Cells were cultured on glass-bottomed 35-mm Petri dishes (MatTek) and placed on the stage of an inverted optical microscope (Olympus IX70-S8F2) equipped with a microinbucator (HCMIS ALA Science) to hold the temperature at 37°C. A commercial stand-alone AFM (Bioscope, Veeco) adapted to the optical microscope and provided with a low-spring constant (0.03 nN/nm) AFM cantilever (Veeco) was used to assess the Young’s modulus (E) of single cells, following a protocol described in detail elsewhere (Alcaraz et al., 2003 (link), 2011 (link)). The spring constant of the cantilever was calibrated using the thermal fluctuations method (Roca-Cusachs et al., 2008 (link)). In brief, three standard force versus displacement curves (F vs. z) were recorded on the perinuclear region of each cell at moderate loading force (∼1 nN) and low speed (∼5 µm/s). The E of a single cell was computed by fitting a suitable contact elastic model to each F-versus-z curve and averaging it over the three recordings. The final E for a cell population in each experimental condition was calculated from at least nine measurements for each independent experiment (n ≥ 2).