Three-dimensional analysis of syncytial-type cell plates during endosperm cellularization visualized by high resolution electron tomography

The three-dimensional architecture of syncytial-type cell plates in the end osperm of Arabidopsis has been analyzed at similar to6-nm resolution by mea ns of dual-axis high-voltage electron tomography of high-pressure frozen/fr eeze-substituted samples. Mini-phragmoplasts consisting of microtubule clus ters assemble between sister and nonsister nuclei. Most Golgi-derived vesic les appear connected to these microtubules by two molecules that resemble k inesin-like motor proteins. These vesicles fuse with each other to form hou rglass-shaped intermediates, which become wide (similar to 45 nm in diamete r) tubules, the building blocks of wide tubular networks. New mini-phragmop lasts also are generated de novo around the margins of expanding wide tubul ar networks, giving rise to new foci of cell plate growth, which later beco me integrated into the main cell plate. Spiral-shaped rings of the dynamin- like protein ADL1A constrict but do not fission the wide tubules at irregul ar intervals. These rings appear to maintain the tubular geometry of the ne twork. The wide tubular network matures into a convoluted fenestrated sheet in a process that involves increases of 45 and 130% in relative membrane s urface area and volume, respectively. The proportionally larger increase in volume appears to reflect callose synthesis. Upon fusion with the parental plasma membrane, the convoluted fenestrated sheet is transformed into a pl anar fenestrated sheet. This transformation involves clathrin-coated vesicl es that reduce the relative membrane surface area and volume by similar to 70%. A ribosome-excluding matrix encompasses the cell plate membranes from the fusion of the first vesicles until the onset of the planar fenestrated sheet formation. We postulate that this matrix contains the molecules that mediate cell plate assembly.