Visualization on intercellular movement of chromatin in intact living anthers of transgenic tobacco expressing histone 2B-CFP Fusion Protein

Intercellular movement of chromatin termed cytomixis is a conspicuous phenomenon reported in both vegetative and reproductive tissues in many plant species, but has not widely been accepted as a normal cellular process. To further address this problem, morphologically normal and fertile transgenic tobacco plants expressing fusion protein of Arabidopsis histone 2B-cyan fluorescent protein (AtH2B-CFP) were constructed, which gave CFP fluorescence in the cell nuclei of both vegetative and floral tissues. The expression of H2B-CFP had no effect on the regulation of cell cycle. The dynamic process of separation of CFP-tagged chromosomes during mitotic anaphase in intact living root tip cells could be observed. In the epidermal and endothelial cells of intact living anther, their nuclei were not round-shaped but with many surface protuberances. These protuberances and the overall shape of nucleus were always in a state of active movement, and cytomixis was observed to occur between some cells, the dynamic process of which was recorded by time-laps imaging, which were very similar to the fixed pictures observed previously with light and electron microscope. We found that squash and enzymatic digestion completely inhibited all the visible nuclear movement, illustrating that harmful factors such as enzyme digestion and mechanical damage repress rather than stimulate cytomictic activity, indicating that cytomixis is a normal cellular process not caused by fixation or mechanical damage.