The calcium ions, as an important second messenger, plays key roles in many cell functions. The present study is aimed to examine the functional roles of calcium and its downstream components in the regulation of cell division. In the first part of this research, we used a calcium sensitive fluorescent probe and a confocal microscope to characterize the calcium signals in association with cell division in zebrafish embryos. We found that there were multiple types of calcium transients appearing at the different stages of cell cleavage. Judging from the spatial and temporal correlation of these calcium transients with the mitotic events, these calcium signals appear to be involved in several mitotic functions, including positioning the cleavage furrow, furrow extension, and furrow deepen...[ Read more ]

The calcium ions, as an important second messenger, plays key roles in many cell functions. The present study is aimed to examine the functional roles of calcium and its downstream components in the regulation of cell division. In the first part of this research, we used a calcium sensitive fluorescent probe and a confocal microscope to characterize the calcium signals in association with cell division in zebrafish embryos. We found that there were multiple types of calcium transients appearing at the different stages of cell cleavage. Judging from the spatial and temporal correlation of these calcium transients with the mitotic events, these calcium signals appear to be involved in several mitotic functions, including positioning the cleavage furrow, furrow extension, and furrow deepening and maintenance. Studies by injecting calcium signal inhibitors further supported this interpretation. In the second part of the research, we examined the involvement of a calcium binding protein, calmodulin. in cell division. Here we used a GFP-fusion gene method and a highly sensitive imaging system to study the dynamic redistribution of calmodulin (CaM) in living mammalian cells. Our results indicated that GFP-CaM underwent specific patterns of redistribution during cell division. At late anaphase, a fraction of GFP-CaM was found to concentrate at the cell cortex of the equator where the cleavage furrow was to appear. The timing of this cortical CaM elevation coincided with the onset of cytokinesis. In addition, we utilized an activity-sensitive fluorescent probe (TA-CaM) to study the spatial-specific activation of CaM during cell division. The cortical fraction of CaM beneath the cleavage furrow was found to be selectively activated during cytokinesis. These findings strongly suggest that in addition to the calcium ions, CaM also plays an important role in regulating cell division. In the third part of this research, we used electroporation and microinjection techniques to introduce specific inhibitors into living cells to examine the functional roles of several downstream components of the calcium signaling pathway. Our results suggest that the calcium signal may be involved in the regulation of metaphase-anaphase transition through the Ca²⁺ /CaM-dependent protein kinase II. At cytokinesis, calcium signal is required for the formation and maintenance of the actomyosin contractile band. Taken all together, these findings support a hypothetical model that was proposed based on a number of very recent studies. Details of this model are given in Chapter 1.