On the Role of Src Family Kinases in Cyclic Strain Induced Cell Reorientation

Abstract

Many cells recognize mechanical signals from their environment which is of vital importance for multicellular organisms. Cyclic stretch is a prominent mechanical signal experienced by various cell types around blood vessels, within the lung epithelia or around the intestine. In response to this signal, some cells reorient their actin cytoskeleton and main cell axis almost perpendicular to the direction of stretch. Despite the vital necessity of cellular adaptation to cyclic stretch, only incomplete ideas exist about the underlying mechanosensoric signal cascade. In this project we cultivated mouse embryonic fibroblasts in flexible cell culture chambers fabricated from silicone where they adhered via focal adhesions (FAs). Cyclic stretching of these chambers during culture provided well-defined mechanical stimuli to the cells. After straining, cells were fixed, stained for F-actin and adjacent micrographs of large areas of the sample were taken. For all cells imaged (some hundreds per chamber) we determined the preferential orientation of the actin fibers by digital image processing. This results in data sets with high statistical significance. Because FAs have been shown to be strongly influenced by the Src-family kinases and have been furthermore implied in mechanosensing we examined the influence of Src-family kinase knockout on cellular reorientation upon cyclic stretch. We found that lacking phosphorylation leads to strongly diminished cellular reorientation.