The Architecture of Traveling Actin Waves Revealed by Cryo-Electron Tomography

Marion Jasnin, Florian Beck, Mary Ecke, Yoshiyuki Fukuda, Antonio Martinez-Sanchez, Wolfgang Baumeister, Günther Gerisch - Department of Molecular Structural Biology, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany

Actin waves are dynamic supramolecular structures involved in cell migration, cytokinesis, adhesion, and neurogenesis. Although wave-like propagation of actin networks is a widespread phenomenon, the actin architecture underlying wave propagation remained unknown. In situ cryo-electron tomography of Dictyostelium cells unveils the wave architecture and provides evidence for wave progression by de novo actin nucleation. Subtomogram averaging reveals the structure of Arp2/3 complex-mediated branch junctions in their native state, and enables quantitative analysis of the 3D organization of branching within the waves.

How Amira-Avizo Software is used

The 3D reconstruction with a final pixel size of 1.684 nm was obtained by weighted-back projection using IMOD and rendered in 3D using Amira. Filaments were automatically segmented using the actin segmentation package in Amira.[…] Binned twice tomograms were subjected to nonlocal-means filtering using Amira software provided by Thermo Fisher Scientific. Actin filaments were traced by an automated segmentation algorithm based on a generic filament as a template, with a diameter and length of 8 and 42 nm, respectively. To reduce background noise, short filamentous structures with lengths below 70 nm (or 40 nm in Figure 7) were filtered out.