Tomography of actin cytoskeletal networks

Structural biology research is increasingly focusing on unraveling structural variations at the micro-, meso-, and macroscale aiming at interpreting dynamic biological processes and pathways. Toward this goal, high-resolution transmission cryoelectron microscopy (cryo-EM) and cryoelectron tomography (cryo-ET) are indispensable, as these provide the ability to determine 3D structures of large, dynamic macromolecular assemblies in their native, fully hydrated state in situ. Underlying such analyses is the implicit assumption that specific structural states yield specific cellular outputs. The dependence on this structure-function paradigm is not unique to studies pertaining a particular pathway or biological process but it sets the foundation for all cell biological analyses of macromolecular assemblies. Yet, the paradigm still awaits formal proof. The field of high-resolution electron microscopy (HREM) is in dire need of establishing approaches and technologies to systematic and quantitative determining structure-function correlates in physiologically relevant environment. Here, using the actin cytoskeletal networks as an example, we will provide snapshots of current advances in defining the structures of these highly dynamic networks in situ. We will further detail some of the major stumbling blocks on the way to quantitatively correlate the dynamic state to network morphology in the same window of time and space.