An Approach to Visualize the Deformation of the Intermediate Filament Cytoskeleton in Response to Locally Applied Forces

The intermediate filament (IF) cytoskeleton plays an important role in integrating biomechanical pathways associated with the actin and microtubule cytoskeleton. Vimentin is a type III IF protein commonly found in fibroblast cells and plays a role in transmitting forces through the cytoskeleton. Employing simultaneous laser scanning confocal and atomic force microscopy (AFM), we developed a methodology to quantify the deformation of the GFP-vimentin-labeled IF cytoskeleton as a function of time in response to force application by the AFM. Over short times (seconds), IFs deformed rapidly and transmitted force throughout the entire cell in a highly complex and anisotropic fashion. After several minutes, mechanically induced displacements of IFs resemble basal movements. In well-adhered cells the deformation of IFs is highly anisotropic as they tend to deform away from the longitudinal axis of the cell. This study demonstrates that simultaneous AFM and LSCM can be employed to track the deformation and dissipation of force through the IF cytoskeleton.