Do oxidative breaks accumulate at gene regulatory regions in disease?

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The recent revolution in genomics aspires to provide better understanding of how changes in the DNA sequence cause disease, which will improve health outcomes by providing better diagnostics and therapeutics. This could be achieved by developing smart sequencing panels for early diagnosis and exploiting vulnerabilities in DNA repair to treat cancer or correcting single gene disorders using gene editing and replacement in neurological disease. Although many patterns of mutation (signatures) arise due to defective DNA repair, some remain of unknown aetiology, and how they lead to disease phenotype is often unknown. Therefore, there is a need to define biological mechanisms generating such mutations and determine their link to human disease. So far, the role of many mutation signatures has only been studied in protein-coding parts of the genome. However, recently, we have found that oxidative DNA breaks and abasic sites arise in the regulatory parts of mammalian genomes and described a mechanism for their repair (Ray et al., Nature 609 :1038-1047, 2022). Although increased oxidative stress is associated with several neurological disorders, the mechanistic pathway from this to disease is currently unclear. We propose that oxidative DNA breaks may accumulate at gene regulatory regions and that these regions may exhibit increased mutational burden reminiscent of defective oxidative break repair, eventually leading to their malfunction and disease phenotypes. In this project, we will investigate oxidative DNA breaks at regulatory genomic regions in two neurological diseases caused by defects in the DNA repair proteins, tyrosyl DNA phosphodiesterase I (TDP1) and ataxia telangiectasis mutated (ATM). We will do so by: 1. Determining the profile of oxidative DNA breaks in patient derived neurons using genomic mapping methods that we recently described in Ray et al., 2022. 2. Establishing the source of endogenous oxidative breaks at gene regulatory regions in patient derived neurons, by genetic and chemical manipulations 3. Exploring the impact of unrepaired oxidative breaks on the early stages of the transcription cycle, using methods that we recently described in Ray et al., 2022.4. Determining whether the mutational burden is increased at gene regulatory regions in patient derived cells Revealing how oxidative breaks impact gene regulatory regions and transcriptional responses will increase our understanding of the aetiology of these two neurological diseases and pave the way of similar approaches in other neurological disease that feature increased oxidative stress but not directly linked to mutations of DNA repair proteins, such as Alzheimer's disease and dementia.