Role of the kinetochore protein Ndc10 in mitotic checkpoint activation in Saccharomyces cerevisiae

Mitotic checkpoints delay cell cycle progression in response to alterations in the mitotic apparatus, thus ensuring correct chromosome segregation. Wh ile improper spindle orientation activates the Bub2/Bfa1-dependent checkpoi nt in budding yeast, delaying exit from mitosis, lack of bipolar kinetochor e-microtubule attachment activates a signal transduction cascade that preve nts both anaphase onset and exit from mitosis by inhibiting the Cdc20/APC ( Anaphase Promoting Complex)-mediated proteolysis of securin and inactivatio n of mitotic cyclin-dependent kinases (CDKs), respectively. Proteolysis of the securin Pds1 is necessary to liberate the separase Esp1, which then tri ggers sister chromatid separation, whereas inactivation of mitotic CDKs is a prerequisite for exit from mitosis and for starting a new round of DNA re plication in the next cell cycle. In budding yeast, this latter checkpoint response involves the proteins Mad 1 2, 3, Bub1 and Bub3, whose vertebrate counterparts localize to unattached kinetochores. Mutations that alter othe r kinetochore proteins result in mitotic checkpoint activation, while the n dc10-I mutation not only impairs kinetochore function, but also disrupts th e checkpoint response, indicating a role for Ndc10 in this process. Here we present evidence that Ndc10 is not part of the Bub2/Bfa1-dependent pathway , and its role in the checkpoint response might also be different from that of the other Mad and Bub proteins. Indeed, Ndc10. unlike other mitotic che ckpoint proteins, is not required for the mitotic block induced by overexpr ession of the Mps1 protein kinase, which is implicated in mitotic checkpoin t control. Furthermore, the delay in mitotic exit caused by non-degradable Pds1, which does not require Mad and Bub proteins, depends on Ndc10 functio n. We propose that a pathway involving Ndc10 might monitor defects in the m itotic apparatus independently of the Mad and Bub proteins. Since the Esp1 separase is required for exit from mitosis in both ndc10-I and nocodazole-t reated mad2 Delta cells, the two signal transduction cascades might ultimat ely converge on the inactivation of Esp1.