Figure 1.
Extending Surface Morphometrics to measure lipid bilayer thickness within tomograms. (A) Tomograms were reconstructed and segmented to produce voxel segmentations that labeled distinct cellular membranes. Voxel segmentations were processed through the Surface Morphometrics pipeline (Barad et al., 2023) to generate triangulated surface meshes. The tomogram density was interpolated systematically from the midpoint of each triangle on the surface mesh, following the normal vector in both the positive and negative directions (dashed arrows). These measurements are used to generate a voxel line scan, which reveals two peaks corresponding to the densities associated with the PHG (dashed vertical lines). We calculated the distance between these peaks as a readout of membrane thickness. Line scans that are extended beyond the lipid bilayer show additional peaks representing protein macromolecules (pink arrowhead). (B) Triangle mesh models for both IMM (orange) and OMM (lavender) are overlaid on top of the binned tomogram reconstruction, which is of sufficient resolution to distinguish lipid head groups visually. The meshes can be used to generate local vectors for density line scan analysis within the tomogram. (C) Line scans generated by sampling along the normal vectors can be combined across a surface to generate a smooth plot with peaks corresponding to the two head groups of the bilayer (blue line). We fit the curve with a dual Gaussian model (orange dashed line, individual Gaussians, pink and green) and used the peak-to-peak distance as the bilayer distance. (D) Local thickness variation analysis was determined via a local weighted averaging scheme, in which the line scan of the central triangle (red) is combined with neighboring triangles up to 12 nm away with weights that decrease with distance from the center. (E) Using per-triangle thickness measurements, we identified the differences in the distribution of thicknesses within different membranes, as depicted in a triangle area–weighted histogram, whose y axis measures the relative area of membrane in each thickness bin.

Extending Surface Morphometrics to measure lipid bilayer thickness within tomograms. (A) Tomograms were reconstructed and segmented to produce voxel segmentations that labeled distinct cellular membranes. Voxel segmentations were processed through the Surface Morphometrics pipeline (Barad et al., 2023) to generate triangulated surface meshes. The tomogram density was interpolated systematically from the midpoint of each triangle on the surface mesh, following the normal vector in both the positive and negative directions (dashed arrows). These measurements are used to generate a voxel line scan, which reveals two peaks corresponding to the densities associated with the PHG (dashed vertical lines). We calculated the distance between these peaks as a readout of membrane thickness. Line scans that are extended beyond the lipid bilayer show additional peaks representing protein macromolecules (pink arrowhead). (B) Triangle mesh models for both IMM (orange) and OMM (lavender) are overlaid on top of the binned tomogram reconstruction, which is of sufficient resolution to distinguish lipid head groups visually. The meshes can be used to generate local vectors for density line scan analysis within the tomogram. (C) Line scans generated by sampling along the normal vectors can be combined across a surface to generate a smooth plot with peaks corresponding to the two head groups of the bilayer (blue line). We fit the curve with a dual Gaussian model (orange dashed line, individual Gaussians, pink and green) and used the peak-to-peak distance as the bilayer distance. (D) Local thickness variation analysis was determined via a local weighted averaging scheme, in which the line scan of the central triangle (red) is combined with neighboring triangles up to 12 nm away with weights that decrease with distance from the center. (E) Using per-triangle thickness measurements, we identified the differences in the distribution of thicknesses within different membranes, as depicted in a triangle area–weighted histogram, whose y axis measures the relative area of membrane in each thickness bin.

This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal