Fig. 5: βII-spectrin mobility displays zonal dependency and relies on myosin-II-dependent contractility.

a Schematic representation of different cell zones highlighting βII-spectrin and actin complementarity. Differential zonal βII-spectrin dynamics was analyzed by FRAP in double-transfected MEFs: RFP-Actin marked the different regions, circular (5-µm diameter) or squared (5 × 5 µm). ROIs were bleached and fluorescence recovery followed for 3 min post bleach. The same cell was simultaneously analyzed for at least 2 of the 4 different zones. The resulting normalized recovery curves are presented in (b), together with a representative MEF considered for the analysis (all the 4 zones under investigation were present). c The resulting mobile fractions and half-time recoveries are plotted in the two graphs: spectrin cortex (blue), stress fibers (red), actin curvatures (green), and spectrin curvatures (purple) (statistical analysis: one-way Anova with multiple comparisons, *p = 0.014, **p = 0.0038, n = 18 cells). d Schematic representation of the dual-FRAP assay of GFP-βII-spectrin expressing MEFs, performed before (blue) and after 5 min of treatment (red). Normalized recoveries for each drug treatment are presented in (e. f) Mobile fractions and half-time recoveries are plotted in the graphs for each drug tested: untreated (blue) and after 5’ treatment (red) (statistical analysis: paired Student’ s t test, two-tailed, *p < 0.05, **p = 0.0021, ****p < 0.0001, n = 12 cells). All data are presented as mean values ± SD.