EHD2 is required for the stabilization of caveolae and the control of gene transcription during tension variations at the plasma membrane. (A) GSEA was performed to identify gene sets positively (+) or negatively (–) enriched by cyclic stretch in Hs578T cells depleted or not from EHD2. (B) Quantification of Cav1, Cav2, cavin1, cavin2, and flotillin-1 (Flot1) mRNA levels in HeLa cells transfected with control siRNA (CTRL) or siEHD2, after 30 min cyclic stretch. (C) Quantification of Cav1, Cav2, cavin1, and cavin2 mRNA levels in HeLa cells transfected with control siRNA (CTRL), siMOKA, or siKLF7 after 30 min cyclic stretch. (D) Representative TIRF images (left) and quantification (right) of changes in cell-surface Cav1 spot numbers in control siRNA (CTRL) or siEHD2-transfected Cav1-EGFP HeLa cells under resting (Iso), under hypo-osmotic (Hypo), and after return to iso-osmotic (Rec) conditions. Cells are delineated by dashes. (E) Quantification of changes in cell-surface endogenous Cav1 spot numbers in HeLa cells depleted (siEHD2) or not (CTRL) for EHD2 and transfected or not with EHD2-EGFP (+ EHD2) under Iso, Hypo, and Rec conditions. (F) Relative changes of the mean tether force under Hypo and Rec conditions in control siRNA (CTRL) and siEHD2 HeLa cells. (G) Quantification of cell surface Cav1 spot numbers at rest and after 30 min cyclic stretch in control siRNA (CTRL) or siEHD2 transfected HeLa cells. *, P < 0.05; **, P < 0.001; ***, P < 0.001; ****, P < 0.0001; two tailed t test. In B and C, Bonferroni’s multiple comparison test; n = 3 independent experiments; in D–G, two-way ANOVA and Tukey’s multiple comparisons test; n = 3; data are mean ± SEM. Numbers of cells are indicated on histogram bars. Scale bar = 10 µm.