Rapid evolution by clonal selection within populations of fibroblasts propagated on a novel soft substrate

Mechanical properties such as substrate stiffness are a ubiquitous feature of a cell9s ecological and evolutionary context, and may be a significant source of natural selection. Many types of animal somatic cells exhibit canonical phenotypic plasticity when grown on substrates of differing stiffness, both in vitro and in vivo. Whether such plasticity is a multivariate optimum due to hundreds of millions of years of animal evolution, or instead is a compromise between conflicting selective demands, is unknown. We addressed these questions by means of experimental evolution of replicate populations of mouse fibroblasts propagated for ~90 cell generations on soft or stiff substrates. The ancestral cells grow twice as fast on stiff substrate as on soft substrate, and exhibit the canonical phenotypic plasticity. Soft-selected lines derived from a genetically diverse ancestral population increased growth rate on soft substrate to the ancestral level on stiff substrate, and evolved the same multivariate phenotype. Conversely, growth rate and phenotypes did not change in cell populations derived from clonal cells. These results imply that the changes were the result of genetic evolution and not phenotypic plasticity. Whole-transcriptome analysis revealed consistent differentiation between ancestral and soft-selected populations, and that both emergent phenotypes and gene expression tended to revert in the soft-selected lines. However, the selected populations appear to have achieved the same phenotypic outcome by means of at least two distinct transcriptional architectures related to mechanotransduction and proliferation.