Final Report Summary - CELLMECHANOCONTROL (The physical basis of cellular mechanochemical control circuits) The project “The physical basis of cellular mechanochemical control circuits” aims to understand how cells integrate mechanical signals in their regulatory circuitry. The approach integrates several aspects of this problem. First, a quantitative understanding of cellular mechanical properties had to be achieved. For this purpose we combined micromechanical measurements, summarized under the term “microrheology”, with physical modeling of the complex geometry and material properties of cells. We have established a microrheology method using optical trapping and laser–interferometric displacement- and force-detection to observe thermal fluctuations and to be able to exert controlled force at the same time. Second, cellular force generation needed to be understood. Cells generate forces through specialized mechano-enzymes (motor proteins) interacting with cytoskeletal polymer networks. The structures in which motors and polymer networks are organized vary from cell to cell and range from relatively disordered cortical actin network with non-muscle myosin motors to highly organized sarcomeric interdigitating actin and myosin structures in heart and skeletal muscle cells. We have observed such subtle transitions in structural and dynamic order in a model system of fully functional and active cell extract enclosed in emulsion droplets where it spontaneously forms a cortical cytoskeletal layer that possesses many properties of the real cell cortex. We have in this project also developed statistical methods to diagnose non-equilibrium fluctuation both in disordered and in ordered systems, based on the breaking of time-reversal symmetry or (equivalently) the breaking of detailed balance. This method can easily be applied to microscopic video recordings of cells and can give unambiguous answers. A further unique tool that we have developed in the frame work of the project are novel fluorescent probes, based on single-walled carbon nanotubes. These nanoparticles fluoresce in the near infrared, a wavelength range that is ideal for biological imaging due to the absence of background fluorescence and scattering, and they are highly photostable, so that individual particles can be tracked for long times. We have successfully incorporated these probes in cells and can specifically tag proteins in the cells. Using these probes, we could show that cells use myosin motor activity to constantly stir themselves, thus speeding up intracellular transport and avoiding traffic jams. Initial experiments have shown that the probes can also enter the cell nucleus where we can now use them to report on transcriptional dynamics.