Feedback between p21 and reactive oxygen production is necessary for cell senescence

Cellular senescence—the permanent arrest of cycling in normally proliferating cells such as fibroblasts—contributes both to age‐related loss of mammalian tissue homeostasis and acts as a tumour suppressor mechanism. The pathways leading to establishment of senescence are proving to be more complex than was previously envisaged. Combining in‐silico interactome analysis and functional target gene inhibition, stochastic modelling and live cell microscopy, we show here that there exists a dynamic feedback loop that is triggered by a DNA damage response (DDR) and, which after a delay of several days, locks the cell into an actively maintained state of ‘deep’ cellular senescence. The essential feature of the loop is that long‐term activation of the checkpoint gene CDKN1A (p21) induces mitochondrial dysfunction and production of reactive oxygen species (ROS) through serial signalling through GADD45‐MAPK14(p38MAPK)‐GRB2‐TGFBR2‐TGFβ. These ROS in turn replenish short‐lived DNA damage foci and maintain an ongoing DDR. We show that this loop is both necessary and sufficient for the stability of growth arrest during the establishment of the senescent phenotype. Synopsis The phenomenon of cellular ‘senescence’—the permanent arrest of division in normally proliferating mammalian cells such as fibroblasts—is thought to be a central component of the ageing process. Senescence contributes both to age‐related loss of tissue homeostasis, as the loss of division capacity leads to impaired cell renewal, and also to protect against cancer, because it acts to block the uncontrolled proliferation of cells that may give rise to a malignant tumour. Replicative senescence is triggered by uncapped telomeres or by ‘unrepairable’ non‐telomeric DNA damage. Both lesions initiate the same canonical DNA damage response (DDR) (d'Adda di Fagagna, 2008 ). This response is characterized by activation of sensor kinases (ATM/ATR, DNA‐PK), formation of DNA damage foci containing activated H2A.X (γH2A.X) and ultimately induction of cell cycle arrest through activation of checkpoint proteins, notably p53 (TP53) and the CDK inhibitor p21 (CDKN1A). This signalling pathway continues to contribute actively to the stability of the G0 arrest in fully senescent cells long after induction of senescence (d'Adda di Fagagna et al , 2003 ). However, senescence is more complex than mere CDKI‐mediated growth arrest. Senescent cells alter their expression of literally hundreds of genes (Shelton et al , 1999 ), pr