Spindle Architectural Features Must Be Considered Along With Cell Size to Explain the Timing of Mitotic Checkpoint Silencing

Abstract

Mitosis proceeds through a defined series of events that is largely conserved, but the amount of time needed for their completion can vary in different cells and organisms. In many systems, mitotic duration depends on the time required to satisfy and silence the spindle assembly checkpoint (SAC), also known as the mitotic checkpoint. Because SAC silencing involves trafficking SAC molecules among kinetochores, spindle, and cytoplasm, the size and geometry of the spindle relative to cell volume are expected to affect mitotic duration by influencing the timing of SAC silencing. However, the relationship between SAC silencing, cell size, and spindle dimensions is unclear. To investigate this issue, we used four DLD-1 tetraploid (4N) clones characterized by small or large nuclear and cell size. We found that the small 4N clones had longer mitotic durations than the parental DLD-1 cells and that this delay was due to differences in their metaphase duration. Leveraging a previous mathematical model for spatiotemporal regulation of SAC silencing, we show that the difference in metaphase duration, i.e., SAC silencing time, can be explained by the distinct spindle microtubule densities and sizes of the cell, spindle, and spindle poles in the 4N clones. Lastly, we demonstrate that manipulating spindle geometry can alter mitotic and metaphase duration, consistent with a model prediction. Our results suggest that spindle size does not always scale with cell size in mammalian cells and cell size is not sufficient to explain the differences in metaphase duration. Only when a number of spindle architectural features are considered along with cell size can the kinetics of SAC silencing, and hence mitotic duration, in the different clones be explained.