Increased substrate stiffness disrupts nuclear-cytoskeletal mechanical coupling in senescent cells.

The physical coupling between the nucleus and the cytoskeleton is essential for the mechanobiological adaptation of cells to mechanical cues presented by the surrounding extracellular matrix (ECM). Although aging is known to influence both cellular and ECM mechanical properties, it remains poorly understood how cellular senescence, a hallmark of aging, affects cellular mechano-adaptation. Here, we use substrate stiffness as a mechano-modulatory cue across three distinct models of senescence and demonstrate that senescent fibroblasts are limited in their capacity to integrate mechanical signals of increasing stiffness. The senescent nucleus undergoes progressive actin-mediated deformation and flattening as substrate stiffness increases, until a mechanical threshold is reached that provokes a decoupling of the nucleus from the cytoskeleton. This mechanical disengagement of the nucleus on stiff substrates is accompanied by a loss of cytoskeletal organization, abnormal focal adhesion (FA) maturation, and nuclear softening. We further suggest that the loss of nuclear compression is linked to changes in the nuclear localization of the key mechanosensitive transcriptional regulator Yes-associated protein (YAP). Our findings reveal a fundamental biophysical limitation in the mechano-adaptive response of senescent cells to high-stiffness environments, conditions typically associated with advanced tissue maturation and pathological scarring, which may underlie altered nuclear mechanotransduction and contribute to their specific role in both physiological and pathological contexts.