Inhibition of annexin A2 cleavage decreases annexin A2 ⁺ MV secretion during membrane repair. (A) EV production index from ANXA2-Nluc cells treated with the indicated combinations of 200 ng/ml SLO and 20 μM calpain inhibitor, ALLN. Error bars indicate three experimental replicates. (B) Immunoblot analysis of lysates from WT cells (wt), annexin A2 KO cells, and annexin A2 KO cells expressing WT annexin A2-Nluc (KO + WT) or ANXA2[P20D, P21D]-Nluc (KO + Mt). (C) EV production index from WT ANXA2-Nluc (wt) or ANXA2[P20D, P21D]-Nluc (Mt) cells treated with or without SLO. Error bars indicate three experimental replicates. (D) Quantification over time of the repair scab intensity from FM1-43–stained (2.5 μM) annexin A2 KO cells rescued with ANXA2-mScarlet (wt) or ANXA2[P20D, P21D]-mScarlet (Mt). Error bars indicate six experimental replicates. (E) Confocal micrographs of eight laser-ablated annexin A2 KO cells rescued with either WT ANXA2-mScarlet (wt) or ANXA2[P20D, P21D]-mScarlet (Mt). Images are 2 min, 30 s after ablation. Scale bars: 5 μm. (F) Schematic depicting the current model of plasma membrane repair and annexin⁺ MV secretion. EPVI, EV production index. Source data are available for this figure: SourceData F6.