Molecular characterization of the mitotic spindle checkpoint protein Bub1
Final Report Abstract
The kinetochore-bound protein kinase Bub1 performs two crucial functions during mitosis: it is essential for spindle checkpoint signaling and for correct kinetochore-microtubule attachment. Interestingly, Bub1 mutations are found in cancer tissues and cancer cell lines. Using an isogenic RNAi complementation system in HeLa cells I investigated the effect of structural Bub1 mutants on chromosome segregation. I demonstrate that Bub1 can regulate chromosome segregation in a kinetochore-independent manner, albeit at lower efficiency. I also identified a novel conserved motif within Bub1 (AA 458-476) that is essential for spindle checkpoint signaling via the recruitment of the checkpoint proteins Mad1 and Mad2, but does not regulate chromosome-microtubule attachment. I showed that Bub1 kinase activity is crucial for kinetochore-microtubule attachment and plays a minor role in spindle checkpoint signaling. Finally, I showed that several cancer-related Bub1 mutants weaken but do not abrogate the spindle checkpoint, suggesting that checkpoint weakening rather than the loss of checkpoint signaling is implicated in tumorigenesis.
