Redistribution of Golgi stacks and other organelles during mitosis and cytokinesis in plant cells

andreas.nebenfuehr{at}colorado.edu, Jennifer A. Frohlick, staeheli{at}spot.colorado.edu

Abstract

We have followed the redistribution of Golgi stacks during mitosis and cytokinesis in living tobacco BY-2 suspension culture cells by means of a green fluorescent protein-tagged soybean a-1,2 mannosidase, and correlated the findings to cytoskeletal rearrangements and to the redistribution of ER, mitochondria and plastids. Although the fusion protein is usually targeted to the cis-Golgi in interphase cells, distinct fluorescence in the ER in addition to the Golgi labeling is present in mitotic cells, indicating altered transport between ER and Golgi during cell division. In preparation for cell division the Golgi stacks redistribute to the perinuclear cytoplasm, the phragmosome. During metaphase, Golgi stacks aggregate near the spindle poles as well as in an equatorial region under the plasma membrane. This latter localization, the "Golgi belt", accurately predicts the future site of cell division, and thus forms a novel marker for this region after the disassembly of the preprophase band. During cytokinesis, this distribution changes and a higher density of Golgi stacks is found near the phragmoplast, the site of cell plate formation. These sites of preferential Golgi stack localization are specific for this organelle and largely exclude mitochondria and plastids, although some mitochondria can approach the phragmoplast. This segregation of organelles is first observed in metaphase and persists until completion of cytokinesis. Maintenance of the distinct localizations does not depend on intact actin filaments or microtubules. The redistribution of Golgi stacks during mitosis and cytokinesis is consistent with the hypothesis that Golgi stacks are repositioned to ensure equal partitioning between daughter cells as well as rapid cell plate assembly.

Video Sequences

QuickTime videos have been compressed with MPEG. If your computer can't play back these sequences you can download QuickTime software here.

3D-Stereo Reconstructions of Interphase and Metaphase Cells (Fig. 1)

Stacks of epifluorescence images were collected with 1 µm (interphase) or 0.5 µm spacing (metaphase) respectively and deconvolved. Spaces between images were interpolated and stereo 3D reconstructions were created in NIH image using Stereo-4D macros. With this setup the far end of the cells is difficult to visualize, thus the bottom of the cells appears fuzzy and not well defined.

MPEG Movie stereo image interphase cell

(a) Interphase Cell - Golgi stacks are evenly distributed throughout the cortical as well as the prinuclear cytoplasm.

MPEG Movie stereo image of metaphase cell

(b) Metaphase Cell - Golgi stacks are accumulated near the spindle poles. The cortical cytoplasm is largely depleted of Golgi stacks except for a narrow equatorial region, the "Golgi belt".

Time Lapse Video Sequence of Cytokinetic Cell (Fig. 4)

Fluorescent and DIC images of a single cell were taken at 30 s intervals. The cell was maintained in a perfusion camber with a continuous supply of aerated MS medium at a flow rate of 0.5 ml/min.

MPEG Movie time lapse of cytokinetic cell

0 min late anaphase - the chromosomes have reached their final destination at the site of the future daughter nuclei. During the rest of the sequence the upper nucleus moves down into the cell so that the focal plane ends up in the perinuclear cytoplasm. Note the accumulation of Golgi stacks in the cortical cytplasm on the right side of the cell. This Golgi belt marks the future site of cell division.

7-10 min telophase/early cytokinesis - the first sign of a forming cell plate between the daughter nuclei is a dark, straight line, slightly sloping down from left to right. Note how rapidly the phragmoplast and the cell plate form over a width of approx. 20 µm.

25-35 min change from barrel- to ring-shaped phragmoplast - the 'early' phragmoplast appears to break down. Note the accumulation of Golgi stacks adjacent to the phragmoplast. At the end of this phase the ring-shaped 'late' phragmoplast becomes apparent at the right end of the cell plate. (The corresponding part of the phragmoplast on the left side is not well developed at this stage, presumably due to the asymmetric position of the nucleus.)

40-65 min expansion of the ring-shaped phragmoplast - the 'late' phragmoplast moves towards the plasma membrane, continuously surrounded by Golgi stacks. Note the presence of Golgi stacks close to the more mature cell plate between the daughter nuclei.

65-94 min fusion of the cell plate with the plasma membrane - the ring-shaped 'late' phragmoplast (now visible on both sides of the cell) expands into the cortical region of the cell and disappears as the cell plate reaches the plasma membrane.

For further information please contact:

Andreas Nebenführ
University of Colorado
MCD Biology
Boulder, CO 80309-0347

phone: +1-303-492-8893
fax: +1-303-492-7744
email: andreas.nebenfuehr{at}colorado.edu
www: http://mcdb.colorado.edu/~nebenfue

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