QUANTITATIVE ANALYSIS OF CHANGE IN GEOMETRY OF MICROTUBULES IN PLANT CYTOKINESIS

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Cytokinesis in Arabidopsis meristem cells has been investigated with a 3D resolution of about 7nm by electron tomography of high-pressure frozen/freeze-substituted samples. Our data demonstrate that cell plates assemble from three distinct structures: phragmoplast initials, a solid phragmoplast, and a ring-shaped phragmoplast. Phragmoplast initials arise during late anaphase from clusters of polar microtubules (MTs) at the cell's midplane. Cell plate assembly sites initiate from a ribosome-excluding, cell plate assembly matrix (CPAM) and Golgi derived vesicles. The CPAM seems to be responsible for both regulating cell plate growth and anchoring MT (+) ends. This association directs vesicles to the CPAM, and thereby to the growing cell plate. After vesicles accumulate within the CPAM, cell plate formation starts with the tethering of vesicles by exocyst-like complexes. Vesicle fusion produces hourglass-shaped vesicles that are stretched to dumbbells by dynamin spring expansion. At the same time, phragmoplast initials and their CPAMs expand laterally to make the solid phragmoplast. Later arriving vesicles fuse with the bulbous ends of the dumbbells, producing a tubulo-vesicular membrane network (TVN). Upon completion of the TVN, the CPAM and MTs disassemble and then reform in a peripheral ring. This creates the centrifugally expanding peripheral cell plate growth zone, which leads to cell plate fusion with the cell wall. Simultaneously, the central TVN matures into a tubular network (TN), and ultimately into a planar fenestrated sheet (PFS), through the removal of membrane via clathrin-coated vesicles and by callose synthesis. Small secondary CPAMs with anchored MTs arise de novo over large fenestrae to focus local growth to these regions. When all of the fenestrae are closed, the new cell wall is complete. Segui-Simarro, J.M., J.R. Austin II, E.A. White and L.A. Staehelin (2004) Plant Cell 16:836-856.