Cultivated animal cells often display prominent actin bundles, also known as stress fibers, throughout their cytoskeleton. These bundles are thought to provide mechanical tension and might take part in cellular mechanosensing. To test the functions of these bundles we cultivated endothelial cells on soft silicone elastomer films. Strain was applied to cells by either globally stretching the whole elastomer film [1] or by locally deforming it with a microtool [2]. In both methods actin bundles are stretched. Resulting molecular rearrangements were studied by immuncytochemistry and by live cell imaging of cells transfected with zyxin, alpha-actinin or VASP fused to fluorescent proteins. We observed strain-dependent formation of zyxin-rich striations within actin bundles similar to [3]. Colocalization of zyxin, alpha-actinin and vinculin on actin bundles was quantified by digital image processing and the time course of dissociation was determined. From these data we conclude that zyxin and VASP act in close concert while alpha-actinin plays a different role.[1] Faust et al PLoSOne 6 (2011) e28963[2] Kirchenbuchler et al J. Phys. Cond. Mat. 22 (2010) 194109[3] Hoffman et al. Mol. Biol. Cell 23 (2012) 1846-1859