Nonthermal driving forces in cells revealed by nonequilibrium fluctuations

Abstract: The mechanical properties within living cells play a critical role in the adaptive regulation of their biological functions upon environmental and internal stimuli. While these properties exhibit nonequilibrium dynamics due to the thermal and nonthermal forces that universally coexist in actin-myosin-active proliferative cells, quantifying them within such complex systems remains challenging. Here, we develop a nonequilibrium framework that combines fluorescence correlation spectroscopy (FCS) measurements of intracellular diffusion with nonequilibrium theory to quantitatively analyze cell-specific nonthermal driving forces and cellular adaptability. Our results reveal that intracellular particle diffusion is influenced not only by common thermal forces but also by nonthermal forces generated by approximately 10-100 motor proteins. Furthermore, we derive a physical parameter that quantitatively assesses the sensitivity of intracellular particle responses to these nonthermal forces, showing that systems with more active diffusion exhibit higher response sensitivity. Our work highlights the biological fluctuations arising from multiple interacting elements, advancing the understanding of the complex mechanical properties within living cells.