Fig. 7: Characterization of dolines and working model description.
From: Caveolin-1 dolines form a distinct and rapid caveolae-independent mechanoadaptation system
a,b, TIRF microscopy snapshots of Cav1GFP cluster formation (a), comparing PTRFKO MEFs co-electroporated with either Cav1GFP and empty vector (Cav1 alone cells) or Cav1GFP and PTRF vectors (control cells) (b). Mean ± s.e.m. n = 13 independent fields pooling together three cells (Cav1 alone); and n = 15 independent fields pooling together four cells (control). Two-tailed Student’s t-test (P = 0.0317). c,d, Maximal projection of TIRF microscopy image series (from Supplementary Videos 13 and 14) of Cav1GFP clusters from Cav1 alone (left) or control (right) PTRFKO MEFs (see a and b), after treatment with methyl-β-cyclodextrin. Time lapse of a splitting cluster shown below. In d, mean ± s.e.m. n = 22 independent experiments pooling together 22 cells (Cav1 alone); and n = 20 independent experiments pooling together 20 cells (control), after treatment with methyl-β-cyclodextrin. Two-tailed Student’s t-test (P = 0.0221). e, Cluster densities across genotypes and osmotic conditions from dSTORM imaging, in three size groups. Data points outside the whiskers are considered outliers and plotted as empty circles. Boxes span Q1 to Q3 quartiles, with whiskers indicating lowest/greatest observed data point within 1.5× IQR below/above Q1/Q3. Middle line represents median; asterisks denote average value. f, Representative quantification of biochemical fractionation of Cav1 complexes from PTRFKO + PTRF (control cells, top) and PTRFKO + Cav1 MEFs (bottom) on 10–40% continuous sucrose gradients as indicated in ref. 65 (blots against Cav1 below the corresponding quantification graphs). Rightmost panels, cells treated with methyl-β-cyclodextrin. Results are representative of three independent experiments. g, A working model for Cav1-based PM tension buffering: Cav1 dolines provide gradual buffering to a wide range of mechanical perturbations; in contrast, caveolae, as a result of PTRF size restriction, constitute a mechanical switch that provides acute buffering to higher forces, only flattening beyond a certain tension threshold (high-range force sensor). Source numerical data and unprocessed blots are available in source data. STORM images set are available at Zenodo (DOI: 10.5281/zenodo.7062213).
