Extended Data Fig. 2: Assessment of the role of PIEZO1 and PIEZO2 in strain-induced plasma membrane rupture.
From: NINJ1 regulates plasma membrane fragility under mechanical strain
a, LDH release of PIEZO1, PIEZO2 knock-down and control HeLa cells. n = 4–8 trials per group. b, Percentage of DRAQ7+ cells of PIEZO1, PIEZO2 knock-down and control HeLa cells. n = 4–8 trials per group from ~150 cells of each group. 2-way ANOVA followed by Bonferroni corrections. n.s., not significant versus Scrambled. c, d, LDH release and the percentage of DRAQ7+ cells from the HEK-293T transiently-transfected with human PIEZO1 and PIEZO2. Vector-transfected cells, as well as TRE3G-NINJ1 HEK-293T stable cells under 100 ng ml−1 Dox induction were used as controls. Assay was conducted 24 h after transfection. n = 3 trials for each group, 2-way ANOVA followed by Bonferroni corrections. ** p <0.01 versus Vector at the same strain level. e, The lysis tension of PIEZO1 and NINJ1-expressing vesicles. Owing to the drastically larger size of PIEZO1 (38 transmembrane segments, 287 kDa, compared to 3 transmembrane and 16 kDa for NINJ1), protein levels were derived from fluorescence intensity of mCherry fused to C-terminal of PIEZO1, corrected for number of transmembrane segments to reflect the footprint, and plotted as size-corrected molecular density on the membrane in arbitrary units (A.U.) for a fair comparison with NINJ1. 30 vesicles harbouring WT NINJ1 and 13 vesicles with PIEZO1 were measured. 10 additional vesicles that only overexpress GPI-eGFP were included as the zero intensity points for both NINJ1 and PIEZO1. Unless otherwise noted, all data are presented as mean ± s.e.m.
