Student Theses and Dissertations
Date of Award
2025
Document Type
Thesis
Degree Name
Doctor of Philosophy (PhD)
Thesis Advisor
Titia de Lange
Keywords
replicative senescence, ATM signaling, telomere shortening, TRF2 deficiency, hypoxia, DNA damage response
Abstract
Somatic human cells have a limited proliferative capacity, dividing a certain number of times before entering a predictable proliferative arrest. Telomeres become shorter with every division until a few become 'critically-short' and activate the DNA Damage Response (DDR), thereby inducing this replicative senescence. However, the nature of the activated DDR is unclear. ATM kinase activation at critically short telomeres has been implicated in the induction of senescence, suggesting that the shortest telomeres lack su]icient TRF2 to repress ATM signaling. However, ATM-deficient cells also undergo replicative senescence, raising the question of POT1 deficiency and subsequent ATR signaling. Moreover, low oxygen culture conditions impart an extension of fibroblast proliferative lifespan, indicating the nature of the critically-short telomere, or the response to that telomere, may be context-dependent. The permanence of senescence is also unclear. Traditionally, senescence has been considered an irreversible state, but evidence suggests interventions specific to the senescent-inducing signal may allow some arrested cells to re-enter the cell cycle. Finding the main driver of replicative senescence – and interrupting that signal – would allow the purported inflexibility of proliferative arrest to be tested. Using specific inhibitors of ATM, ATR, Chk2, and Chk1, we show that replicative senescence is solely due to ATM signaling, and that deficiency in TRF2 is the primary reason for ATM activation at critically short telomeres. In further support of this, we find that ataxia-telangiectasia cells (deficient in functional ATM expression) tend to bypass a traditional replicative senescence, instead entering telomeric crisis. Using FUCCI live-cell imaging, we establish that ATM inhibition allows a subset of wildtype senescent cells to re- enter the cell cycle and progress through several cell divisions. RNA-seq and p38 inhibitor culturing show ATM-inhibited post-senescent fibroblasts eventually enter a second replicative arrest due to p38 α/β-p53-p21 pathway induction. We also show that cells maintained at low (3%) oxygen have a greater tolerance for critically-short telomeres, despite unaltered shelterin expression. Instead, this greater tolerance of short telomeres is due to a diminished ability of ATM to respond to DSBs, despite normal expression of ATM pathway components. Our data is consistent with hypoxia-induced sequestration of ATM in inactive dimers, explaining why cells are more tolerant to critically short telomeres and have an extended proliferative lifespan when maintained at physiological oxygen levels.
License and Reuse Information
This work is licensed under a Creative Commons Attribution-NonCommercial-Share Alike 4.0 International License.
Recommended Citation
Stuart, Alexander James, "Replicative Senescence is ATM-Driven, Reversible, and Delayed by Reduced ATM Activity at Low Oxygen" (2025). Student Theses and Dissertations. 826.
https://digitalcommons.rockefeller.edu/student_theses_and_dissertations/826
Comments
A thesis presented to the faculty of The Rockefeller University in partial fulfillment of the requirements for the degree of Doctor of Philosophy