Characterizing the epigenetic response to stress using next generation sequencing

Röh-Karamihalev, Simone (2023): Characterizing the epigenetic response to stress using next generation sequencing. Dissertation, LMU München: Fakultät für Biologie

Abstract

Stress-related disorders have complex etiologies and high incidences, with mood disorders expected to become the leading cause of disability in the coming years. The mechanisms leading to resilience or susceptibility are not well understood and involve both genetic and environmental aspects, especially exposure to stress and adversity, integrated in the form of epigenetic changes such as DNA methylation or histone modification. In recent years, next generation sequencing (NGS) has become a powerful tool for epigenetic profiling. This thesis aims to improve on existing, as well as designing new NGS methods with increased sensitivity and accuracy for improved detection of small epigenetic changes even in composite tissues such as blood. We apply these methods to study DNA methylation changes in the context of stress by genotype interaction within the FKBP5 gene locus. In the scope of minimizing the bias and maximizing the sensitivity of NGS technology, the first part of this work represents an in-depth characterization and reduction of GC bias on the SOLiD sequencers relative to the currently dominant Illumina sequencing technology. Subsequently, we present a new method, termed HAM-TBS, which enables highly sensitive assessment of methylation levels of a target set of CpG sites in studies with cohort-level sample sizes. Using HAM-TBS, we can tailor the set of CpGs to fit the biological question at hand. To our knowledge, this is the most sensitive method to detect methylation levels in NGS data to date allowing us to resolve small changes. With it, we supply a fully tested panel of amplicons targeting selected CpGs in functionally relevant regions within the FKBP5 locus including glucocorticoid receptor elements, CTCF binding sites, topologically associating domain boundaries and the transcription start site. Finally, we apply HAM-TBS using the established amplicon panel to assess dynamic methylation changes in the FKBP5 gene locus following a stress challenge. We expose a cohort of healthy individuals to dexamethasone (DEX), a synthetic glucocorticoid activating the stress reactive hypothalamic-pituitary- adrenal axis. We found dynamic and lasting methylation changes in blood as well as a genotype-dependent response following DEX exposure. This work demonstrates the potential of NGS technologies to aid in improving our understanding of epigenetics in the context of the stress response system.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Keywords: next generation sequencing, epigenetics, FKBP5, HPA axis, DNA methylation, targeted bisulfite sequencing
• Themengebiete: 500 Naturwissenschaften und Mathematik ; 500 Naturwissenschaften und Mathematik > 570 Biowissenschaften, Biologie
• Fakultäten: Fakultät für Biologie
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 21. Dezember 2023
• 1. Bericht­erstatter:in: Schmidt, Mathias