candidate genes we identify (WP4). This will allow us to quantify novel transcript isoforms in HD (RQ9), which are associated with disease, CAG repeat length, and the epigenetic landscape


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