A fragile code:DNA damage as a fundamental driver of aging

Over the past century, increased average lifespans have resulted in a rapidly aging population. Aging is accompanied by a rising incidence of chronic, age-related diseases such as cancer, neurodegenerative conditions, and frailty, all of which place considerable strain on healthcare systems, impact individual well-being, and threaten socioeconomic stability. Understanding aging's underlying mechanisms is vital to mitigating these challenges and improving quality of life. This thesis mainly focusses on the role of DNA damage in the process of aging and explores its role in four interrelated aspects of aging: transcriptional stress, frailty, ovarian aging, and cellular senescence. By integrating experimental data, mathematical modeling, and genetic analysis, this work aims to elucidate how persistent DNA damage contributes to functional decline with age and to age-associated pathologies. 1. Transcriptional-stress Transcription, the process by which DNA is converted into RNA, is essential for proper gene expression and cellular function. However, DNA damage can block transcription, particularly in long genes, leading to transcriptional stress. This results in imbalanced mRNA production and impaired cellular homeostasis, both hallmarks of aging cells. In Chapter 2, a mathematical model was developed to simulate the accumulation of transcription-blocking lesions (TBLs) over time. Validated in both human cells and mouse tissues, the model offers a quantitative framework for understanding how DNA damage disrupts transcription. These results support the concept that transcriptional dysregulation due to DNA damage is a progressive, quantifiable feature of aging. 2. Frailty Frailty is a multidimensional syndrome characterized by decreased physiological reserves and increased vulnerability to stressors, often used to assess biological aging beyond chronological age. In Chapter 3, this thesis proposes that transcriptional stress is a driving force behind frailty. Using data from human cohorts, transcriptional stress was linked to frailty scores and gene expression signatures. Additionally, frailty was assessed in DNA-repair-deficient mouse models (Xpg-/- and Ercc1d/-), which are known to experience high levels of transcriptional stress. Furthermore, we identified specific genes associated with frailty and observed increased transcriptional stress upon their knockdown. Together, these results suggest that accumulating DNA damage and transcriptional stress are important contributors to the frailty phenotype observed in aging. 3. Ovarian-aging Ovarian aging refers to the gradual decline in the quantity and quality of oocytes, a key factor determining female fertility and the timing of menopause. This process is influenced by a combination of genetic, environmental, and lifestyle factors. Chapter 4 focuses on the MCM8-E341K genetic variant, which is associated with a delayed age at natural menopause (ANM) and an increased risk of breast cancer. To understand the cellular impact of this variant, a CRISPR-Cas9-engineered human cell line carrying the E341K mutation was generated. Functional studies revealed that this variant impairs DNA repair, specifically leading to replication stress and altering the cellular response to stalled or collapsed replication forks. These findings link defective DNA repair to extended reproductive lifespan on the one hand, and increased cancer susceptibility on the other, highlighting a dual-edged nature of aging-related genetic variants. 4. Cellular-senescence Cellular senescence is a state in which cells permanently cease dividing in response to stress, including unresolved DNA damage. While this mechanism initially serves as a protective barrier against tumorigenesis, the accumulation of senescent cells over time contributes to tissue dysfunction and chronic inflammation, promoting both aging and disease. A key challenge in the field is the reliable identification and tracking of senescent cells in real time. In Chapter 5, a novel live-cell lamin-based senescence reporter was developed to monitor therapy-induced senescence (TIS) and senescence escape in cancer cell lines. This tool enables dynamic tracking of senescent states and could be instrumental for future studies targeting senescence in cancer and aging research. In summary, this thesis sheds light on the intricate relationship between DNA damage and four critical aspects of aging. By studying transcriptional stress, frailty, ovarian aging, and cellular senescence, this work contributes to a more integrated understanding of how DNA damage drives aging-related decline. The insights gained here may ultimately inform strategies to delay aging, prevent age-related diseases, and promote healthier, more resilient aging across the population.