The uncoupling phenotype and Rvs167 misrecruitment are due to an increase in PM tension. (A) Correlation between the kinetics of the increase in PM tension upon TORC2 inhibition and appearance of the various endocytosis phenotypes. PM tension was monitored through the lifetime of the FliptR probe measured by FLIM, and error bars represent the propagated error of mean values calculated from three independent experiments (with n > 10 cells). (B) Sla1-GFP and Abp1-mCherry kymograph recorded across a FPS1Δ cell upon hypo-osmotic shock. Dot lines represent the PM. (C) Time series of merged Sla1-GFP and Abp1-mCherry signals in FPS1Δ cells upon hypo-osmotic shock. The percentage of cells presenting clustered endocytosis sites are indicated before and after the shock and were calculated from three independent experiments (**, P < 0.01). (D) Rvs167-GFP and Abp1-mCherry kymograph along the PM of a FPS1Δ cell upon hypo-osmotic shock. (E) Average intensity of Rvs167-GFP foci before and after hypo-osmotic shock. Error bars represent SD of mean values calculated for n = 100 events from three independent experiments (****, P < 0.0001). (F and H) Abp1-mCherry and either Sla1-GFP (F) or Rvs167-GFP (H) kymographs recorded as indicated along the PM of a TOR1-1 AVO3Δ^CT cell, pretreated with Rapamycin for 1 h, upon PalmC injection. (G) Average intensity of Rvs167-GFP foci before and after PalmC treatment in a TOR1-1 AVO3Δ^CT cell, pretreated with Rapamycin for 1 h. Error bars represent SD of mean values calculated for n = 100 events from three independent experiments (****, P < 0.0001). Scale bars, 5 µm. Source data are provided in Table S1.