Cryoelectron Tomography Reveals Nanoscale Organization of the Cytoskeleton and Its Relation to Microtubule Curvature Inside Cells

Abstract

Microtubules (MTs) are the most rigid elements of the cytoskeleton with invitro persistence lengths (Lp) in the range of 1-6mm. In cellular environments, however, MTs often appear strongly curved. This has been attributed to the forces acting upon them in situ where they are embedded in composite networks of different cytoskeletal elements. Hitherto, the nanoscale organization of these networks has remained largely uncharacterized. Cryo-electron tomography (cryo-ET) allowed to visualize and analyze the in situ structure of cytoskeletal networks in pristinely preserved cellular environments and at high resolution. Here, we studied the molecular organization of MTs and their interactions with the composite cytoskeleton in frozen-hydrated HeLa and P19 cells at different cell-cycle stages. We describe modulation of MT curvature correlated with the surrounding molecular architecture, and show that nanoscale defects occur in curved MTs. The data presented here contribute to constructing realistic models of cytoskeletal biomechanics.