Stiffness-mediated genetic selection of cells

Abstract Studying a cell’s ability to sense and respond to mechanical cues has emerged as a field unto itself over the last several decades and is now appreciated by engineers and biologists alike. For most cell types investigated thus far, when cultured on soft substrates they have slower growth rates, smaller spread areas, lower traction, and lower migration rates than when cultured on stiff substrates. However, there is a critical gap in the literature. These prior studies, with two notable exceptions, have all taken cell lines previously propagated on plastic surfaces and assayed cell response to substrate stiffness over the course of a few days to a week. These studies, which have been seminal and established the field of cell mechanosensing, are limited to short term phenotypic responses. Taking an intellectual leap, this proposal will test a novel concept of cellular evolution driven by substrate stiffness. We hypothesize that the mechanical properties of the substrate will select for variants fit for growth on soft or stiff substrates out of a genetically heterogeneous population of cells, just as drug treatments select for genetically resistant cells. We will test this hypothesis by performing sustained culture of cells on soft and hard hydrogels. Across two aims we will quantify how cell phenotypes (growth, motility, metastasis) evolve as cells are cultured for increasing lengths of time on soft and stiff hydrogels, sequence genomic changes during this mechano-evolution, and alter the link between mechano-evolution and phenotypic consequences through targeting mechanosensors.