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Plakoglobin Is Required for Effective Intermediate Filament Anchorage to Desmosomes
Devrim Acehan, Christopher Petzold, Iwona Gumper, David D Sabatini, Eliane J Müller, Pamela Cowin and David L Stokes
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Influence of different methods for EM preparation. (a-d) Cells scraped from culture dishes after a preliminary chemical fixation. PG+/+ cells (a) retain intact epithelial sheets, whereas PG-
/-
cells (b) are greatly disrupted by this procedure. Higher magnification images of these PG+/+ cells (c) reveal numerous desmosomes (circles), whereas very few desmosomes are visible in PG-
/-
cells (d). (e-f) When grown and prepared on Transwell membranes, these differences are less apparent, though PG-
/-
cells (f) exhibit a wider intercellular gap and a larger number of filopodia. Arrowheads indicate the cell borders. Scale bars are 1
m.
Supplementary Figure S2 (jpeg 1,052K)
Tomographic images of desmosomes. (a) Extended, "tandem" desmosomes are prevalent in PG- /- cells. This tomographic slice shows the underdeveloped cytoplasmic plaque and that the intermediate filament bundles (segmented in blue) do not connect to the desmosomes. Ribosomes (dark globular densities) and oblique sections of microtubules (arrowhead) can be traced throughout the tomogram. (b) Desmosomes from PG+/+ cells reveal connections to a large bundle of intermediate filaments. This is a slice from the tomographic data set with partial segmentation of the intermediate filament network (blue). A dense, unsegmented bundle of intermediate filaments is running vertically and frays into smaller bundles or individual filaments, which connect with the cytoplasmic plaque of each desmosome visible in the image. Segmentation is based on all the slices that makeup the tomogram, only one of which is shown in gray-scale here. Ribosomes and other large cellular assemblies are excluded from the cytoplasmic plaque of the desmosomes. Scale bars correspond to 100nm.
Supplementary Figure S3 (jpeg 915K)
A gallery of tomographic slices from PG+/+ and PG- /- cells. Whereas wild type desmosomes consistently display extensive connections with the intermediate filament network (a-c), PG deficient desmosomes have poorly formed outer and inner dense plaques and do not connect to intermediate filaments (d-f). Scale bars are 100 nm.
Supplementary Movie S1 (mpeg 3,899K)
Animated series of slices through the tomographic volume in Figure 5a.
Supplementary Movie S2 (mpeg 4,658K)
Animated series of slices through the tomographic volume in Figure 5d.
Supplementary Movie S3 (mpeg 4,729K)
Animated series of slices through the tomographic volume in Figure 7a.