Mechanotransduction associated to lysosome localization and dynamics during B cell activation. (A) Representative images of fixed B cells interacting with soft or stiff substrates containing BCR⁺ ligands at different time points. LAMP1 is shown in green, and the cell border (white dashed outline) was defined by F-actin phalloidin staining. Scale bar: 10 μm. (B) Top: Schematics depicting central and peripheral areas of the IS where LAMP1⁺ lysosomes were quantified. (see Materials and methods). Bottom: Represents the MFI percentage of LAMP1⁺ lysosomes in each region. (C) Number of lysosomes at the IS from cells shown in A. (D) Representative images of MR signal in B cells seeded on soft or stiff substrates containing BCR⁺ ligands. (E) MR fluorescence intensity through Z stacks for cells displayed in D. (F) Representative time-lapse images of B cells expressing cathepsin-RFP activated on stiff or soft conditions. (G and H) Displacement and mean speed of lysosome tracks from cells detailed in F. Data shown consider n ≥ 30 cells pooled from N = 3 independent experiments. Scale bars are 10 μm. (B and C): Two-way ANOVA with Sidak’s multiple comparison test. (G and H): t test. P values illustrated with asterisks are ** <0.01, *** <0.001, and **** <0.0001. Error bars are mean ± SEM.