VIFs defasciculate from bundles to interact with MTs. (A) 3D rendering of VIFs (blue) and MTs (gray) traced from the EM volume specified in Fig. 7 A. The ventral plasma membrane is shown in light gray. (B) Magnified inset of a VIF bundle and neighboring, minimally co-aligned MTs. (C) 3D reconstruction of a VIF bundle (blue) aligned with an MT (gray). VIFs at the bundle surface splay out, locally aligning with the MT (black arrowheads). (D) Scatter plot with marginal histograms indicating average VIF orientation relative to the nearest MT versus average VIF distance to the nearest MT. (E) Rendering of perinuclear VIFs color-coded by local alignment with MTs. VIF segments that are both proximity to and aligned with a neighboring MT (<100 nm NND and <20° relative orientation) are red. All other nonaligned segments are white. (F) Sum intensity projection of a center-aligned 3,500-nm MT trajectory. Scale bar = 50 nm. (Right) Orthogonal view of an MT center–aligned track. By aligning the stack around the MT centroid, we have computationally straightened the MT, allowing for visualization of the local neighborhood around a single MT. (G) MT-centered FIB-SEM slice with VIF cross sections highlighted by blue circles and distances from the MT delineated by the dashed lines. Scale bar = 50 nm; different z slices from this stack are represented in Fig. 7 C. (Right) Cartoon schematic relating the cross-sectional profile of MTs and vimentin to the expected appearance in the longitudinal axis. (H) 3D reconstruction of a computationally straightened MT (gray) and local vimentin filament bundle (blue). (I) Total number of vimentin filaments within 249 nm of the centroid of an MT over its 3,500-nm length. (J) Average distance of vimentin filaments from a single MT over its 3,500-nm length within the reconstruction volume. Line = mean, error bars = standard deviation. (K) Distribution of distance between MTs and their nearest vimentin neighbors. The line indicates a mean distance of 309 nm.