Abstract
To survive in a mechanically active environment, cells must adapt to variations of applied membrane tension. A collagen-coated magnetic bead model was used to apply forces directly to the actin cytoskeleton through integrin receptors. We demonstrate here that by a calcium-dependent mechanism, human fibroblasts reinforce locally their connection with extracellular adhesion sites by inducing actin assembly and by recruiting actin-binding protein 280 (ABP-280) into cortical adhesion complexes. ABP-280 was phosphorylated on serine residues as a result of force application. This phosphorylation and the force-induced actin reorganization were largely abrogated by inhibitors of protein kinase C. In a human melanoma cell line that does not express ABP-280, actin accumulation could not be induced by force, whereas in stable transfectants expressing ABP-280, force-induced actin accumulation was similar to human fibroblasts. Cortical actin assembly played a role in regulating the activity of stretch-activated, calcium-permeable channels (SAC) since sustained force application desensitized SAC to subsequent force applications, and the decrease in stretch sensitivity was reversed after treatment with cytochalasin D. ABP-280-deficient cells showed a > 90% increase in cell death compared with ABP-280 +ve cells after force application. We conclude that ABP-280 plays an important role in mechanoprotection by reinforcing the membrane cortex and desensitizing SACs.
Highlights
Nearly 2-fold higher proportion of actin in filamentous form [7]
Consistent with these data we have demonstrated that fibroblasts undergo localized actin assembly during isolated force application through focal adhesion complexes [6]
force application (F-actin) Accumulation—We studied force-induced actin assembly locally at the force transfer sites by isolating the proteins associated with collagen-coated magnetic beads
Summary
Nearly 2-fold higher proportion of actin in filamentous form [7]. This example indicates that cells are able to sense and adapt to environmental tension in part through cytoskeletal adaptations. Inhibitors—We assessed the dependence of actin rearrangement on calcium ions by incubating cells with BAPTA/AM at 3 M for 45 min at 37 °C prior to force application. F-actin Accumulation—We studied force-induced actin assembly locally at the force transfer sites by isolating the proteins associated with collagen-coated magnetic beads.
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