Abstract
In higher plants, vacuoles increase their volume according to cell enlargement and become large vacuoles that occupy most of the cell volume. To observe the vacuolar dynamics in tobacco BY-2 cells and Physcomitrella patens, we have established transgenic cell lines expressing GFP-AtVam3p fusion protein. We have also developed software package named REANT for three-dimensional reconstruction of vacuoles. We adopted a new algorithm based on the combinations of region correspondence techniques and contour-connecting approaches. As a result, we found unique vacuolar structures including TVM (tubular structure of vacuolar membrane) in mitotic cell and vacuolar networks developed in elongating miniprotoplasts. The experiments using the inhibitor for actin microfilaments indicated that the microfilaments were indispensable for the development and the maintenance of these vacuolar structures. Similar tubular vacuoles were also found in the protonemal cells of P. patens, however, the structural dependency of vacuoles on cytoskeletons was different from BY-2 cell.
