engleski

Cryoelectron Tomography: Implications for Actin Cytoskeleton Research

Disclosing undistorted spatial organization of the actin cytoskeleton with molecular resolution is fundamental for understanding the cellular ultrastructure. Whereas important insights into the architecture of microfilament networks have been gained by negative staining, critical point drying, and freeze-fracturing methods, it is cryoelectron tomography that provides, for the first time, a comprehensive three-dimensional view of the intact actin cytoskeleton in situ. In particular, topological relationships such as microfilament three-dimensional proximity, angles at filament branching points, and modes of microfilament interaction with the plasma membrane can be visualized with an unprecedented accuracy using this technique. Further improvements are expected to bring the resolution into the realm of 2– 3 nm, where automatic pattern- recognition methods can be applied to identify actin-binding complexes. Combining cryoelectron tomography with ultra-fine immunolabeling and high-resolution fluorescence microscopy will make it possible to correlate structural data on the nanometer scale with molecular specificity and dynamical information.

actin; cytoskeleton; cryofixation; electron microscopy; tomography

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