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Origin and Role of Nuclear Somatic Mutations in Frontotemporal Dementia


Type

Thesis

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Authors

Nair, Sulekha 

Abstract

Ageing is a natural process that occurs in all cells of multicellular organisms, including non-dividing cells like neurons, and is associated with the accumulation of mutational burden and a loss of genomic maintenance over time. Recent advances in technology have brought attention to the accumulation of somatic mutations in neurons that may cause ageing and several specific mutational signatures have been associated with disease states. Frontotemporal Dementia (FTD) is a young-onset form of dementia displaying great heterogeneity with respect to clinical phenotype and pathology. Several genes have been implicated in FTD, but their true mutational prevalence has been poorly understood due to the limitations of accuracy and reproducibility of bulk and single-cell sequencing technologies at the sub-clonal levels, respectively. Duplex Sequencing, the error-corrected next generation sequencing (NGS) technique that combines the strengths of bulk and single-cell sequencing, was applied to genomic DNA extracted from three different frozen brain regions of a patient with FTD-Tau pathology and an age-matched control. Two main approaches to DS were explored – the amplicon-based and the double capture-based methods – and two different protocols were investigated for each method. Optimizations of several parameters were required, particularly of the library amplification step, to obtain the optimal number of reads to make a duplex consensus of true mutations in each sample. Although the amplicon-based method was easier and less time-consuming, the double capture-based method that used a greater number of biotinylated probes proved able to remove more false-positives and detect true mutations more accurately. Due to the selective degeneration of the frontal and temporal lobes in FTD, we hypothesized that the number of somatic mutations detected in the temporal sample of the FTD-Tau patient would be higher than the other brain regions of the patient which would in turn be greater than the age-matched control. While the overall number of mutations detected in the iii FTD-Tau patient is higher than in the age-matched control, the overall number of mutations detected in the temporal lobe were comparable to those detected in the occipital lobe and the medulla brain regions of the patient. However, the per-bp mutation rate of the temporal lobe of the patient was the highest of the six brain regions analysed, indicating that this brain region may be more susceptible to mutation accumulation in the disease state. Despite being a sporadic case of FTD, the patient exhibited a statistically significant presence of early-arising somatic single nucleotide variations (SNVs) (variable allele frequency, VAF ≥ 0.05 and VAF < 0.25), ultralow frequency somatic SNVs (VAF < 0.01), and germline SNVs (VAF > 0.25 and VAF < 1.00), indicating a complex interplay of mutational accumulation being associated with the FTD-Tau disease state in the patient. Somatic SNVs (VAF < 0.25) particularly found within the intronic, exonic, ncRNA, and the UTR regions have a higher presence in the patient than the control, indicating a relationship between somatic mutational burden in genomic regions not directly involved in the transcription and translation of the tau gene and disease state in the patient. This study is important in that it sheds some light on the origin and role of nuclear mutations, particularly that of ultralow frequency somatic SNVs, in the pathogenesis of the FTD-Tau disease state in a sporadic patient case versus an age-matched control by successfully applying the error-corrected NGS technique, duplex sequencing, on DNA extracted from different brain regions of both the individuals.

Description

Date

2022-05-30

Advisors

Chinnery, Patrick

Keywords

Duplex Sequencing, Frontotemporal Dementia, FTD, FTD-Tau pathology, Nuclear somatic mutations, Single nucleotide variation detection, SNV detection, Somatic mutations, Ultralow frequency mutations

Qualification

Doctor of Philosophy (PhD)

Awarding Institution

University of Cambridge
Sponsorship
Cambridge Trust - Cambridge-India Ramanujan Scholaship and the Cambridge Brain Bank supported by the NIHR Cambridge Biomedical Research Centre.