Studies on premature centriole disengagement during a prolonged mitotic arrest

Abstract

Centrioles duplicate and segregate in a tight link to the cell cycle. During S phase, a new centriole grows orthogonally from preexisting centriole and then establishes engagement. At the end of mitosis, paired mother and daughter centrioles eventually disengaged from each other, which allow mother centriole to be able to duplicate in the following S phase. Disruption of the centriole cycle often results in aneuploidy and cancers. Here, I report that centrioles are prematurely disengaged when a cell is arrested at M phase for a period of time. Premature centriole disengagement still occurs in mitotic cells of which the separase activity is reduced by depletion of CDC20 or separase itself. However, centrioles remain engaged with treatment of BI2536, a PLK1 inhibitor, suggesting that the PLK1 activity is critical even for premature centriole disengagement in M-phase-arrested cells. I also observed that SAS6 is displaced in prematurely disengaged daughter centrioles, suggesting that SAS6 displacement precedes premature centriole disengagement. BI2536 inhibits the SAS6 displacement as well as premature centriole disengagement. Furthermore, the centriole disengagement rate was reduced in M-phase-arrested cells with the centriole-directed SAS6-PACT protein. Finally, I observed reduction of the centriole disengagement rate in M-phase-arrested cells with a higher dose of nocodazole and taxol, and, therefore, whose cellular microtubules were completely disrupted. Furthermore, disengagement was rapidly rescued when each drug was washed out in CDC20-depleted cells. Based on the results, I propose that centrioles in M-phase-arrested cells are prematurely disengaged following two sequential steps: (i) removal of SAS6 from engaged centrioles with a help of PLK1 and (ii) a microtubule-mediated pushing force.