Jonne Raaijmakers, Mar Soto, Vincent Blomen, Thijn Brummelkamp, Rita Maia, Andre Koch, René H. Medema
Netherlands Cancer Institute, Amsterdam, The Netherlands
The mitotic checkpoint controls faithful chromosome segregation, and inhibition of the mitotic checkpoint kinase Mps1/TTK causes excessive chromosomal instability and tumor cell death. Inhibition of Mps1 is more detrimental in tumor cells, as compared to non-transformed cells. Nonetheless, full inhibition of Mps1 is lethal in all cell lines tested thus far. This is not so for the other mitotic checkpoint kinases, Bub1 and BubR1, for which we were able to establish knock-out cell lines using Crispr/Cas9. We have used the Bub1 and BubR1 knock-out cells to study the exact role of these mitotic checkpoint kinases. We find that Bub1 is not essential for mitotic checkpoint function, but contributes to the recruitment of several other kinetochore components to facilitate chromosome alignment. Intriguingly, its kinase activity does not appear to be important for this latter function. Conversely, BubR1 is essential for mitotic checkpoint function, but does not seem to be required for chromosome alignment.
We have shown that inhibitors of Mps1 act synergistic with spindle poisons, indicating that they could be of clinical use. We have therefore studied resistance to Mps1-inhibitors and identified several mutant forms of Mps1 that render the kinase resistant to distinct small molecule inhibitors. We have constructed a transgenic mouse strain carrying such a mutation and are currently using these mice to study the effect of induced chromosome instability on tumorigenesis and aging in mice. In parallel, we are performing live cell imaging combined with single cell sequencing to follow the fate of daughter cells that inherit the wrong complement of chromosomes. Contrary to the accepted model, we find that aneuploid progeny can continue to divide in the presence of functional p53. Also, we find that a missegregated chromosome is less likely to be propagated through subsequent generations, allowing cells to regain a normal karyotype.
