Use The Cali Technical Study Intermediate Spindle Assembly For Cytokinesis And Mitotic Exit Event Regulation

In eucaryotic cell, during anaphase, the mitotic spindle reorganizes in preparation for cytokinesis which happens after sister chromatids segregation. The central spindle, termed as an array of antiparallel microtubules, are bundled at their overlapping plus ends, regulates cleavage furrow initiation and the completion of cytokinesis.Failures in cytokinesis can lead to tetraploidy, a state that has for a long time been suspected to contribute to cancer formation. In our previous studies, fluorescence intensity analysis showed that there is compact relation beween central spindle and chromosome bridges. Cells with chromosome bridges leads to week and unstable central spindle at mid zone. Considering cells with chromosome bridges can regress the furrow and lead to failure of mitotic exit, we come up with a hypothesis that central spindle plays an important role in cytokinesis regulation. It is still unclear how the cleavage furrow respond to the central spindle in specific tempral manner, and we thus asked if the integrity of central spindle is required for mitotic exit, as well as the main molecular mechanism of them.Generally, cytokinesis is a brief process which is hard for us to catch with during the living cell observation research of cell mitosis. Chromophore-assisted laser inactivation (CALI) is a technique that can inactivate chromophore-binding protein specifically in a spatially and temporally defined manner. Here we use CALI technique to inactivate the central spindle in definite stage of cytokinesis to identify the role of the central spindle in a spatiotemporal pattern. In this study, we demonstrate that:(1) CALI inactivation of central spindle at anaphase B results in delay of cytokinesis, however CALI inactivation of central spindle at anaphase A leads to disordered the intracellular location of RhoA at cleavage furrow which induces the arrest of cytokinesis. We got the evidence of that the assembly of central spindle coordinated with contractile ring and can regulate the furrow formation at anaphase A onset.(2) CALI induced depletion of central spindle disordered the location of AuroraB and INCENP, which is considered as the cause of the inhibiton of chromosome decondensation. Furthermore, destruction of central spindle also induced the delay of degradation of CyclinB3. We have the hypothesis that the integrity of the central spindle structure is required for mitotic exit, and the structure possibly plays a key role of anaphase-telophase transition checkpoint, such as chromosome decondensation and nuclear envelope assembly.In conclusion, our study defines a new check point mechanism of anaphase-telophase transiton in animal cells that prevents tetraploidization by furrow regression in response to central spindle defect.