Experimental Characterization And Physical Modeling Of Myosin At The Cellular And Tissue Scales

Cells have the ability to sense peripheral mechanical stimuli and convert them into physiological and biochemical signals.This ability enables cells to adapt to the surrounding physical microenvironment through remodeling of cytoskeletal microstructure and activates multiple signaling pathways,and alter gene expression.These phenomena have two important processes:mechanosensing and mechanotransduction.In these processes,mechanical signals(forces or deformations)are transferred from the cell's external environment to proteins and organelles within the cell.Actin cytoskeleton in the cellular cortex is a major player in sensing mechanical stimuli and activating downstream signaling pathways.A quantitative understanding of the mechanosensing mechanisms of individual cytoskeletal proteins is essential for dissecting the role of these proteins in various signaling pathways and physiological systems.At the cellular level,the results of mechanosensing are often expressed in the deformations of cells.Cells can be spherical,ellipsoid,butterfly,or columnar.These shapes change and transform under the action of external forces and internal forces in the cell.The maintenance of cell shape and its changes are largely determined by the cytoskeleton.Among the three known cytoskeletal systems,microfilaments play an important role in controlling cell morphology.At the molecular scale,the deformations of filaments include intermolecular deformations and intramolecular deformations.Intramolecular deformations depend on specific molecular structures,such as peptide sequences,folded states,and combinations of dimers(or oligomers).So far,a quantitative interpretation of the protein behaviors based on the deformation of cells under compression is still absent.For example,the characterization of the mechanosensory behaviors of myosin Hs in published literature was more or less qualitative since the exact amount of compression on each cell was not well defined and the dynamic responses of the proteins over time were not characterized.Moreover,how the anisotropic deformations in a compressed cell affect the spatial distribution of the mechanosensitive proteins has not been fully investigated.Furthermore,the study on mechanobiology feedback loop of myosin is not sufficient.In this study,we used several well-studied myosin ? mutants to test the ability of the compression assay for the characterizations of the mechanosensory responses of cytoskeletal proteins in many cells at one time.Based on elasticity theories,we calculated the tensions and strains along the cell cortex.Using this information as input,we simulated the mechanosensory accumulation of myosin ?s and quantitatively reproduced the experimental observations.Combining the compression assay with confocal microscopy,we monitored the polarization of myosin ?oligomers at the sub-cellular level.The polarization was found to be largely determined by the ratio of the two principal strains of the cellular deformations.At last,we showed that this technique could be used for the investigation of other mechanosensory proteins.Our results demonstrated that the compression assay is a fast low-cost technique for the high throughput screening of mechanosensitive cytoskeletal proteins in cells.Moreover,a mechanobiology model based on myosin mechanobiology feedback loop is studied and established.The mechanical signal feedback loop and myosin biochemical signal pathway are fully coupled.Mechanical signals affect the reaction energy barrier during myosin inactivation in the form of mechanical energy.At the same time,the local concentration of myosin changes the stress in the corresponding region of the cell.Then the oscillatory behavior of cell area and myosin concentration in cell layer with different topological morphologies under simple stress was calculated by using the mechanobiology model.The results show that the model can completely reproduce the oscillatory characteristics of cell area and myosin concentration observed in drosophila embryo development.At the same time,the model also reveals that the mechanical signal can make the system in the stable state break away from the stable region,produce oscillation,and evolve into the next stable state.The changing mechanical signal makes the system unable to reach a stable state,but in a state of long-term oscillation.In addition,the results show that the mechanical signal can also affect the waveform and period of the oscillation.